Bioinformatics

In Silico Phylogenetic Study Of The Human Tp53 Gene:

Insights Into Evolutionary Dynamics

Journal: Bioinformatics

Manuscript ID BIOINF-2024-2085

Manuscript type: Original Paper

Fo
Date Submitted by the
15-Sep-2024
Author:

Complete List of Authors: Pigili, Akhil Kumar; Sri Venkateswara Institute of Medical Sciences,
rP

Department of bioinformatics
Katika, Anil Kumar Babu; Central Tribal University of Andhra Pradesh,
Department of Biotechnology
ee

Phylogenetics, Visualization, Statistics, Genetic networks, Computational

Portal Keywords:
genomics, Cancer
rR

p53, tumour suppressor gene, Phylogenetics, evolutionary analysis,

Keywords:
Sequence alignment, BLAST
ev
iew
Page 1 of 6 Bioinformatics

1
2
3
4 In Silico Phylogenetic Study Of The Human Tp53 Gene: Insights
5 Into Evolutionary Dynamics
6
7
8 Akhil Kumar Pigili1, Anil Kumar Babu Katika2
9
1Department of Bioinformatics, Sri Venkateswara institute of medical sciences, Tirupati, India
10
2Department of Biotechnology, Central Tribal University Of Andhra Pradesh, Vizianagaram, India
11
12
13
14 The tumour suppressor protein p53, sometimes known as the "protector of the genome," is widely
15 recognised for its crucial function in averting cancer through the regulation of DNA repair, disruption
16 of the cell cycle, and programmed cell death known as apoptosis. Recent research has shown that
17
p53 plays a more complex role in genomic equilibrium than its conventional roles. The Tp53 gene's
18
19 evolutionary linkages are examined in this work while evolutionary studies are crucial to
20 contemporary scientific research. Studies on evolution provide links that are useful for current
21 study. The Homo sapiens tumour protein Tp53 transcript variant-1 mRNA FASTA sequence used in
Fo

22 this work was obtained from the NCBI, after the Multiple Sequence Alignment using UPGMA and
23
24
MEGA were used to conduct evolutionary experiments with other species. Consequently, this
research. Thus, this work will aid current biological research in developing and comprehending the
rP

25
26 evolutionary dynamics and pathways of the Tp53 gene in Clinical Research.
27
28
ee

29
30 Keywords: p53, tumour suppressor gene, phylogenetics, evolutionary analysis, sequence
31 alignment, BLAST
rR

32
33
34
35 Introduction: The p53 gene, also known as TP53 or tumour protein, encodes a protein that
ev

36 controls the cell cycle and thereby inhibits cancer formation. In species with many cell lines,
37 the capacity of cells to suppress cancer is essential. P53's role in preserving rigidity by
38
iew

39 preventing genome changes has given it the nickname "the guardian of the genome" (Strachan
40 and Read, 1999). It has an atomic mass of 53 kilo Dalton, which is typical for cell proteins. It
41 is located on the final region of the human seventeenth chromosome. The structure of p53
42 includes the p53 protein is made up of phosphoproteins that include 393 amino acids. The
43
44
tumour silencer p53 is well-known for its role in cell cycle capture, apoptosis, senescence, and
45 Ferro ptosis. As of late, more evidence has emerged suggesting p53 is also effectively involved
46 in the reconstruction of the cellular digestive system. The quantity of scientific evidence
47 supporting p53's multiple activities in metabolism, normal tissue functioning, and tumour
48
49
progression is expanding at an incredible rate. Owing to the significance of this gene, further
50 research into the regulation and function of p53 is still required in order to provide more
51 effective treatments and diagnostics. Understanding the trends and mechanisms of evolution is
52 crucial for current scientific study, and this gene's phylogenetic analysis plays a key role in that
53
regard. It has shown to be a crucial component in bringing together a wide variety of scientific
54
55 disciplines to analyse and explore evolutionary patterns. The distribution pattern of p53 genetic
56 variation, which is present in several creatures including humans, is also investigated in this
57 work.
58
59
60
 Bioinformatics Page 2 of 6

