Functional and structural characterization of missense mutations in <i>PAX6</i> gene

doi:10.1007/s11515-015-1346-2

Front. Biol.

2015, Vol. 10

Issue (4) : 377-385 https://doi.org/10.1007/s11515-015-1346-2

RESEARCH ARTICLE

Functional and structural characterization of missense mutations in PAX6 gene

S. Udhaya Kumar,N. Priyanka,P. Sneha,C. George Priya Doss(

)

Medical Biotechnology Division, School of Biosciences and Technology, VIT University, Vellore, Tamil Nadu 632014, India

Download: PDF(661 KB) HTML
Export: BibTeX | EndNote | Reference Manager | ProCite | RefWorks

Abstract

The PAX6 gene belongs to the Paired box (PAX) family of transcription factors that is tissue specific and required for the differentiation and proliferation of cells in embryonic development. PAX6 regulates the pattern formation in early developmental stages. This function of PAX6 protein enables the successful completion of neurogenesis and oculogenesis in most animals such as mice, Drosophila and some other model organisms including humans. A variation in the sequence of PAX6 gene may alter the function and structure of the protein. Such changes can produce adverse effects on functioning of the PAX6 protein which were clinically observed to occur in a broad range of ocular defects such as aniridia in humans. We employed in silico prediction methods such as SIFT, PolyPhen 2; I mutant 3.0, SNAP, SNPs&GO, and PHD-SNP to screen the pathogenic missense mutation in PAX6 and DNA binding sites by BindN and BindN+ . Furthermore, we employed KD4V server to examine the structural and functional modifications that occur in the PAX6 protein as a result of mutation. Based on the results obtained from the in silico prediction methods, we carried out modeling analysis for V53L, I56T, G64V, and I87R to visualize the impact of mutation in structural context.

Keywords PAX6 missense mutation DNA-protein

Corresponding Author(s): C. George Priya Doss

Just Accepted Date: 06 January 2015 Issue Date: 14 August 2015

Cite this article:

S. Udhaya Kumar,N. Priyanka,P. Sneha, et al. Functional and structural characterization of missense mutations in PAX6 gene[J]. Front. Biol., 2015, 10(4): 377-385.

URL:

https://academic.hep.com.cn/fib/EN/10.1007/s11515-015-1346-2
https://academic.hep.com.cn/fib/EN/Y2015/V10/I4/377

Fig.1 Mutational mapping in PAX6. The position of the four highly deleterious mutations have been mapped on the structure of PAX6 protein (PDB ID: 6PAX) using PyMOL 1.7.2.1. The DNA bound to the protein is visualized in cartoon shape in grey color followed by helices in cyan, beta strands in magenta and the loop in salmon color. The native amino acids V53, I56, G64 and I87 are shown in red spheres.

