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 Handbook of
Plant Growth
pH as the Master Variable

edited by
Zdenko Rengel
University of Western Australia
Perth, Western Australia, Australia

M A R C E L

MARCEL DEKKER, INC. NEW YORK • BASEL

D E K K E R

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 ISBN: 0-8247-0761-3

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Neither this book nor any part may be reproduced or transmitted in any form or by any
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Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 Preface

Few would argue that pH is truly a master variable that permeates just about any
area of study of living organisms and extends into the physical and chemical
world in which living organisms come about, grow, develop, reproduce, and die.
There is arguably no other master variable that captures the complexities of the
interactions between the biological, physical, and chemical aspects of the world
to a similar extent.
This book aims to provide a unifying view of the role of pH in plant growth,
taking into account molecular, biochemical, functional, structural, and develop-
mental factors in such growth, as well as environmental processes involved in
plant interaction with the biotic and abiotic environment. The book tries to cap-
ture the multitude of roles played by H + ions in the processes that sustain life
on this planet. It deals with pH in plant symplasm, plant apoplasm, the rhizo-
sphere, the ecosystem, and soil biotic and abiotic components, thus covering plant
life from the general environment all the way down to cell organelles and mole-
cules. The book covers four main subjects: (1) dynamics of H + fluxes across
membranes (plasma membrane, tonoplast, chloroplast thylakoids and mitochon-
dria), (2) the role of H + activity (pH) in cellular, subcellular, and whole plant
processes, (3) the role of pH and H + fluxes in soil biotic processes involving
microorganisms as well as in soil-plant-microbe interactions, and (4) the interde-
pendence of pH changes and soil abiotic processes (ion availability). The book
covers a wide range of topics spanning many scientific disciplines, for example,
plant biology, cell physiology, botany, microbiology, ecology, soil science,
agronomy, and forestry.
All chapters have been reviewed according to the standards of high-impact

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 international journals. I would like to thank the authors, who patiently went with
me through a number of revisions of their chapters. I would also like to thank
the Marcel Dekker, Inc., staff for capable handling of numerous issues and for
their dedication to producing a high-quality multidisciplinary book.

Zdenko (Zed) Rengel

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 Contents

Preface
Contributors

1. H+-ATPases in the Plasma Membrane: Physiology

and Molecular Biology
Thomas Jahn and Michael Gjedde Palmgren

2. H+-ATPase and H+-PPase in the Vacuolar Membrane:

Physiology and Molecular Biology
Masayoshi Maeshima and Yoichi Nakanishi

3. The Cytoplasmic pH Stat

Robert J. Reid and F. Andrew Smith

4. Confocal pH Topography in Plant Cells: Shifts of Proton

Distribution Involved in Plant Signaling
Werner Roos
5. pH as a Signal and Regulator of Membrane Transport
Hubert H. Felle

6. The Role of the Apoplastic pH in Cell Wall Extension

and Cell Enlargement
Robert E. Cleland

1. Mechanisms and Physiological Roles of Proton Movements

in Plant Thylakoid Membranes
W. S. Chow and Alexander B. Hope

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 8. Dynamics of H Fluxes in Mitochondria! Membrane
Francis E. Sluse and Wiestawa Jarmuszkiewicz
9. H 4 Fluxes in Nitrogen Assimilation by Plants
Fernando Gallardo and Francisco M. Cdnovas
10. Crassulacean Acid Metabolism: A Special Case of pH
Regulation and H 4 Fluxes
Karl-Josef Dietz and Dortje Golldack
11. Dynamics of H + Fluxes in the Plant Apoplast
Joska Gerendds and Burkhard Sattelmacher
12. H" Currents around Plant Roots
Miguel A. Pineros and Leon V. Kochian
13. Role of pH in Availability of Ions in Soil
Zclenko Rengel
14. Regulation of Microbial Processes by Soil pH
David E. Crowley and Samuel A. Alvey
15. The Role of Acid pH in Symbiosis between Plants
and Soil Organisms
Karen G. Ballen and Peter H. Graham
16. Distribution of Plant Species in Relation to pH of Soil
and Water
Jacqueline Baar and Jan G. M. Roelofs

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 Contributors

Samuel A. Alvey Department of Environmental Sciences, University of Cali-

fornia, Riverside, Riverside, California
Jacqueline Baar* Department of Aquatic Ecology and Environmental Biol-
ogy, University of Nijmegen, Nijmegen, The Netherlands
Karen G. Ballen Biology Department, Augsburg College, Minneapolis, Min-
nesota
Francisco M. Canovas Department of Molecular Biology and Biochemistry,
Andalusian Institute of Biotechnology, University of Malaga, Malaga, Spain
W. S. Chow Research School of Biological Sciences, Australian National Uni-
versity, Canberra, Australia
Robert E. Cleland Department of Botany, University of Washington, Seattle,
Washington
David E. Crowley Department of Environmental Sciences, University of Cali-
fornia, Riverside, Riverside, California
Karl-Josef Dietz Department of Physiology and Biochemistry of Plants, Uni-
versity of Bielefeld, Bielefeld, Germany
Hubert H. Felle Botanisches Institut I, Justus Liebig University, Giessen, Ger-
many

* Current affiliation: Department of Applied Plant Research, Wageningen University and Research
Center, Horst, The Netherlands

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 Fernando Gallardo Department of Molecular Biology and Biochemistry, An-
dalusian Institute of Biotechnology, University of Malaga, Malaga, Spain
Joska Gerendas Institute for Plant Nutrition and Soil Science, University of
Kiel, Kiel, Germany
Dortje Golldack Department of Physiology and Biochemistry of Plants, Uni-
versity of Bielefeld, Bielefeld, Germany
Peter H. Graham Department of Soil, Water, and Climate, University of Min-
nesota, St. Paul, Minnesota
Alexander B. Hope School of Biological Sciences, Flinders University, Ade-
laide, South Australia, Australia
Thomas Jahn Department of Agricultural Sciences, The Royal Veterinary and
Agricultural University, Copenhagen, Denmark
Wiestawa Jarmuszkiewicz Department of Bioenergetics, Adam Mickiewicz
University, Poznan, Poland
Leon V. Kochian U.S. Plant, Soil and Nutritional Laboratory, USDA-ARS,
Cornell University, Ithaca, New York
Masayoshi Maeshima Graduate School of Bioagricultural Sciences, Nagoya
University, Nagoya, Japan
Yoichi Nakanishi Graduate School of Bioagricultural Sciences, Nagoya Uni-
versity, Nagoya, Japan
Michael Gjedde Palmgren Department of Agricultural Sciences, The Royal
Veterinary and Agricultural University, Copenhagen, Denmark
Miguel A. Pineros U.S. Plant, Soil and Nutrition Laboratory, USDA-ARS,
Cornell University, Ithaca, New York
Robert J. Reid Department of Environmental Biology, Adelaide University,
Adelaide, South Australia, Australia
Zdenko Rengel Department of Soil Science and Plant Nutrition, The Univer-
sity of Western Australia, Perth, Western Australia, Australia
Jan G. M. Roelofs Department of Aquatic Ecology and Environmental Biol-
ogy, University of Nijmegen, Nijmegen, The Netherlands
Werner Roos Institute of Pharmaceutical Biology, Martin Luther University,
Halle (Saale), Germany
Burkhard Sattelmacher Institute for Plant Nutrition and Soil Science, Univer-
sity of Kiel, Kiel, Germany

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 Francis E. Sluse Laboratory of Bioenergetics, University of Liege, Liege, Bel-
gium
F. Andrew Smith Department of Soil and Water, Adelaide University, Ade-
laide, South Australia, Australia

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 1
H+-ATPases in the Plasma Membrane:
Physiology and Molecular Biology

Thomas Jahn and Michael Gjedde Palmgren

The Royal Veterinary and Agricultural University, Copenhagen,
Denmark

1 INTRODUCTION
A typical plant cell expresses four ATP-fueled proton pumps (H+-ATPases), each
one targeted to a specific cellular membrane. The F0F, and CF0CF, H+-ATPases,
present in the mitochondrial inner membrane and the thylakoid membrane, re-
spectively, operate under physiological conditions to synthesize ATP at the ex-
pense of H + gradients. Vacuolar H+-ATPase and plasma membrane H+-ATPase,
on the other hand, generate H + gradients at the expense of ATP. The plant plasma
membrane H+-ATPase has been extensively discussed in a number of recent re-
views [1-5]. In the present chapter we give an overview of recent structural and
functional aspects of the plasma membrane H+-ATPase, including its regulation
and its role in intra- and extracellular pH regulation.