1
2
3 MATERIALS AND METHODS
4
5 Retrieval of nucleotide sequences of p53 gene
6
7 The nucleotide sequence of Homosapiens tumour protein p53 (Tp53) transcript variant-1 mRNA was
8 retrieved from the NCBI website (https://www.ncbi.nlm.nih.gov) with the accession number of gene
9 bank in FASTA format.
10
11 Determination of identity and similarity (%)
12 Local Sequence Alignment
13
14 The BLAST (Basic Local Alignment Search Tool) technique was utilized to evaluate the
15
identification and similarity (%) of the chosen animal. Homo sapiens cancer protein Tp53
16
17 transcript variant-1 mRNA was obtained from the NCBI website in the FASTA format, and
18 the BLAST N (Nucleotide Blast) was carried out from the NCBI BLAST site
19 (https://blast.ncbi.nlm.nih.gov) website. BLAST (Altschul et al., 1990) was carried out for the
20 Tp53 gene.
21
Fo

22 Phylogenetic Analysis
23
24
The CLUSTALW tool was used to align the Tp53 gene sequences multiple and pairwise. Using
rP

25
26 the web program genome.jp, a phylogenetic analysis of the H. sapiens tumour protein p53
27 (TP53) and transcript variant-1 mRNA sequence was performed using CLUSTAL W. The
28 program created a phylogenetic tree that illustrated the ancestral connection between the
ee

29 sequences. The 1000 replications of the bootstrap method were used to recreate the
30
evolutionary tree. Sequences that are located in the same cluster have a close relationship. The
31
tree displays many groupings that indicate how they relate to one another.
rR

32
33
34 Results and Discussions
35
ev

36 Sequence retrieval
37 H. sapiens tumour protein p53 (TP53), transcript variant-1 mRNA was retrieved from the NCBI
38
iew

in FASTA format. The sequence of the transcript variant-1 mRNA (NM_000546.4) is as the
39
40 following:
>NC_000017.11:c7687490-7668421 Homo sapiens chromosome 17, GRCh38.p14
41
Primary Assembly
42
43
44
45 GATTGGGGTTTTCCCCTCCCATGTGCTCAAGACTGG GACCTGCCCTGTGCAGCTGTGGGTTGATTCCACACC
46 CGCTAAAAGTTTTGAGCTTCTCAAAAGTCTAGAGCCA CCCGCCCGGCACCCGCGTCCGCGCCATGGCCATCT
47 CCGTCCAGGGAGCAGGTAGCTGCTGGGCTCCGGGG ACAAGCAGTCACAGCACATGACGGAGGTTGTGAGGC
48 ACACTTTGCGTTCGGGCTGGGAGCGTGCTTTCCACG GCTGCCCCCACCATGAGCGCTGCTCAGATAGCGATG
49 ACGGTGACACGCTTCCCTGGATTGGCAGCCAGACTG GTCTGGCCCCTCCTCAGCATCTTATCCGAGTGGAAG
50
CCTTCCGGGTCACTGCCATGGAGGAGCCGCAGTCA GAAATTTGCGTGTGGAGTATTTGGATGACAGAAACA
51
52 GATCCTAGCGTCGAGCCCCCTCTGAGTCAGGAAACA C TTTTCGACATAGTGTGGTGGTGCCCTATGAGCCGCC
53 TTTTCAGACCTATGGAAACTACTTCCTGAAAACAACG TGAGGTTGGCTCTGACTGTACCACCATCCACTACAA
54 TTCTGTCCCCCTTGCCGTCCCAAGCAATGGATGATTT CTACATGTGTAACAGTTCCTGCATGGGCGGCATGAA
55 GATGCTGTCCCCGGACGATATTGAACAATGGTTCAC CCGGAGGCCCATCCTCACCATCATCACACTGGAAGA
56 TGAAGACCCAGGTCCAGATGAAGCTCCCAGAATGCC CTCCAGTGGTAATCTACTGGGACGGAACAGCTTTGA
57
AGAGGCTGCTCCCCCCGTGGCCCCTGCACCAGCAG GGTGCGTGTTTGTGCCTGTCCTGGGAGAGACCGGC
58
59 CTCCTACACCGGCGGCCCCTGCACCAGCCCCCTCCT GCACAGAGGAAGAGAATCTCCGCAAGAAAGGGGAG
60 GGCCCCTGTCATCTTCTGTCCCTTCCCAGAAAACCTA CCTCACCACGAGCTGCCCCCAGGGAGCACTAAGCG
CCAGGGCAGCTACGGTTTCCGTCTGGGCTTCTTGCA AGCACTGCCCAACAACACCAGCTCCTCTCCCCAGCC
TTCTGGGACAGCCAAGTCTGTGACTTGCACGTACTC AAAGAAGAAACCACTGGATGGAGAATATTTCACCCTT
CCCTGCCCTCAACAAGATGTTTTGCCAACTGGCCAA CAGATCCGTGGGCGTGAGCGCTTCGAGATGTTCCGA
Page 3 of 6 Bioinformatics