SNPs/Variants	Position	SIFT	PolyPhen2	I-Mutant3.0	SNAP	SNPs&GO	PhD-SNP
VAR_003808	N17S	0.04	0.535	Large decrease	Non-neutral	Disease	Disease
VAR_003809	G18W	0	1	Large increase	Non-neutral	Disease	Disease
VAR_047860	R19P	0	1	Large decrease	Non-neutral	Disease	Neutral
VAR_003810	R26G	0	1	Decrease	Non-neutral	Disease	Disease
VAR_008694	I29S	0	1	Large decrease	Non-neutral	Disease	Neutral
VAR_003811	I29V	0	0.996	Large increase	Neutral	Disease	Neutral
VAR_008695	A33P	0	1	Large decrease	Non-neutral	Disease	Neutral
VAR_008697	I42S	0	0.999	Large decrease	Non-neutral	Disease	Neutral
VAR_008698	S43P	0	0.999	Large increase	Non-neutral	Disease	Disease
VAR_003812	R44Q	0	0.999	Large decrease	Non-neutral	Disease	Neutral
VAR_047861	L46R	0	1	Large increase	Non-neutral	Disease	Disease
VAR_047862	C52R	0	1	Large increase	Non-neutral	Disease	Disease
VAR_008700	V53D	0	1	Large Increase	Non-neutral	Disease	Disease
VAR_008699	V53L	0	0.999	Large decrease	Non-neutral	Disease	Disease
VAR_047863	I56T	0	1	Large decrease	Non-neutral	Disease	Disease
VAR_008701	T63P	0	1	Large decrease	Non-neutral	Disease	Neutral
VAR_008702	G64V	0	1	Large decrease	Non-neutral	Disease	Disease
VAR_017540	P68S	0	1	Decrease	Non-neutral	Disease	Disease
VAR_047864	G73D	0	1	Large Increase	Non-neutral	Disease	Disease
VAR_008703	A79E	0	1	Large Decrease	Non-neutral	Disease	Disease
rs372222637	V83I	0	0.998	Large decrease	Neutral	Disease	Neutral
VAR_047865	I87K	0	0.989	Large decrease	Non-neutral	Disease	Neutral
VAR_003813	I87R	0	1	Large decrease	Non-neutral	Disease	Disease
VAR_015065	P118R	0	1	Large increase	Non-neutral	Disease	Disease
VAR_008704	S119R	0	1	Large decrease	Non-neutral	Disease	Disease
VAR_017541	R125C	0	0.214	Large decrease	Non-neutral	Disease	Disease
VAR_008705	V126D	0	0.999	Large decrease	Non-neutral	Disease	Disease
VAR_003814	R128C	0	0.999	Large decrease	Non-neutral	Disease	Disease
rs371018133	M137I	0.36	0.011	Large decrease	Neutral	Disease	Neutral
rs201983312	Y143C	0	0.926	Large decrease	Neutral	Disease	Disease
rs372143889	R159C	0.06	0.007	Large increase	Non-neutral	Disease	Disease
rs141021880	T166S	0.36	0	Large decrease	Neutral	Disease	Neutral
rs151086737	S167L	0.43	0.012	Large decrease	Non-neutral	Disease	Neutral
VAR_003815	Q178H	0.33	0.77	Large decrease	Neutral	Disease	Neutral
rs138803897	E181K	0.09	0.001	Large decrease	Neutral	Disease	Neutral
rs374396492	G194R	0.01	0.791	Large increase	Neutral	Disease	Disease
VAR_008706	R208Q	0	1	Large increase	Non-neutral	Disease	Disease
VAR_003816	R208W	0	1	Large increase	Non-neutral	Disease	Disease
VAR_047866	R242T	0	0.992	Large increase	Non-neutral	Disease	Disease
VAR_017542	F258S	0	0.986	Large decrease	Non-neutral	Disease	Disease
rs201846044	R267G	0.01	1	Large increase	Non-neutral	Disease	Disease
rs199610944	R267I	0	1	Large decrease	Non-neutral	Disease	Neutral
rs201439078	K270E	0.01	0.751	NA	Non-neutral	Disease	Disease
VAR_017543	S292I	0.39	0.907	Large decrease	Non-neutral	Disease	Disease
VAR_047867	A321T	0.56	0.736	Large decrease	Non-neutral	Disease	Neutral
VAR_008707	S353A	0.27	0.214	Large increase	Neutral	Disease	Neutral
VAR_017544	S363P	0.08	0	Large increase	Neutral	Disease	Neutral
rs201690439	T373S	0.73	0.232	Large decrease	Neutral	Disease	Neutral
VAR_015066	P375Q	0.1	0.498	Large decrease	Neutral	Disease	Disease
VAR_017545	Q378R	0.73	0.22	Large decrease	Neutral	Disease	Neutral
VAR_017546	M381V	0.51	0	Large decrease	Neutral	Disease	Neutral
VAR_047868	G387D	0.71	0.058	Large decrease	Non-neutral	Disease	Neutral
VAR_017547	T391A	0.62	0.001	Large decrease	Neutral	Disease	Neutral
VAR_067698	G395R	0	0.891	Large increase	Non-neutral	Disease	Neutral
VAR_008708	Q422R	0	0.993	Large decrease	Non-neutral	Disease	Neutral