2 EVOLUTION OF PROTON PUMPS

Plasma membrane H+-ATPase is not evolutionarily related to any other plant
proton pumps. This enzyme is composed of a single polypeptide of around 100

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 kDa (around 950 amino acid residues) and belongs to the superfamily of P-type
ATPases, mainly cation pumps characterized by forming a phosphorylated reac-
tion cycle intermediate and by being inhibited by vanadate [6,7].
Plasma membrane H -ATPases are ubiquitous in higher plants, algae, and
fungi but have not been identified in animals and eubacteria. This would suggest
a relatively late evolutionary origin of this class of proton pumps. A sequence
related to plasma membrane H 1 -ATPase has been identified in the genome of the
archaebacterium Methanococcus jannaschii [8]. In the four other archaebacterial
genomes available so far, no sequences with homology to H f -ATPases have been
observed. Also, a plasma membrane H + -ATPase-like gene has been cloned from
Leishmania donovani, a protozoan [9|. In present-day bacteria, P-type ATPases
are involved in pumping K f , Mg 2+ , Ca 24 , Cu 2+ , and Cd 2+ , suggesting that, early
in evolution, P-type ATPases mainly pumped divalent cations. Evolution of
mechanisms for extrusion of metal ions from cells might have been necessary
to prevent metal salts from precipitating in the cell or giving rise to toxic effects.
Other FT pumps such as F0F, and CF0CF, (both F-type) H + -ATPases are
multisubunit pumps that probably evolved early in evolution in order to extrude
excess H" generated in anaerobic metabolism [10]. Later, other mechanisms for
pH homeostasis evolved, and F0F, ATPases acquired a new role in ATP synthesis
by operating in the reverse direction.

3 MOLECULAR BIOLOGY OF THE PLASMA MEMBRANE

H+-ATPase
Every single plant species investigated has a large number of plasma membrane
H+-ATPase isoforms. Twelve different isoforms have been identified in Arabi-
dopsis (according to the P-type ATPase database—PAT-base; http://biobase.
dk/~axe/Patbase.html), and so far there is evidence for the expression of eight
of these isoenzymes. At least nine isoforms are present in the genome of the
tobacco Nicotianu plumbagin(folia [4]. The expression patterns of Arabidopsis
and tobacco plasma membrane H + -ATPase isoforms have been studied exten-
sively using reporter genes [11,12] and immunohistochemical localization of epi-
tope-tagged H -ATPase [13]. The picture that has emerged supports the view
that each isoform is expressed in a tissue- and development-specific manner [11-
13]. Furthermore, it has been established that in certain cell types more than one
isoform might be expressed at the same time in the same cell [ 1 1 ] .
It is not known why there are so many plasma membrane FT-ATPases in
a given plant. When expressed in the yeast Saccharomyces cerevisiae, different
isoforms of Arabidopsis [14] and tobacco [15] plasma membrane H+-ATPases
exhibit quantitative differences with respect to a number of kinetic parameters.
However, depending upon the isoform, heterologously expressed plant FP-ATP-
ase is phosphorylated in this host at the penultimate threonine residue, which

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 might alter the properties of the enzyme [16,17]. Therefore, it is still unclear
whether the functional differences observed between the H+-ATPases produced
in yeast are relevant in planta.
The current completion of eukaryotic genome sequencing programs has
showed that multigene families are a normal feature of multicellular organisms.
For example, more than 60% of the genes on chromosome 2 of Arabidopsis
thaliana have a significant match with another Arabidopsis gene [18]. Gene dupli-
cation events might give rise to a large number of functionally more or less
equivalent isoforms. Evolution of different promoter regions for such isoenzymes
might be the simplest means for providing tissue- and development-specific ex-
pression of a given enzyme.

3.1 Structure and Function of Plasma Membrane H+-

ATPase
How a single polypeptide of rather limited size is able to execute the whole
sequence of functions of ATP hydrolysis, energy coupling, energy transfer, as
well as highly specific binding and release of the transported substrate is not well
understood.
Hydrophobicity analyses in combination with biochemical studies have
suggested the presence of 10 transmembrane helices, with most of the protein
mass (about 70% of the H+-ATPase) facing the cytoplasmic side of the plasma
membrane [19]. The cytoplasmic part of the enzyme involves the N- and the C-
termini, a relatively small loop of about 135 residues between transmembrane
segments 2 and 3, a large loop of about 345 residues between transmembrane
domains 4 and 5, and two very small loops of less than 20 residues connecting
transmembrane helices 6-7 and 8-9. The large central cytoplasmic loop includes
a conserved DKTGT sequence in which the aspartate becomes reversibly phos-
phorylated during catalysis. In addition to phosphorylation, the large central loop
has been implicated in binding of ATP [7].
Our picture of the structure of the H+-ATPase has been improved by data
obtained by two-dimensional (2D) crystallization of the Neurospora crassa ho-
mologue (NcPMAl) [20]. This fungal H+-ATPase forms well-ordered 2D crys-
tals in which the protein appears in a hexameric structure. The polypeptide
crosses the membrane 10 times and a big cytoplasmic domain is connected with
the membrane-embedded part of the enzyme.
The transmembrane part of the H+-ATPase most likely contains the H +
binding site(s). Intensive mutagenesis analysis of various P-type ATPases has
pointed to a role for transmembrane oxygen atoms in binding and coordination
of the transported cations. In all plasma membrane H+-ATPases, one aspartate
residue is conserved in transmembrane helix 6 (D684 in Arabidopsis AHA2;
D730 in Saccharomyces cerevisiae PMA1). Thus, this group is a candidate resi-

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 due directly involved in H + coordination. In a recent study, all charged residues
in the postulated transmembrane segments of S. cerevisiae PMA1 were mutagen-
ized and the effects on proton pumping by the H+-ATPase were tested [21].
Nearly half of the mutant ATPases (including D730) were misfolded and conse-
quently not properly secreted to the plasma membrane. In this case, the role of
D730 in binding of H + was difficult to ascertain. However, mutagenesis studies
with PMA1 have demonstrated that H + pumping can take place in the absence
of D730 provided that second-site mutations are introduced to remove the posi-
tive charge of R695 in neighboring transmembrane helix 5 [22]. Therefore, D730
is apparently not part of the H + translocation mechanism but rather is important
for proper folding of the enzyme, probably by forming a salt bridge with R695.
Two additional charged residues, E703 in transmembrane segment 5 and E803
in transmembrane segment 8, appear to be important for the coupling between
ATP hydrolysis and H + pumping in PMA1 [21], but these residues are not con-
served in plant plasma membrane H+-ATPases.
The C-terminus of the plasma membrane H+-ATPase functions as an au-
toinhibitor of enzyme activity [23,24]. Successive deletions [25] and single point
mutations have revealed the presence of two inhibitory regions within the C-
terminal domain [26-29] as well as a binding site for regulatory 14-3-3 protein
(see later).