1
2
3 GAGCTGAATGAGGCCTTGGAACTCAAGGATGCCCAG TAGGTAGAGGGAGTTGTCAAGTCTCTGCTGGCCCAG
4 TAGGTAGAGGGAGTTGTCAAGTCTCTGCTGGCCCAG CCAAACCCTGTCTGACAACCTCTTGGTGAACCTTAGT
5
CCAAACCCTGTCTGACAACCTCTTGGTGAACCTTAGT ACCTAAAAGGAAATCTCACCCCATCCCACACCCTGG
6
7 ACCTAAAAGGAAATCTCACCCCATCCCACACCCTGG AGGATTTCATCTCTTGTATATGATGATCTGGATCCAC
8 AGGATTTCATCTCTTGTATATGATGATCTGGATCCAC CAAGACTTGTTTTATGCTCAGGGTCAATTTCTTTTTTC
9 CAAGACTTGTTTTATGCTCAGGGTCAATTTCTTTTTTC TTTTTTTTTTTTTTTTTTCTTTTTCTTTGAGACTGGGTC
10 TTTTTTTTTTTTTTTTTTCTTTTTCTTTGAGACTGGGTC TCGCTTTGTTGCCCAGGCTGGAGTGGAGTGGCGTGA
11
TCGCTTTGTTGCCCAGGCTGGAGTGGAGTGGCGTGA TCTTGGCTTACTGCAGCCTTTGCCTCCCCGGCTCGA
12
13 TCTTGGCTTACTGCAGCCTTTGCCTCCCCGGCTCGA GCAGTCCTGCCTCAGCCTCCGGAGTAGCTGGGACC
14 GCAGTCCTGCCTCAGCCTCCGGAGTAGCTGGGACC ACAGGTTCATGCCACCATGGCCAGCCAACTTTTGCA
15 ACAGGTTCATGCCACCATGGCCAGCCAACTTTTGCA TGTTTTGTAGAGATGGGGTCTCACAGTGTTGCCCAG
16 TGTTTTGTAGAGATGGGGTCTCACAGTGTTGCCCAG GCTGGTCTCAAACTCCTGGGCTCAGGCGATCCACCT
17 GCTGGTCTCAAACTCCTGGGCTCAGGCGATCCACCT GTCTCAGCCTCCCAGAGTGCTGGGATTACAATTGTG
18
GTCTCAGCCTCCCAGAGTGCTGGGATTACAATTGTG AGCCACCACGTCCAGCTGGAAGGGTCAACATCTTTT
19
20 AGCCACCACGTCCAGCTGGAAGGGTCAACATCTTTT ACATTCTGCAAGCACATCTGCATTTTCACCCCACCCT
21 ACATTCTGCAAGCACATCTGCATTTTCACCCCACCCT TCCCCTCCTTCTCCCTTTTTATATCCCATTTTTATATC
Fo

22 TCCCCTCCTTCTCCCTTTTTATATCCCATTTTTATATC GATCTCTTATTTTACAATAAAACTTTGCTGCCACCTGT
23 GATCTCTTATTTTACAATAAAACTTTGCTGCCACCTGT GTGTCTGAGGGGT
24
rP

25
26
27
Local sequence alignment
28
ee

29 Homo sapiens cancer protein Tp53 transcript variant-1 mRNA was obtained from the NCBI website
30
31 in the FASTA format, and the BLAST N (Nucleotide Blast) was carried out from the NCBI BLAST
rR

32 site (https://blast.ncbi.nlm.nih.gov) website. BLAST (Altschul et al., 1990) was carried out for the
33 Tp53 gene and the results of the BLAST were formulated in a table (table.1) showcasing the E-value,
34 Accession Number, Percent Identity and organism and Sequence details.
35
ev

36
37
38
iew

39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58 Fig.1 Genomic Context of Tp53 Gene
59
60
 Bioinformatics Page 4 of 6