Tab.1 Summary of nsSNP along with their prediction scores of by computational methods in PAX6 gene

Tab.2 Prediction of physico-chemical properties by KD4V server

Tab.3 DNA Binding site determination using PDBSUM, BindN and BindN+ tool

Fig.2 Closer view of the deleterious mutations and their interactions. The structure of the PAX6 protein is shown using cartoon feature in PyMOL 1.7.2.1. The helices have been shown in cyan colour, the beta stands in magenta and the loop in salmon color. Further, the helices and beta strands have been labeled to better understand the location of the mutations in the protein structure. (A) At position 53, the native amino acid valine is substituted with leucine. The native and mutant residues are undergoing polar contact with the surrounding amino acids L57, K55, C52, S49 and N50. (B) At position 56, the native amino acid isoleucine is substituted with threonine. The native residue undergoes polar contact with Y60, R59 and C52, whereas the mutant residue undergoes polar contact with L57. (C) At position 64, the native amino acid glycine is substituted with valine. The native and mutant residues undergo polar contact with the surrounding amino acid Y60. Further the amino acids S65 and T63 are close to the affected residue and will be affected by variations occurring in that position. (D) At position 87, the native amino acid isoleucine is substituted with arginine. At this position, the native residue undergoes polar contact with the surrounding amino acids Y90, K91, V83 and V84.