4 PHYSIOLOGY OF THE PLASMA MEMBRANE H+-ATPase

The plant plasma membrane H + -ATPase is thought to play a crucial role in a
number of essential physiological processes [1,2]. Among them are energization
of nutrient uptake, phloem loading, opening of stomata, as well as the regulation
of extra- and intracellular pH (Fig. 1).
The plasma membrane H+-ATPase is very abundant in cells that are spe-
cialized for nutrient acquisition. Here it plays a role in generation of the electro-
chemical gradient of H* that provides the driving force for uptake of solutes
through channel proteins and H + -coupled carriers. Cells specialized for nutrient
uptake are, for example, those of the root epidermis, phloem companion cells,
and the transfer cells of the xylem. The root is a specialized organ that functions
in uptake of nutrients from the soil and translocation of those nutrients to other
parts of the plant. In roots, plasma membrane H+-ATPase has been shown to be
highly abundant in the epidermis and vascular tissues [30-32]. In the corn root
epidermis and outer cortical cells, the plasma membrane H + -ATPase has been
shown to be asymmetrically localized, with very high abundance in plasma mem-
brane domains facing the root-soil interface [32]. Asymmetric localization of
plasma membrane H + -ATPase has also been shown in other cell types [33].
Phloem companion cells are rich in mitochondria and thus have the capacity
to synthesize large amounts of ATP. The ATP produced is believed to fuel mainly

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 H+

Phosphate
Ca2+

FIGURE 1 Plant plasma membrane H+-ATPase generates a membrane poten-

tial (negative on the inside of the cell) and a ApH gradient (acidic on the out-
side). This electrochemical gradient in turn can be used as an energy supply
for transport proteins. Typical values for pH and membrane potential are indi-
cated.

plasma membrane H+-ATPase, which is abundant in this cell type. In tobacco,

promoter-GUS fusion studies have shown PMA4 to be expressed in companion
cells but also in other tissues [12]. Based on epitope-tagging studies, Arabidopsis
thaliana plasma membrane H+-ATPase AHA3 appears to be specifically ex-
pressed in phloem companion cells [13]. The H + gradient established across the
plasma membrane of companion cells is believed to energize sucrose uptake
through H+-coupled sucrose transporters in the phloem [34]. Sucrose transporters
were first localized in phloem companion cells in Plantago major [35] and A.
thaliana [36] as the result of promoter-GUS fusion studies. Later, using immuno-
localization and in situ hybridization techniques, sucrose transporters were local-
ized in enucleated sieve elements of tobacco, potato, and tomato [37]. In the
latter case, sucrose transporter SUT1 messenger RNA (mRNA) and potentially
SUT1 protein is targeted through plasmodesmata from the companion cells to
the sieve elements. How energization of sucrose uptake through the plasma mem-
brane of enucleate sieve elements takes place is still an open question. One possi-
bility is that the mRNA of plasma membrane H+-ATPase is transported through
plasmodesmata from companion cells to the sieve elements, where H+-ATPase
is then synthesized. ATP would have to be transported in the same way as enucle-
ate sieve elements lack mitochondria. Alternatively, the membrane potential gen-

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 crated by the H + -ATPase in companion cells is transmitted to sieve elements,
where it is used to drive uptake through sucrose transporters located in the plasma
membrane of these cells. The possibility that plasma membrane H 4 -ATPase is
in fact present in the enucleate sieve elements but has escaped detection by the
methods employed so far cannot be discarded. For example, modification of H + -
ATPase mRNA by insertion of a tag or a reporter gene might interfere with
targeting to sieve elements.
An additional number of cell types in the plant body are specialized for
intense active transport and contain high amounts of plasma membrane HT-ATP-
ase. Most prominent are stomatal guard cells and pulvinar cells. Guard cell local-
ization of plasma membrane H ' -ATPases has been demonstrated for MHA2 in
Zea mays [38], VHA1 and VHA2 in Vicia faba [39], and PMA2 and PMA4 in
Nicotiana plumbaginifolia [12]. Activation of the H+-ATPase in guard cells re-
sults in an increased uptake of potassium that precedes water uptake [40]. Due
to the special wall anatomy of the cells, osmotic swelling of the cells results in
opening of the stomatal pore, allowing transpiration and gas exchange with the
environment of the plant. In accordance with this model, cosuppression of PMA4
gene expression in N. plumbaginifolia results in failure of stomatal opening [41].
Pulvinar cells are specialized cells functioning as osmotic motors driving leaf
movements. Increased expression of plasma membrane H+-ATPase in these cells
[42] suggests a role for this enzyme in energizing the massive ion fluxes taking
place here.

4.1 Role in Regulation of Apoplastic pH

Activation of proton pump activity in plant tissues results in an increase in acidi-
fication of external solutions, whose pH can be lowered by at least one unit con-
comitant with this hyperpolarization of the plasma membrane [43-45]. Although
accurate determination of apoplastic pH is difficult, it has been accomplished
by the use of pH-dependent fluorescent dyes [46,47] or sharp double-barreled
microelectrodes [48]. Although the cell wall is buffered and able to maintain
a steep pH gradient toward the external solution, compounds affecting plasma
membrane H + -ATPase positively such as fusicoccin or negatively such as vana-
date provoke acidification and alkalization of the apoplast, respectively. However,
the deviation from normal values (typically between pH 5.1 and 5.9) is not very
dramatic and ranges from O.I to 0.6 pH units [46-48]. A more extensive discus-
sion of factors contributing to regulation of apoplastic pH can be found in Chapter
11. Here the specific roles of plasma membrane H+-ATPase are discussed.
Acidification at the root surface is important for increasing nutrient accessi-
bility. This is due to the fact that most nutrients in the soil are not readily available
for uptake by the plant. Soil particles can be of either organic or inorganic origin,
but in both cases they are characterized by having negative charges on their sur-

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 face. The negative charge of soil particles causes cationic mineral nutrients such
as K + , Ca2+, Mg 2+ , Mn 2+ , Fe3+, and A13+ to be absorbed to their surface, whereas
anions in principle do not bind to soil particles and remain dissolved. However,
sulfate (SO 4 2 ~) is typically bound strongly to Ca2+, whereas phosphate (PO43~)
forms very strong complexes with soil particles by replacing hydroxyl ions (OH*)
that are complexed to Fe2+, Fe3+, and A13+. Solubilization of nutrients bound to
soil particles occurs by cation exchange with H + , which is very efficient in ex-
changing bound cations. Factors affecting soil pH are the decomposition of or-
ganic matter, acidic rain, and active extrusion of organic acids and H + from the
roots. Because the plasma membrane H+-ATPase is responsible for the extrusion
of H + from roots to the soil, its role in nutrient mobilization cannot be overesti-
mated.
Acidification of the apoplast is believed to play an important role in plant
physiology, namely in cell elongation growth. The plant cell wall is a complex
cross-linked network of carbohydrate polymers limiting cell expansion. There-
fore, for the cell to expand, this rigid structure has to be softened by breaking
cross-linking bonds. This process appears to be strictly pH dependent. Growth-
promoting substances such as the plant hormone auxin and the fungal toxin fusi-
coccin have promoted H + extrusion from plant tissues [49-51]. This increased
H + secretion has been ascribed to increased activity of the plasma membrane
H+-ATPase. Rayle and Cleland [52] and Hager et al. [53] independently formu-
lated the acid growth theory of auxin action that adopts elements of the growth
theory proposed by Ruge [54]. According to this model, auxin-induced growth
is triggered by acidification of the cell wall, resulting in disruption of chemical
or physical bonds in the cell wall matrix. Loosening of cell wall bonds would
then allow turgor-driven cell expansion in a well-ordered manner. Initiation of
root hairs has been shown to be accompanied by, and strictly dependent on, the
formation of local changes in apoplastic and cytoplasmic pH around the initiation
zone [55]. This strongly suggests a role of the plasma membrane H+-ATPase in
the regulation of cell expansion.
A molecular mechanism for the acid-induced softening of cell walls is be-
ginning to emerge. A number of tissues, such as hypocotyl segments, respond
with an increase in plastic extensibility when incubated in buffers with a pH of
4-4.5. When such segments are treated with boiling to denature proteins, the cell
walls refuse to expand. However, Cosgrove and coworkers [56-58] could show
that addition of extracts of native cell wall proteins to the incubation solution
restored the pH-sensitive response. This bioassay has been used to identify a class
of cell wall proteins, so-called expansins. The ability of expansins to promote cell
wall relaxation is strictly regulated by the pH of the apoplast. Expansins comprise
a big family with a high degree of functional conservation and, although their
biochemical function has not been clarified, they show sequence similarity to a
family of endoglucanases [59]. Endoglucanases, xyloglucan endotransglycosyl-

