Determination of the consequences of VHL mutations on VHL transcripts in renal cell carcinoma

Taylor,Claire; Craven,Rachel   A.; Harnden,Patricia; Selby,Peter  J.; Banks,Rosamonde  E.

doi:10.3892/ijo.2012.1561

Determination of the consequences of VHL mutations on VHL transcripts in renal cell carcinoma

Authors:
- Claire Taylor
- Rachel A. Craven
- Patricia Harnden
- Peter J. Selby
- Rosamonde E. Banks
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Affiliations: Cancer Research UK Cancer Centre Genomics Facility, Leeds Institute of Molecular Medicine, St. James's University Hospital, Leeds, UK, Clinical and Biomedical Proteomics Group, Leeds Institute of Molecular Medicine, St. James's University Hospital, Leeds, UK, Department of Histopathology, St. James's University Hospital, Leeds, UK
Published online on: July 20, 2012 https://doi.org/10.3892/ijo.2012.1561
Pages: 1229-1240
Copyright: © Taylor et al. This is an open access article distributed under the terms of Creative Commons Attribution License [CC BY_NC 3.0].

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Abstract

Genetic and epigenetic changes in the von Hippel-Lindau (VHL) tumour suppressor gene are common in sporadic conventional (clear cell) renal cell carcinoma (ccRCC). The effects on VHL expression are unknown but increased understanding may be relevant clinically, either in terms of prognosis or in therapy selection. We have examined the expression of VHL mutant RNA in 84 ccRCC tumours previously screened for mutations in genomic DNA, 56 of which contained 52 unique mutations or polymorphisms. Based on the predicted change to the primary amino acid sequence, 24 of the mutations were missense, 11 resulted in frameshifts with premature truncation, 9 resulted in immediate truncation at the site of the mutation and 2 were frameshifts which extended the reading frame beyond the normal stop codon. Nine tumours had intronic variants, including substitution of invariant residues at splice sites, deletion of nucleotides spanning the exon-intron junction, an intronic variant of unknown function and the polymorphism c.463+43A>G. Four variants were identified which were present in genomic DNA but not in mRNA. Three of these, all encoding apparent missense changes to the primary amino acid sequence, were located close to the ends of exons, reduced the strength of the splice site and function as null rather than missense variants. One nonsense variant was not detectable in mRNA but all other mutations resulting in premature truncation codons (PTCs) were, suggesting truncating VHL mutations may potentially generate truncated VHL protein. An intronic variant, c.341‑11T>A, previously regarded as of unknown function, is associated with an increased level of skipping of exon 2 and may, therefore, reduce production of pVHL. Our data show that the biological consequences of VHL mutations are not necessarily predictable from the sequence change of the mutation and that for the majority of VHL mutations, the potential for the generation of mutant protein exists.