1	Acharya V, Nagarajaram H A (2012). Hansa: an automated method for discriminating disease and neutral human nsSNPs. Hum Mutat, 33(2): 332–337 https://doi.org/10.1002/humu.21642 pmid: 22045683
2	Adzhubei I A, Schmidt S, Peshkin L, Ramensky V E, Gerasimova A, Bork P, Kondrashov A S, Sunyaev S R (2010). A method and server for predicting damaging missense mutations. Nat Methods, 7(4): 248–249 https://doi.org/10.1038/nmeth0410-248 pmid: 20354512
3	Azuma N, Yamaguchi Y, Handa H, Tadokoro K, Asaka A, Kawase E, Yamada M (2003). Mutations of the PAX6 gene detected in patients with a variety of optic-nerve malformations. Am J Hum Genet, 72(6): 1565–1570 https://doi.org/10.1086/375555 pmid: 12721955
4	Bromberg Y, Rost B (2007). SNAP: predict effect of non-synonymous polymorphisms on function. Nucleic Acids Res, 35(11): 3823–3835 https://doi.org/10.1093/nar/gkm238 pmid: 17526529
5	Bromberg Y, Yachdav G, Rost B (2008). SNAP predicts effect of mutations on protein function. Bioinformatics, 24(20): 2397–2398 https://doi.org/10.1093/bioinformatics/btn435 pmid: 18757876
6	George Priya Doss C, Rajith B, Chakraborty C (2013). Predicting the impact of deleterious mutations in the protein kinase domain of FGFR2 in the context of function, structure, and pathogenesis—a bioinformatics approach. Appl Biochem Biotechnol, 170(8): 1853–1870 https://doi.org/10.1007/s12010-013-0315-y pmid: 23754559
7	Calabrese R, Capriotti E, Fariselli P, Martelli P L, Casadio R (2009). Functional annotations improve the predictive score of human disease-related mutations in proteins. Hum Mutat, 30(8): 1237–1244 https://doi.org/10.1002/humu.21047 pmid: 19514061
8	Capriotti E, Calabrese R, Casadio R (2006). Predicting the insurgence of human genetic diseases associated to single point protein mutations with support vector machines and evolutionary information. Bioinformatics, 22(22): 2729–2734 https://doi.org/10.1093/bioinformatics/btl423 pmid: 16895930
9	Capriotti E, Fariselli P, Rossi I, Casadio R (2008). A three-state prediction of single point mutations on protein stability changes. BMC Bioinformatics, 9(2 Suppl 2): S6 https://doi.org/10.1186/1471-2105-9-S2-S6 pmid: 18387208
10	Davis L K, Meyer K J, Rudd D S, Librant A L, Epping E A, Sheffield V C, Wassink T H (2008). Pax6 3′ deletion results in aniridia, autism and mental retardation. Hum Genet, 123(4): 371–378 https://doi.org/10.1007/s00439-008-0484-x pmid: 18322702
11	George Priya Doss C, Rajith B, Garwasis N, Mathew P R, Raju A S, Apoorva K, William D, Sadhana N R, Himani T, Dike I P (2012). Screening of mutations affecting protein stability and dynamics of FGFR1—A simulation analysis. Applied & Translational Genomics., 1: 37–43 https://doi.org/10.1016/j.atg.2012.06.002
12	Gr?nskov K, Rosenberg T, Sand A, Br?ndum-Nielsen K (1999). Mutational analysis of PAX6: 16 novel mutations including 5 missense mutations with a mild aniridia phenotype. Eur J Hum Genet, 7(3): 274–286 https://doi.org/10.1038/sj.ejhg.5200308 pmid: 10234503
13	Halder G, Callaerts P, Gehring W J (1995). Induction of ectopic eyes by targeted expression of the eyeless gene in Drosophila. Science, 267(5205): 1788–1792 https://doi.org/10.1126/science.7892602 pmid: 7892602
14	Hanson I, Churchill A, Love J, Axton R, Moore T, Clarke M, Meire F, van Heyningen V (1999). Missense mutations in the most ancient residues of the PAX6 paired domain underlie a spectrum of human congenital eye malformations. Hum Mol Genet, 8(2): 165–172 https://doi.org/10.1093/hmg/8.2.165 pmid: 9931324
15	Hanson I M, Fletcher J M, Jordan T, Brown A, Taylor D, Adams R J, Punnett H H, van Heyningen V (1994). Mutations at the PAX6 locus are found in heterogeneous anterior segment malformations including Peters’ anomaly. Nat Genet, 6(2): 168–173 https://doi.org/10.1038/ng0294-168 pmid: 8162071
16	Hill R E, Favor J, Hogan B L, Ton C C, Saunders G F, Hanson I M, Prosser J, Jordan T, Hastie N D, van Heyningen V (1991). Mouse small eye results from mutations in a paired-like homeobox-containing gene. Nature, 354(6354): 522–525 https://doi.org/10.1038/354522a0 pmid: 1684639
17	Kaplan W, Littlejohn T G (2001). Swiss-PDB Viewer (Deep View). Brief Bioinform, 2(2): 195–197 https://doi.org/10.1093/bib/2.2.195 pmid: 11465736