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 ase, and other enzymes also modify cell wall structure during cell elongation but
appear to act secondarily to expansin action [59].
In corn coleoptiles, the growth hormone auxin induces expression of an
isoform of plasma membrane H f -ATPase [38] concomitant with increased turn-
over of the enzyme [60], indicating that H 4 -ATPase is an element of auxin-in-
duced growth in this tissue. However, auxin-induced growth stimulation of to-
bacco leaf strips does not appear to involve plasma membrane H+-ATPase,
although cell wall loosening in some form does occur [61]. This would suggest
that cell wall loosening induced by auxin does not require H+-ATPase per se but
may involve other proteins. Thus, a connection between auxin and the expression
of cell wall-modifying proteins is beginning to emerge. The expression of tomato
genes encoding xyloglucan endotransglycosylase (LeEXTl) and an endo-1,4-
beta-glucanase (CelT) is auxin regulated in etiolated hypocotyls [62]. Similarly,
expression of expansin genes from tomato [63] and Finns taeda [64] is up-
regulated in hypocotyls during incubation with auxin.

4.2 Role in Regulation of Cytoplasmic pH

Several factors contribute to the formation of a pH-stat that keeps cytoplasmic
pH more or less constant. These factors are discussed in detail by Reid and Smith
(Chapter 3). However, the specific role of plasma membrane H+-ATPase will be
discussed here.
As the result of each catalytic cycle, the plasma membrane H + -ATPase
transfers at least one proton out of the cytosol and into the apoplast [65]. There-
fore, it would be expected that activation of proton pumping results in alkalization
of the cytoplasm. Assuming a cellular volume of 20 pL, a typical cell contains
about 10X H + at pH 7. A number of 1 million plasma membrane H + -ATPase
molecules per cell is probably not unreasonable; for example, much smaller kid-
ney cells each contain more than a million Na + /K + -ATPase molecules in the
plasma membrane [66]. Extrapolating from the maximal turnover rate of 6000
per minute for the purified enzyme (T. Jahn et al., unpublished data), 108 H"
would be pumped out of the cell per second. Obviously, this should have dramatic
implications for intracellular pH.
The pH dependence of the activity of the plasma membrane H + -ATPase
does not allow it to operate under alkaline conditions. The curve describing the
pH versus activity profile of the enzyme is bell shaped with an optimum that is
typically pH 6.5 (Fig. 2). The activity drops sharply when the pH becomes either
more alkaline or acidic than the optimal pH. This pH dependence is highly depen-
dent upon the ATP levels in the cell. Similarly, the ATP affinity of the pump is
strictly linked to pH. The closer the pump is at its pH optimum, the higher its
affinity for ATP is [25].

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 100

50

6.0 7.0
PH

FIGURE 2 The pH optimum of plasma membrane l-T-ATPase from oat roots

is slightly acidic but moves toward physiological pH as the ATPase is acti-
vated, e.g., by lysophospholipids or free fatty acids. (O) No additions; (•) 24
u^/mL lysophosphatidylcholine; (x) 6 [iM (18:3) linolenic acid; (•) 6 uM (20:
4) arachidonic acid. (Adapted from Ref. 82.)

The shape of the pH dependence profile implies that as the pH in the cell
becomes alkaline, the activity of the pump diminishes. Similarly, if the pH in
the cytosol acidifies from a typical value of pH 7.2, the pump increases in activity
severalfold, resulting in adjustment of the pH back to normal. In this way the
plasma membrane H+-ATPase by itself serves as an efficient pH-stat of the cell.
This is illustrated by the difficulty in changing cytoplasmic pH, even by treating
the plant cell with a compound known to activate H+-ATPase activity. Fusicoccin
is a powerful agent causing activation of plasma membrane H+-ATPase (see
later). An addition of 1 fiM fusicoccin to root hairs of Medicago saliva resulted
in an increase in plasma membrane potential of 30 mV, which is probably due
to activation of H+-ATPase, but the pH of the cytoplasm remained constant at
around pH 7 [67]. Other factors that might lead to a decrease in cytoplasmic pH
are low oxygen and acidic soil, both of which are factors that might result in
cytoplasmic acidosis. The plasma membrane H+-ATPase operates during anoxia
and might therefore play a role under these conditions [68]. Overexpression of
a modified form of AHA3 H+-ATPase in transgenic Arabidopsis thaliana led to

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

 increased growth at low pH compared with wild-type plants [69]. These data
suggest that proton extrusion via the plasma membrane H + -ATPase can contrib-
ute to stabilization of cytoplasmic pH.
A role for plasma membrane FT-ATPases in intracellular pH regulation is
supported by the data obtained from heterologous expression of regulatory mu-
tants of plant plasma membrane H + -ATPase in the yeast Saccharomyces cerevis-
iae. The more activated the plant H '-ATPase, the higher its capability is to confer
acid tolerance to yeast cells lacking the endogenous H + -ATPase PMAl [4,26].
In addition, plant isoforms with different pH optima when expressed in yeast
support growth at different external pH values [15].
Sudden changes in the cytoplasmic pH are often seen in plant cells attacked
by certain pathogens. One example is the rapid acidification of the cytosol of
tobacco cells [70] and Chenopodium rub mm [71] after treatment with various
fungal elicitors. The role of plasma membrane H + -ATPase under these conditions
is unclear. It is also questionable whether these changes in pH are related to
elicitor-induced gene expression |71|.

5 REGULATION OF THE PLASMA MEMBRANE H+-ATPase

Because of the fundamental roles plasma membrane H*-ATPase plays in plant
physiology, the enzyme is likely to be subject to tight regulation. Indeed, regula-
tion of the PT-ATPase occurs at different levels. Salt stress [72,73], hydroponic
growth conditions [74], and addition of exogenous glucose [75] or auxin [38]
are factors that have been found to lead to increased expression of plasma mem-
brane H + -ATPase in a number of species. Increased H 4 -ATPase activity concom-
itant with an increase in immunodetectable protein occurs in response to
wounding [76] and aging [77]. The number of fusicoccin binding sites can be
taken as a measure of the number of H ' -ATPases in complex with the activator
14-3-3 protein (see later). Auxin application to corn coleoptiles increased the
number of fusicoccin receptors [78]. Under cold stress, an increase in fusicoccin
binding activity in plasma membranes of suspension cultures of sugar beet cells
was accompanied by an increase in H 4 -ATPase activity [79].
Some plasma membrane H -ATPases with short open reading frames in
the 5'-untranslated region appear to be regulated at the translational level [4,80].
The physiological significance of this phenomenon is at present unknown.
At least fusicoccin (see later), blue light [81], lysophosphatidylcholine and
free fatty acids [82,83], and products of phospholipase A2 action [84,85] activate
proton pumping and ATPase activity by the plasma membrane H+-ATPase via
a mechanism not involving increased gene expression. Considerable attention has
been paid to the study of posttranslational regulation of H ' -ATPase activity; this
work is summarized next.