1
2
Table.1 BLAST Table of Homo sapiens cancer protein Tp53 transcript variant-1 mRNA Gene
3
4
5 S/N AC Number E- Organism Description Seq PW
6 Value Length Identity
7 1 0 100
NM_000546.6 Homo sapiens Homo sapiens tumour protein p53 (TP53), transcript variant 1, mRNA 2512
8
9 2 0 100
HG975427.1 Homo sapiens TPA: Homo sapiens Processed transcript p53-mRNA (p53 gene) 2586
10 3 0 99.88
11 NM_001276761.3 Homo sapiens Homo sapiens tumour protein p53 (TP53), transcript variant 2, mRNA 2509

Fo
12 4 0 99.88
13 AF307851.1 Homo sapiens Homo sapiens p53 protein mRNA, complete cds 2521

rP
14 5 0 Homo sapiens tumour protein p53, mRNA (cDNA clone MGC:646 IMAGE:3544714), 99.88
15 BC003596.1 Homo sapiens complete cds 2508

ee
16 6 0 99.36
17 XM_001172077.6 Pan troglodytes PREDICTED: Pan troglodytes tumour protein p53 (TP53), transcript variant X2, mRNA 2537
18 7 0 99.36

rR
XM_004058511.5 Gorilla gorilla gorilla PREDICTED: Gorilla gorilla gorilla tumour protein p53 (TP53), transcript variant X2, mRNA 2529
19
8 0 99.2
20 XM_003810066.5 Pan paniscus PREDICTED: Pan paniscus tumour protein p53 (TP53), transcript variant X2, mRNA 2555

ev
21 9 0 99.09
22 DQ191317.1 Homo sapiens Homo sapiens p53 protein (TP53) mRNA, complete cds, alternatively spliced 2584

iew
23 10 0 99.09
24 FJ207420.1 Homo sapiens Homo sapiens mutant p53 mRNA, complete cds 2513
25 11 0 99.09
DQ286964.1 Homo sapiens Homo sapiens p53 protein (TP53) mRNA, complete cds, alternatively spliced 2529
26
27 12 0 100
NM_001407262.1 Homo sapiens Homo sapiens tumour protein p53 (TP53), transcript variant 9, mRNA 2615
28 13 0 100
29 JN900492.1 Homo sapiens Homo sapiens tumour suppressor TP53 (TP53) mRNA, complete cds, alternatively spliced 2697
30 14 0 100
31 NM_001126118.2 Homo sapiens Homo sapiens tumour protein p53 (TP53), transcript variant 8, mRNA 2629
32 15 0 99.3
33 XM_016931470.3 Pan troglodytes PREDICTED: Pan troglodytes tumour protein p53 (TP53), transcript variant X1, mRNA 2634
34 16 0 99.13
XM_057300362.2 Pan paniscus PREDICTED: Pan paniscus tumour protein p53 (TP53), transcript variant X1, mRNA 2623
35
17 0 100
36 NM_001407269.1 Homo sapiens Homo sapiens tumour protein p53 (TP53), transcript variant 12, mRNA 2642
37 18 0 100
38 NM_001276696.3 Homo sapiens Homo sapiens tumour protein p53 (TP53), transcript variant 3, mRNA 2645
39 19 0 100
40 NM_001276695.3 Homo sapiens Homo sapiens tumour protein p53 (TP53), transcript variant 4, mRNA 2572
41
42
43
44
45
46
Page 5 of 6 Bioinformatics

1
2
3 Phylogenetic Analysis
4
5
Finding evidence of directional selection in molecular evolution is a common step in
6
7 phylogenetic research (Koref et al., 2003; Hsu et al., 2005; Hofmann et al., 2003; Yang and
8 Bielawski, 2000). TP53 evolution was examined in a variety of taxa, and sequence modifications
9 were shown to be adaptive. The UPGMA rooted tree diagram of H. sapiens tumour protein p53
10 (TP53), transcript variant-1 mRNA sequence showed different clusters formation. Organism that
11 originated from same ancestors having same e-value and 100% pair wise identity, are placed in
12 same clusters whereas those which are distant from each other are placed in separate clusters.
13
14
Phylogenetic reconstructions and an alignment of the H. sapiens tumor protein p53 (TP53) transcript
15 variant-1 mRNA sequence were carried out using the "build" function of ETE3 3.1.3 (Huerta-Cepas
16 et al., 2016) as it was implemented on the GenomeNet (https://www.genome.jp/tools/ete/).The
17 multiple sequence alignment was supplied by the user.
18 The PhyML v20160115 model and settings were used to infer the ML tree: -pinv e --alpha e --
19 nclasses 4 -o tlr -f m --bootstrap 1000 (Guindon et al., 2010). Out of 1000, branch supports are
20
calculated.
21
Fo