18	Lill M A, Danielson M L (2011). Computer-aided drug design platform using PyMOL. J Comput Aided Mol Des, 25(1): 13–19 https://doi.org/10.1007/s10822-010-9395-8 pmid: 21053052
19	Luu TD, Rusu A, Walter V, Linard B, Poidevin L, Ripp R, Moulinier L, Muller J, Raffelsberger W, Wicker N, Lecompte O, Thompson JD, Poch O, Nguyen H (2012). KD4v: comprehensible knowledge discovery system for missense variant. Nucl Acids Res, W71–W75
20	Matsuo O Y N. Noji S, Ohuchi H, Koyama E, Myokai F, Matsuo N, Taniguchi S, Doi H, Iseki S, Ninomiya Y, Fujiwara M,Watanabe T , Eto K (1993). A mutation in the pax6 gene in rat small eye is associated with impaired migration of mid-brain crest cells. Nat Genet, 3: 299–304 https://doi.org/10.1038/ng0493-299 pmid: 7981749
21	Maulbecker C C, Gruss P (1993). The oncogenic potential of Pax genes. EMBO J, 12(6): 2361–2367 pmid: 8099544
22	McCulley T J, Mayer K, Dahr S S, Simpson J, Holland E J (2005). Aniridia and optic nerve hypoplasia. Eye (Lond), 19(7): 762–764 https://doi.org/10.1038/sj.eye.6701642 pmid: 15359227
23	Mi H, Guo N, Kejariwal A, Thomas P D (2007). PANTHER version 6: protein sequence and function evolution data with expanded representation of biological pathways. Nucleic Acids Res, 35(Database issue): D247–D252 https://doi.org/10.1093/nar/gkl869 pmid: 17130144
24	Mishra R, Gorlov I P, Chao L Y, Singh S, Saunders G F (2002). PAX6, paired domain influences sequence recognition by the homeodomain. J Biol Chem, 277(51): 49488–49494 https://doi.org/10.1074/jbc.M206478200 pmid: 12388550
25	Ng P C, Henikoff S (2003). SIFT: Predicting amino acid changes that affect protein function. Nucleic Acids Res, 31(13): 3812–3814 https://doi.org/10.1093/nar/gkg509 pmid: 12824425
26	Nishikawa K, Ishino S, Takenaka H, Norioka N, Hirai T, Yao T, Seto Y (1994). Constructing a protein mutant database. Protein Eng, 7(5): 733 https://doi.org/10.1093/protein/7.5.733 pmid: 8073043
27	Osumi N, Hirota A, Ohuchi H, Nakafuku M, Iimura T, Kuratani S, Fujiwara M, Noji S, Eto K (1997). Pax-6 is involved in the specification of hindbrain motor neuron subtype. Development, 124(15): 2961–2972 pmid: 9247338
28	Puk O, Yan X, Sabrautzki S, Fuchs H, Gailus-Durner V, Hrabě de Angelis M, Graw J (2013). Novel small-eye allele in paired box gene 6 (Pax6) is caused by a point mutation in intron 7 and creates a new exon. Mol Vis, 19: 877–884 pmid: 23592925
29	Sherry S T, Ward M, Sirotkin K (1999). dbSNP-database for single nucleotide polymorphisms and other classes of minor genetic variation. Genome Res, 9(8): 677–679 pmid: 10447503
30	Stromo G D ( 2000). DNA binding sites: Representation and discovery. Bioinformatics, 16(1): 16–23
31	The UniProt Consortium (2008). The Universal Protein Resource (UniProt). Nucleic Acids Res, 36: D190–D195
32	Tzoulaki I, White I M, Hanson I M (2005). PAX6 mutations: genotype-phenotype correlations. BMC Genet, 6(1): 27 https://doi.org/10.1186/1471-2156-6-27 pmid: 15918896
33	van Heyningen V, Williamson K A (2002). PAX6 in sensory development. Hum Mol Genet, 11(10): 1161–1167 https://doi.org/10.1093/hmg/11.10.1161 pmid: 12015275
34	Wang L, Brown S J(2006). Bind N: A web based tool for efficient prediction of DNA and RNA binding site in amino acid sequences. Nucleic Acids Res, 34: W243–248
35	Wang L, Huang C, Yang MQ, Yang JY ( 2010). BindN+ for accurate prediction of DNA and RNA-binding residue from protein sequence features. BMC Syst Biol, 4: S3
36	Wawersik S, Maas R L (2000). Vertebrate eye development as modeled in Drosophila. Hum Mol Genet, 9(6): 917–925 https://doi.org/10.1093/hmg/9.6.917 pmid: 10767315
37	Wawrocka A, Sikora A, Kuszel L, Krawczynski M R (2013). 11p13 deletions can be more frequent than the PAX6 gene point mutations in Polish patients with aniridia. J Appl Genet, 54(3): 345–351 https://doi.org/10.1007/s13353-013-0154-0 pmid: 23761016

[1]	Merlin LOPUS, Rao SETHUMADHAVAN, P. CHANDRASEKARAN, K. SREEVISHNUPRIYA, A.W. VARSHA, V. SHANTHI, K. RAMANATHAN, R. RAJASEKARAN. A computational approach to explore the functional missense mutations in the spindle check point protein Mad1[J]. Front Biol, 2013, 8(6): 618-625.

Viewed

Full text

Abstract

Cited

Shared

Discussed