Copyright 2002 by Marcel Dekker, Inc. All Rights Reserved.

Exploring the Variety of Random
Documents with Different Content
 12. In compounds of self : e.g., self-esteem, self-respecting, etc.,
but not in selfhood, selfish, selfsame, or oneself.
13. In connecting ex, vice, general, elect, etc., constituting parts
of titles, with the chief noun: e.g., ex-Governor Draper, Governor-
elect Wilson, etc.
14. In compounds of by : e.g., by-laws, by-products, etc.
15. In connection with prefixes co, pre, and re when followed by
words beginning with the same vowel as that in which they
terminate, but not when followed by a {48} different vowel or a
consonant: e.g., co-operation, but coeducation ; pre-empted, but
prearranged ; re-elected, but recast. Exceptions: combinations with
proper names, long or unusual formations, and words where the
hyphen defines the meaning: e.g., re-creation and recreation, re-
form and reform, re-collect and recollect, pre-Raphaelite.
16. In writing ordinal numbers when compounded with such
words as first-rate, second-hand, etc.
17. In connection with the word quasi prefixed to a noun or to an
adjective: e.g., quasi-corporation, quasi-compliant, etc.
18. In connection with the Latin prepositions extra, infra, semi,
supra, and ultra : e.g., extra-hazardous but extraordinary ; ultra-
conservative but Ultramontane.
19. In spelling out fractional numbers involving more than two
words: e.g., The supply is three-quarters exhausted ; but, This
leaves twenty-five hundredths.
20. In compounding numerals of one syllable with self-explanatory
words of various meanings: e.g., three-legged, four-footed, one-
armed, etc.
Also in combining numerals with nouns, to form an adjective: e.g.
twelve-inch rule, hundred-yard dash, two-horse team, etc.
21. In compounding a noun in the possessive case with another
noun: e.g., jews’-harp, crow’s-nest, etc. {49}
 22. In some compounds with tree : e.g., apple-tree ; but
whippletree, crosstree, etc.
23. In compounding personal epithets: e.g., hard-headed, bow-
legged, etc.
24. Use the hyphen in the following words:
after-years bas-relief birth-rate blood-feud blood-
relations common-sense cross-examine cross-reference
cross-section death-rate feast-day folk-song food-stuff
fountain-head guinea-pig horse-power page-proof pay-
roll poor-law post-office sea-level sense-perception
son-in-law subject-matter man-of-war object-lesson
thought-process title-page wave-length well-being well-
nigh will-power

Do not use the hyphen:

1. When writing points of the compass: e.g., northeast,
southwest ; but north-northeast, etc.
2. In words ending in like, unless compounded with nouns
containing more than one syllable (except when ending in l ): e.g.,
childlike, lifelike ; but business-like, bell-like, etc.
3. In compounds ending with man or woman : e.g., workman,
needlewoman, etc.
4. In phrases such as by and by, by the bye, good morning,
attorney at law, coat of arms, etc.
5. In words ending in boat, house, look, room, side, yard, shop,
mill, field, work, chair, maker, holder, keeper, skin, store, book,
fold, score, penny, pence when the prefixed noun {50} contains but
a single syllable: e.g., twofold, schoolroom, handbook, windmill,
bookkeeper, workshop, etc. Exceptions: Court-house and State
House.
When prefixed noun contains two syllables, use the hyphen: e.g.,
bucket-shop, twenty-fold, ante-room, mason-work, paper-mill, etc.9
 When prefixed noun contains three or more syllables write as two
separate words: e.g., policy shop.
6. In compounds of bi, tri, semi, or demi : e.g., bimonthly,
tricolor, semiannual, demijohn. Exceptions: long or unusual
formations: e.g., bi-centennial, etc.
7. In today, 10 tonight, tomorrow, viewpoint, or standpoint.
9 See page 47, Sec. 5.
10 Cf. the old English words to-morn, to-year, etc.

8. In connection with the negative prefixes un, in, and a : e.g.,

unrepublican, inanimate, etc., but the prefix non requires the
hyphen except in words which have become common: e.g., non-
conductor, but nonsense.
9. In connection with the words over and under employed as
prefixes: e.g., overestimate, undersecretary ; but over-soul, over-
spiritual, under-man.
10. In connection with the Latin prepositions ante, anti, inter,
intra, post, sub, and super : e.g., antedate, antidote, subtitle, etc.;
but ante-room, anti-imperialistic, intra-atomic. {51}
11. In spelling out a series of fractions: e.g., I can secure you one,
two, or three fifths, not one-, two-, or three-fifths.
Nor in writing ordinary fractions: e.g., one half, five eighths, etc.
But another class of fractions require the hyphen: e.g., twenty-
fifth, twenty-two one-hundredths.
12. In compounds ending with holder or monger : e.g.,
stockholder, ironmonger.
13. In compounds beginning with eye : e.g., eyeglass, eyebrow,
eyelash, eyewitness, etc.
14. In compounds with the word school : e.g., schoolmates,
schoolmaster, etc. Exceptions: when made with a participle: e.g.,
school-teaching ; or in combinations where separate words are more
clear: e.g., school committee, school children, etc.
 15. In compounds with deutero, electro, pseudo, sulpho, thermo,
etc., unless the compound is unusual: e.g., electrotype, pseudonym,
etc.
16. In compound adverbs: e.g., meantime, moreover, forever,
everywhere, etc. But in phrases like after a while, in the mean time,
for ever and ever, the words should be separated.
17. In words like anybody, anyhow, anything, anyway, anywhere,
somebody, somehow, something, sometime, somewhat,
somewhere. But any one and some one are written as separate
words.
 {52}