22
23 Fig.1 Phylogenetic Analysis of Human Tp53 Gene
24
rP

25
26
27
28
ee

29
30
31
rR

32
33
34
35
ev

36
37
38
iew

39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55 Conclusion
56
57 We may infer that the p53 protein plays a significant role as a tumour suppressor in bovines as well
58 as other species, including humans. Animals that have a high proportion of identity and resemblance
59
have more conserved amino acid sequences. Therefore, this work can offer a platform for
60
researchers to determine animal comparative genomics.
 Bioinformatics Page 6 of 6

1
2
3 References
4
5
6 1. Pintus, Sergey S., et al. "Phylogenetic analysis of the p53 and p63/p73 gene families." In
7 silico biology 7.3 (2007): 319-332.
8 2. Pintus, S. S., et al. "Phylogenetic analysis of the p53 family." Biophysics 51 (2006): 571-580.
9 3. Sahoo, Pravas Ranjan, et al. "Phylogenetic analysis of p53 tumor suppressor gene of bos
10 taurus through in sillico platform." Ovis 92 (2018): 95.
11
4. Yang Z, Bielawski JP (2000). Statistical methods for detecting molecular adaptation. Trends
12
13
Ecol. Evol. 15(12): 496-503.
14 5. Åberg, Emma, et al. "Evolution of the p53-MDM2 pathway." BMC Evolutionary Biology 17
15 (2017): 1-12.
16 6. Rutkowski, Rachael, Kay Hofmann, and Anton Gartner. "Phylogeny and function of the
17 invertebrate p53 superfamily." Cold Spring Harbor perspectives in biology 2.7 (2010):
18 a001131.
19
7. Fernandes, Andrew D., and William R. Atchley. "Biochemical and functional evidence of
20
21 p53 homology is inconsistent with molecular phylogenetics for distant sequences." Journal
Fo

22 of molecular evolution 67 (2008): 51-67.

23 8. Biscotti, Maria Assunta, et al. "The p53 gene family in vertebrates: evolutionary
24 considerations." Journal of Experimental Zoology Part B: Molecular and Developmental
rP

25 Evolution 332.6 (2019): 171-178.

26 9. Hiller, Thomas, et al. "Comparative analysis of 19 genital human papillomavirus types with
27
28
regard to p53 degradation, immortalization, phylogeny, and epidemiologic risk
ee

29 classification." Cancer Epidemiology Biomarkers & Prevention 15.7 (2006): 1262-1267.

30 10. Jain AK, Allton K, Iacovino M, Mahen E, Milczarek RJ, Zwaka TP et al. p53 regulates cell
31 cycle and microRNAs to promote differentiation of human embryonic stem cells. PLoS
rR

32 Biology. 2012; 10(2):e1001268

33 11. Woods YL, Lane DP. Exploiting the p53 pathway for cancer diagnosis and therapy.
34
Hematology Journal. 2003; 4(4):233-247.
35
ev

36
12. Shengkan J, Sebastian M, Woo SJ, Jennifer RW, Nebojsa M, Andrej S et al. Identification
37 and characterization of a p53 homologue in Drosophila melanogaster. Proceedings of the
38 National Academy of Sciences. 2000; 97(13):7301-7306
iew

39 13. Soltis DE, Soltis PS. The Role of Phylogenetics in Comparative Genetics. Plant Physiology.
40 2003; 132:1790-1800.
41 14. Pintusa SS, Fomin ES, Ivanisenko VA, Kolchanov NA. Phylogenetic Analysis of the p53
42
Family. Biophysics. 2006; 51(4):571-580.
43
44 15. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ. Gapped
45 BLAST and PSI BLAST a new generation of protein database search programme. Nucleic
46 acids research. 1997; 25:3389-3402.
47
48
49
50
51
52
53
54
55
56
57
58
59
60