DIVISION OF WORDS

THE unnecessary division of a word should always be avoided. The

whole word should be carried over whenever possible.
2. A word should be divided in accordance with its natural division
in correct pronunciation, rather than according to derivation: e.g.,
knowl-edge, not know-ledge ; but divide according to meaning and
derivation as far as is compatible with good spacing and
pronunciation: e.g., dis-pleasure, not displeas-ure.
3. A single consonant between two vowels should be joined with
the first vowel if short, but with the latter if long: e.g., riv-er, ri-val ;
but avoid dividing words as short as these if possible.
4. When two consonants come together between two vowels the
consonants should be divided: e.g., mil-lion, struc-ture.
5. When three consonants come together between two vowels the
first of which is short, all which can be pronounced together go with
the last syllable: e.g., han-dle, chil-dren, frus-trate, etc.
6. Whenever practicable, and always when the pronunciation of a
word is peculiar, the division should come upon the vowel: e.g., pro-
duct, colo-nel, sepa-rate, peo-ple, pro-gress. {53}
Exceptions: words ending in -able and -ible, which should carry
the vowel over into the next line.
7. The letters c and g must never be separated from the vowels
e, i, and y upon which their soft sound depends: e.g., re-li-gion, ca-
pa-ci-ty, etc.
 8. X must never begin a syllable, j must never end one, and q
must not be separated from u, which invariably follows it.
9. When used as terminations, ing, en, ed, er, est, and the plural
es are considered as syllables, except when the preceding
consonant is doubled, or when they follow c or g soft: e.g., speak-
ing, tak-en, long-er ; but lat-ter, for-cing, ran-ging.
10. Two-letter divisions should always be avoided. These words
should never be divided: eleven, heaven, power, faster, finer,
houses, given, flower, prayer, soften, liken, verses, listen, often,
voyage, nothing, even, etc.
11. When a derivative word ends in t, the t is carried over when
the accent changes: e.g., instinc-tive (instinct); but not otherwise:
e.g., construct-ive (construct).
12. More than two divisions in successive lines should be avoided.
13. A division at the end of the last full line of a paragraph should
be avoided.
14. A word of four letters is not divisible. Dividing words of five or
six letters should be avoided if possible. {54}
15. An amount that is stated in figures should not be divided.
16. A word in the past tense, pronounced as one syllable, should
not be divided: e.g., beamed.
17. The separation of two initials of a person’s name, or such
combination as B.C., A.M., should be avoided.
18. A divided word that will appear on the last line of one page
and the top line of the next page is to be avoided.
19. Separating a divisional mark (a ) or (b ) from the matter to
which it pertains should be avoided.
20. Adjectives ending in -ical should be divided upon the i : e.g.,
musi-cal, not music-al or mu-sical.
21. Word endings -tion, -sion, -tial, -cial, -tive, etc., are treated
as one syllable.
22. In compound words additional hyphens should be avoided:
e.g., music-lesson, not music-les-son.
 Keep prefixes, roots, suffixes, etc., as distinct as possible.
N OTE. Division of words in French, Italian, German, and Spanish, may be found
in De Vinne’s “Correct Composition,” pp. 434–446.
 {55}

INDENTION AND PARAGRAPHING

IN prose manuscript, each paragraph should be written with regular

indention.
2. When paragraphs are numbered, the figure should be written
where the capital letter of the first word would otherwise be placed.
3. In poetry, lines which overrun should be written with reverse
indention.
4. When poetry is quoted in a prose composition, it should begin
on a new line. If the continuation of the prose writing does not call
for a new paragraph, the next line of prose text begins without
indention.
5. When a prose quotation is introduced, it should be treated as in
Rule 4 if it contains more than a single sentence; otherwise it is “run
in,” with the proper quotation-marks and punctuation.
6. There are different forms of indention in typography which are
used for different purposes, viz.:
Irregular indention.— For this form
of in­den­tion there are no spec­i­fied rules,
and it is used prin­ci­pal­ly in cer­tain styles
of po­et­ry and in dis­play work. {56}
En échelon indention is largely
used for dis­play in post­ers and ad­ver­‐
tise­ments. It con­sists in the di­ag­o­nal ar­‐
range­ment of words, thus:
 CARPETS
CHAIRS
TABLES
STOVES
Hanging indention, which makes the
first line of full width and in­dents all
the fol­low­ing lines one or more ems
on the left, as shown in these four
lines.

Lozenge in­den­tion re­quires an ar­‐

range­ment of the lines in this manner:
 {57}

PARAGRAPHING
Indention and Display are the methods employed in typography to
secure clearness and to add distinction to the printed page, but
clearness in the body of the text is dependent upon the skill of the
writer in properly dividing his composition into paragraphs. Just as
correct punctuation assists the reader in his understanding of a
sentence, so does correct paragraphing add to the understanding of
the composition as a whole. The following rules may be formulated,
based upon the practice of the most careful writers:
1. A sentence which continues the topic of the sentence which
precedes it rather than introduces a new topic should never begin a
paragraph.
2. Each paragraph should possess a single central topic, to which
all the statements in the paragraph should relate. The introduction
of a single statement not so related to the central topic violates the
unity.
3. A sentence or short passage may be detached from the
paragraph to which it properly belongs if the writer wishes
particularly to emphasize it.
4. For ease in reading, a passage which exceeds three hundred
words in length may be broken into two paragraphs, even though no
new topic has been developed.
5. Any digression from the central topic, or any change in the
viewpoint in {58} considering the central topic, demands a new
paragraph.
6. Coherence in a paragraph requires a natural and logical order of
development.
 7. Smoothness of diction in a paragraph calls for the intelligent use
of proper connective words between closely related sentences. A
common fault, however, is the incorrect use of such words as and or
but between sentences which are not closely related.
8. In developing the paragraph, emphasis is secured by a careful
consideration of the relative values of the ideas expressed, giving to
each idea space proportionate to its importance to the whole. This
secures the proper climax.
9. The paragraph, like the composition itself, should possess
clearness, unity, coherence, and emphasis. It is a group of related
sentences, developing a central topic. Its length depends upon the
length of the composition and upon the number of topics to be
discussed.
 {59}

SPACING

EACH line should be spaced evenly throughout.

2. The spaces in a line should never vary more than the difference
between a three to em space and an en quad.11
11 See page 60.

3. Uniformity in appearance shows excellence in printing. To have

one line thin-spaced and the next wide-spaced is in bad taste. Even
in narrow measure this inequality can be avoided with proper care.
4. In fonts of type where ends of hyphens and dashes touch the
adjoining letters, hair-spaces should be used. Also use hair-spaces
before colons, semicolons, interrogation-points, exclamation-marks,
and inside quotation-marks.
5. No space should be left between superior letters, or letters
indicating powers, and inferior figures or letters; or between letters
forming products.
6. No space should be left between the abbreviations A.D. and B.C.,
A.M. and P.M., between titles, such as LL.D., Ph.D., etc., or between
the abbreviations of States, such {60} as N.Y., R.I., N.H., etc., except
in a wide-spaced line.
7. Scripture references should be spaced as follows: II Sam.
1 : 2–6; 2 : 8–12.
8. No space should be left between the symbols $ and £ and the
succeeding figures.
 9. An indention of one or more ems should be allowed at the end
of the last line of a paragraph.
10. The same space should be left on each side of short words,
such as a, an, etc.
11. It is poor typography to thin-space or wide-space a line in
order to avoid a turn-over.
12. Beginning and ending a line with the same word in wide
measure should be avoided, but not at the expense of good spacing.
13. The kerned letters f and j require a hair-space before them
when they begin a line. When kerned letters end a line a hair-space
is necessary after them to prevent breaking off in printing.
14. Should wide spacing be necessary, it should occur where it will
be least noticed; viz., between words ending or beginning with tall
letters.
15. This is an en dash: –
This is an em dash: —
This is a 2 em dash: ——
This is a 3 em dash: ———
{61}
16. This line is hair-spaced.
This line is spaced with 5-em spaces.
This line is spaced with 4-em spaces.
This line is spaced with 3-em spaces.
This line is spaced with en quads.
This line is spaced with em quads.
 {62}

ITALIC

THE first italic types were designed and used by Aldus Manutius,
the celebrated Italian printer, of the fifteenth century. It is said that
the style was suggested by the handwriting of Petrarch. In the first
fonts only the lower case letters were italic, the capitals being
roman; but this irregularity soon gave way to the more regular style
of italic capitals and lower case.
1. Italic is not used for the text of a book, but it is permissible for
the preface, extracts, etc.
2. For poetry and other matter of a literary character italic has a
decorative effect.
3. Italic is used for emphasis in roman composition and vice versa.
4. Italic is used frequently for subheads, running-heads, and side-
heads, as well as for important paragraphs or extracts.
5. Italic is often used instead of roman-quoted for the titles of
books, magazines, newspapers, and names of ships.
6. Signatures or credits are often placed in italic at the end of an
article.
7. Foreign words and phrases are often required to be set in italic,
but there are many which are now so familiar to English readers {63}
that they are kept in the ordinary text. The following is a list of the
most familiar words:
à propos ad valorem addenda aide-de-camp alias
alibi alma mater anno domini ante-bellum beau idéal
billet-doux bon-ton bona fide bravo café canto carte
 blanche chapeau chaperon chargé d’affaires
chiaroscuro cicerone contra corrigenda data débris
début depot diarrhœa diatum dilettante dramatis
personæ ennui entrepôt erratum et cetera facsimile
fête finis gratis hoi polloi imprimatur innuendo
literati mandamus manœuvre mignonette naïve
ollapodrida onus paterfamilias patois per annum per
capita per cent. per centum per se post-mortem pro
rata protégé quondam régime rendezvous rôle
savant seraglio sobriquet ultimatum verbatim vice
vice versa viva voce
8. The following expressions, which are not as a rule so well
known, should be italicized:
ab ovo ancien régime bête noire comme il faut de quoi
vivre de trop en passant fait accompli grand monde
hors de combat inter alia jeu d’esprit locum tenens mise
en scène noblesse oblige raison d’être sans cérémonie
tour de force
{64}
9. Italic is used in the following words, phrases, and abbreviations
employed in literary and legal references: e.g., ibid, idem, loc. cit.,
op. cit., ad loc., s.v., supra, infra, passim, vide, circa (ca. ).
Exceptions: cf., i.e., e.g., v. (versus), viz., etc., which are always
roman.
10. Punctuation marks which are placed after italicized words
should be italic.
Italic is used:
1. For the names of plaintiff and defendant in the citation of legal
causes.
2. In algebraic, geometric, and similar matter to designate
unknown quantities, lines, etc.
 3. For s. and d. (shilling and pence ) following the figures: e.g.,
1s. 6d.
4. For specific names in Botany, Zoölogy, and Geology.
5. In medical matter roman is used instead of italic for scientific
terms.
6. For names of stars or constellations in astronomical matter.
7. In resolutions for the word Resolved.
In MS. one line drawn beneath a word or sentence signifies that it
is to be put in italic.
 {65}

ABBREVIATIONS

DATES

DATES are not usually abbreviated in writing or in regular text

matter; but when necessary the following rules may be followed:
1. Do not use st, d, rd, or th after a date given in figures; e.g.,
June 3, not June 3d or 3rd. If a date is spelled out, the rule is as
follows: e.g., June the third, not June three.
2. Do not use ult., inst., or prox., but always name the month:
e.g., Oct. 25, not 25th ult., Nov. 10, not 10th inst.
3. Use generally accepted abbreviations for the names of months
when the day of the month is also given, and prefer dates in order
of day, month, and year: e.g., 7 Jan., 1912. When the name of the
month is used alone or followed only by the year, do not abbreviate:
e.g., February, 1912, not Feb., 1912.
4. For ordinary purposes use these as the generally accepted
abbreviations for the months and days of the week:
Jan. Feb. Mar. Apr. May June July Aug. Sept.
Oct. Nov. Dec.
Sun. Mon. Tues. Wed. Thurs. Fri. Sat.
5. In tables and wherever matter must be greatly condensed use
the Dewey dates, {66} which are the briefest possible without
ambiguity, in the order of (a ) day of week, (b ) day of month, (c )
month, (d ) year. These abbreviations are not desirable except in
tabular matter.
Ja. F. Mr. Ap. My. Je. Ju. Ag. S. O. N. D.
Sn. M. Tu. W. Th. F. St.

PROPER NAMES
1. Abbreviate company in firm names: e.g., John Brown & Co.
2. Spell out names of companies, railroads, etc., using the
ampersand (&) only between proper names: e.g., Brown & Sharpe
Manufacturing Company ; Norfolk & Western Railroad ; but American
Smelting and Refining Company. If names of railroads are
abbreviated, use no space between the letters: e.g., N.Y., N.H. &
H.R.R.
3. Abbreviate United States when immediately associated with the
name of an officer of the army or navy, as Lieut. John Doe, U.S.A. ;
when it is the name of an organization of the army or navy, as First
Regiment U.S.V. ; when preceding the name of a government vessel,
as U.S.S. Brooklyn.
4. In referring to plays, specify act, scene, and line, also part if
necessary: e.g., 2 Henry IV, I, ii, 1–7.
5. Christian names should be spelled in full, as John, George,
Charles, except in an {67} original signature, or when following copy
in a quotation.
6. When necessary to abbreviate Christian names, use the forms
Dan., Edw., Sam., Thos., etc. Alex, Ben, Ed, and Sam are not
always abbreviations, and copy should be followed as regards
period. Use the following list:
Alex. Alexander
And. Andrew
Anth. Anthony
Ap. Appius
Arch. Archibald
Aug. August, Augustus
Benj. Benjamin
C. Caesar
Caes. Aug. Caesar Augustus
Cath. Catherine
Chas. Charles
Dan. Daniel
Eben. Ebenezer
Edm. Edmund
Edw. Edward
Eliz. Elizabeth
Esd. Esdras
Esth. Esther
Ez. Ezra
Ezek. Ezekiel
Ferd. Ferdinand
Fran. Francis
Fred. Frederic, Frederick
Geo. George
Herbt. Herbert
Hos. Hosea
Jas. James
Jona. Jonathan
Jos. Joseph
Josh. Joshua
Matt. Matthew
Nath. Nathaniel
Pet. Peter
Phil. Philip, Philander
Philem. Philemon
Reg. Reginald
 Richd. Richard
Robt. Robert
Sam. Samuel
Theo. Theodore
Thos. Thomas
Tim. Timothy
Wm. William

TITLES
1. In ordinary body matter use generally accepted abbreviations of
titles when they are immediately prefixed to names.
2. Do not abbreviate a title used as part of a name: e.g., Bishop
Lawrence, not Bp. Lawrence.
3. Such titles as Mr., Mrs., Messrs., Gen., Dr., Hon., Rev., when
prefixed to names, may be abbreviated; but Colonel, Major,
Professor, President, ex-President, etc., are better spelled in full.
Compound titles, such as Major-General, Lieutenant-Colonel, Rear-
Admiral, etc., should also be spelled and both {69} words capitalized.
Where a person has been mentioned by name and title, and is
afterward mentioned by title only, the title should be capitalized.
Where initials of a name are used, abbreviate the title: e.g., Col. T.
G. Benson, of the Second Illinois Regiment, has returned from
Havana. The Colonel’s friends gave him a warm reception.
4. When the names of sovereigns of a country are mentioned only
occasionally, such names may be given in full: e.g., George the Fifth,
Charles the First. When such names occur frequently, they may be
printed with roman numerals without a period: e.g., George V,
Charles I
5. The following is a full list of the usual abbreviations:
A.B. or B.A. (Artium Baccalaureus ) Bachelor of Arts
Abp. Archbishop
A.C. Archchancellor
A.D. Archduke
A.D.C. Aide-de-camp
Adjt. Adjutant
Adm. Admiral
Admr. Administrator
Admx., Admrx. Administratrix
Adv. Advocate
Agt. Agent
Aldm. Alderman
A.M. or M.A. (Artium Magister ) Master of Arts
Amb. Ambassador
A.P.A. American Protective Association
Asst. Assistant
A.T. Archtreasurer
Atty. Attorney
B.A. or A.B. Bachelor of Arts
Bart. Baronet
B.C.L. Bachelor of Civil Law
B.D. (Baccalaureus Divinitatis ) Bachelor of Divinity
B.LL. (Baccalaureus Legum ) Bachelor of Laws
B.M. (Baccalaureus Medicinae ) Bachelor of Medicine
Bp. Bishop
B.R. (Banco Regis or Reginae ) the King’s or
Queen’s Bench
Brig.-Gen. Brigadier-General
Bro(s). Brother(s)
B.S. Bachelor of Science or Bachelor of Surgery
B.V. (Beata Virgo ) Blessed Virgin
Cantab. (Cantabrigia ) Cambridge
Capt. Captain
Capt.-Gen. Captain-General
Cash. Cashier
C.B. Companion of the Bath
C.C.P. Court of Common Pleas
C.E. Civil Engineer
C.J. Chief Justice
C.M.G. Companion of the Order of St. Michael and St.
George
Col. Colonel
Com. Commander, Commodore
Cor. Sec. Corresponding Secretary
Corp. Corporal
C.S. Court of Sessions
C.S. (Custos Sigilli ) Keeper of the Seal
D.C.L. Doctor of Civil Law
D.D. Doctor of Divinity
D.D.S. Doctor of Dental Surgery
Dea. Deacon
Dep. Deputy
D.F. Defender of the Faith
D.M. Doctor of Music
Dr. Doctor
D.Sc. Doctor of Science
D.T. (Doctor Theologiae ) Doctor of Divinity
D.V.M. or M.D.V. Doctor of Veterinary Medicine
E. (after titles ) Edinburgh
Esq. Esquire
F.D. (Fidei Defensor ) Defender of the Faith
F.G.S. Fellow of the Geological Society
Fr. Father
F.R.G.S. Fellow of the Royal Geographical Society
F.R.S. Fellow of the Royal Society
F.R.S.A. Fellow of the Royal Society of Arts
F.S.A. Fellow of the Society of Arts
G.C.B. Knight of the Grand Cross of the Bath
G.C.H. Knight of the Grand Cross of Hanover
G.C.M.G. Knight of the Grand Cross, Order of St. Michael
and St. George
Gen. General
Gov. Governor
Govt. Government
G.R. (Georgius Rex ) King George
H.B.M. His or Her Britannic Majesty
H.M. His or Her Majesty
H.M.S. His or Her Majesty’s Service
Hon. Honorable
H.R. House of Representatives
H.R.E. Holy Roman Emperor
H.R.H. His or Her Royal Highness
H.S.H. His or Her Serene Highness
I.N.R.I. (Jesus Nazarenus Rex Judaeorum ) Jesus of
Nazareth, King of the Jews
Insp. Inspector
Insp. Gen. Inspector General
I.O.O.F. Independent Order of Odd Fellows
J.A. Judge-Advocate
J.P. Justice of the Peace
J. Prob. Judge of the Probate
Jr. or Jun. Junior
K. King
K.A. Knight of St. Andrew, in Russia
K.A.N. Knight of Alexander Newski, in Russia
K.B. King’s Bench; Knight of the Bath
K.B.A. Knight of St. Bento d’Avis, in Portugal
K.B.E. Knight of the Black Eagle, in Prussia
K.C. King’s Council; Knight of the Crescent, in
Turkey
K.C.B. Knight Commander of the Bath
K.C.H. Knight Commander of Hanover
K.C.M.G. Knight Commander of Order of St. Michael and
St. George
K.C.S. Knight of Charles III, in Spain
K.E. Knight of the Elephant, in Denmark
K.F. Knight of Ferdinand of Spain
K.F.M. Knight of Ferdinand and Merit, in Sicily
K.G. Knight of the Garter
K.G.C. Knight of the Grand Cross
K.G.C.B. Knight of the Grand Cross of the Bath
K.G.F. Knight of the Golden Fleece
K.G.H. Knight of the Guelph of Hanover
K.G.V. Knight of Gustavus Vasa of Sweden
K.H. Knight of Hanover
K.J. Knight of St. Joachim
K.L.H. Knight of the Legion of Honor
K.M. Knight of Malta
K. Mess. King’s Messenger
K.M.H. Knight of Merit, in Holstein
K.M.J. Knight of Maximilian Joseph of Bavaria
K.M.T. Knight of Maria Theresa of Austria
K.N.S. Knight of the Royal North Star, in Sweden
K.P. Knight of St. Patrick
K.R.E. Knight of the Red Eagle, in Prussia
K.S. Knight of the Sword, in Sweden
K.S.A. Knight of St. Anne of Russia
K.S.E. Knight of St. Esprit, in France
K.S.F. Knight of St. Fernando of Spain
K.S.F.M. Knight of St. Ferdinand and Merit, in Naples
K.S.G. Knight of St. George of Russia
K.S.H. Knight of St. Hubert of Bavaria
K.S.J. Knight of St. Janarius of Naples
K.S.L. Knight of the Sun and Lion, in Persia
K.S.M. & S.G. Knight of St. Michael and St. George of the
Ionian Isles
K.S.P. Knight of St. Stanislaus of Poland
K.S.S. Knight of the Southern Star of the Brazils;
Knight of the Sword, in Sweden
K.S.W. Knight of St. Wladimir of Russia
Kt. Knight
K.T. Knight of the Thistle
K.T.S. Knight of the Tower and Sword, in Portugal
K.W. Knight of William of the Netherlands
K.W.E. Knight of the White Eagle, in Poland
L. (after titles ) London
L.C. Lord Chancellor
L.C.J. Lord Chief Justice
Leg. Legate
Legis. Legislature
Lieut. Lieutenant
Lieut.-Col. Lieutenant-Colonel
Lieut.-Gen. Lieutenant-General
Litt.D. (Litterarum Doctor ) Doctor of Literature
LL.B. (Legum Baccalaureus ) Bachelor of Laws
LL.D. (Legum Doctor ) Doctor of Laws
M. Monsieur
M.A. Master of Arts
Maj. Major
Maj.-Gen. Major-General
M.B. (Medicinae Baccalaureus ) Bachelor of
Medicine; (Musicae Baccalaureus ) Bachelor
of Music
M.C. Member of Congress
M.D. (Medicinae Doctor ) Doctor of Medicine
Messrs. Messieurs
Mgr. Manager; Monsignor
Min. Plen. Minister Plenipotentiary
Mlle. Mademoiselle
Mme. Madame
M.P. Member of Parliament
M.R. Master of the Rolls
Mr. Mister or Master
Mrs. Mistress
Mus. Doc. Doctor of Music
Oxon. (Oxonensis ) Oxford
P.C. (Patres Con­scrip­ti ) Con­script Fath­ers; Sen­a­‐
tors; Privy Coun­sel­lor
Ph.D. Doctor of Philosophy
Ph.G. Graduate in Pharmacy
P.M. Postmaster
P.M.G. Postmaster-General
P.R.A. President of the Royal Academy
Pres. President
Prof. Professor
Prov. Provost
P.R.S. President of the Royal Society
Q. Queen
Q.M. Quartermaster
R.A. Royal Academician
R.E. Royal Engineers
Reg. Prof. Regius Professor
Rev. Reverend
R.M. Royal Marines
R.N. Royal Navy
R.N.O. (Riddare af Nordstjerneorden ) Knight of the
Order of Polar Star
R.S.S. (Regiae Societatis Socius ) Fellow of the Royal
Society
Rt. Hon. Right Honorable
Rt. Rev. Right Reverend
Rt. Wpful. Right Worshipful
R.W. Right Worthy
R.W.O. (Riddare af Wasa Orden ) Knight of the Order
of Wasa
Sec. Secretary
Sec. Leg. Secretary of Legation
Serg. Sergeant
Serg.-Maj. Sergeant-Major
S.J. Society of Jesus
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