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1	Gnarra JR, Tory K, Weng Y, et al: Mutations of the VHL tumour suppressor gene in renal carcinoma. Nat Genet. 7:85–90. 1994. View Article : Google Scholar : PubMed/NCBI
2	Latif F, Tory K, Gnarra J, et al: Identification of the von Hippel-Lindau disease tumor suppressor gene. Science. 260:1317–1320. 1993. View Article : Google Scholar : PubMed/NCBI
3	Nordstrom-O’Brien M, van der Luijt RB, van Rooijen E, et al: Genetic analysis of von Hippel-Lindau disease. Hum Mutat. 31:521–537. 2010.PubMed/NCBI
4	Vortmeyer AO, Huang SC, Pack SD, Koch CA, Lubensky IA, Oldfield EH and Zhuang X: Somatic point mutation of the wild-type allele detected in tumours of patients with VHL germline deletion. Oncogene. 21:1167–1170. 2002. View Article : Google Scholar : PubMed/NCBI
5	Nickerson ML, Jaeger E, Shi Y, et al: Improved identification of von Hippel-Lindau gene alterations in clear cell renal tumors. Clin Cancer Res. 14:4726–4734. 2008. View Article : Google Scholar : PubMed/NCBI
6	Young AC, Craven RA, Cohen D, et al: Analysis of VHL gene alterations and their relationship to clinical parameters in sporadic conventional renal cell carcinoma. Clin Cancer Res. 15:7582–7592. 2009. View Article : Google Scholar : PubMed/NCBI
7	Richards FM, Crossey PA, Phipps ME, et al: Detailed mapping of germline deletions of the von Hippel-Lindau disease tumour suppressor gene. Hum Mol Genet. 3:595–598. 1994. View Article : Google Scholar : PubMed/NCBI
8	Richards FM, Schofield PN, Fleming S and Maher ER: Expression of the von Hippel-Lindau disease tumour suppressor gene during human embryogenesis. Hum Mol Genet. 5:639–644. 1996. View Article : Google Scholar : PubMed/NCBI
9	Schoenfeld A, Davidowitz EJ and Burk RD: A second major native von Hippel-Lindau gene product, initiated from an internal translation start site, functions as a tumor suppressor. Proc Natl Acad Sci USA. 95:8817–8822. 1998. View Article : Google Scholar : PubMed/NCBI
10	Bradley JF and Rothberg PG: Processed pseudogene from the von Hippel-Lindau disease gene is located on human chromosome 1. Diagn Mol Pathol. 8:101–106. 1999. View Article : Google Scholar : PubMed/NCBI
11	Woodward ER, Buchberger A, Clifford SC, Hurst LD, Affara NA and Maher ER: Comparative sequence analysis of the VHL tumour suppressor gene. Genomics. 65:253–265. 2000. View Article : Google Scholar : PubMed/NCBI
12	Stebbins CE, Kaelin WG Jr and Pavletich NP: Structure of the VHL-ElonginC-ElonginB complex implications for VHL tumor suppressor function. Science. 284:455–461. 1999. View Article : Google Scholar : PubMed/NCBI
13	Rini BI, Campbell SC and Escudier B: Renal cell carcinoma. Lancet. 373:1119–32. 2009. View Article : Google Scholar : PubMed/NCBI
14	Banks RE, Tirukonda P, Taylor C, et al: Genetic and epigenetic analysis of von Hippel-Lindau (VHL) gene alterations and relationship with clinical variables in sporadic renal cancer. Cancer Res. 66:2000–2011. 2006. View Article : Google Scholar : PubMed/NCBI
15	Herman JG, Graff JR, Myohanen S, Nelking BD and Baylin SB: Methylation-specific PCR a novel PCR assay for methylation status of CpG islands. Proc Natl Acad Sci USA. 93:9821–9826. 1996. View Article : Google Scholar : PubMed/NCBI
16	Reese MG, Eeckman FH, Kulp D and Haussler D: Improved splice site detection in genie. J Comp Biol. 4:311–323. 1997. View Article : Google Scholar : PubMed/NCBI
17	Cartegni L, Wang J, Zhu Z, Zhang MQ and Krainer AR: ESEfinder: a web resource to identify exonic splicing enhancers. Nucleic Acids Res. 31:3568–3571. 2003.PubMed/NCBI
18	Valcarcel J, Gaur RK, Singh R and Green MR: Interaction of U2AF⁶⁵ RS region with pre-mRNA of branch point and promotion base pairing with U2 snRNA. Science. 273:1706–1709. 1996.PubMed/NCBI
19	Lefevre SH, Chauveinc L, Stoppa-Lyonnet D, et al: A T to C mutation in the polypyrimidine tract of the exon 9 splicing site of the RB1 gene responsible for low penetrance hereditary retinoblastoma. J Med Genet. 39:e212002. View Article : Google Scholar : PubMed/NCBI
20	Nakai K and Sakamoto H: Construction of a novel database containing aberrant splicing mutations of mammalian genes. Gene. 141:171–177. 1994. View Article : Google Scholar : PubMed/NCBI
21	Krawczak M, Thomas NST, Hundrieser B, Mort M, Wittig M, Hampe J and Cooper DN: Single base-pair substitutions in exon-intron junctions of human genes: nature, distribution, and consequences for mRNA splicing. Hum Mutat. 28:150–158. 2007. View Article : Google Scholar : PubMed/NCBI
22	Martella M, Salviati L, Casarin A, et al: Molecular analysis of two uncharacterised sequence variants of the VHL gene. J Hum Genet. 51:964–968. 2006. View Article : Google Scholar
23	McNeill A, Rattenberry E, Barber R, Killick P, MacDonald F and Maher ER: Genotype-phenotype correlations in VHL exon deletions. Am J Med Genet Part A. 149A:2147–2151. 2009. View Article : Google Scholar : PubMed/NCBI
24	Zhang MQ: Statistical features of human exons and their flanking regions. Hum Mol Genet. 7:919–932. 1998. View Article : Google Scholar : PubMed/NCBI
25	Wang Z and Burge C: Splicing regulation: from a parts list of regulatory elements to an integrated splicing code. RNA. 14:802–813. 2008. View Article : Google Scholar : PubMed/NCBI
26	Pohlenz J, Rosenthal IM, Weiss RE, Jhiang SM, Burant C and Refetoff S: Congenital hypothyroidism due to mutations in the sodium/iodide symporter. J Clin Invest. 101:1028–1035. 1998. View Article : Google Scholar : PubMed/NCBI
27	Pymar LS, Platt FM, Askham JM, Morrison EE and Knowles MA: Bladder tumour-derived somatic TSC1 missense mutations cause loss of function via distinct mechanisms. Hum Mol Genet. 17:2006–2017. 2008. View Article : Google Scholar : PubMed/NCBI
28	Lorson CI, Hahnen E, Androphy EJ and Wirth B: A single nucleotide in the SMN gene regulates splicing and is responsible for spinal muscular atrophy. Proc Natl Acad Sci USA. 96:6307–6311. 1999. View Article : Google Scholar : PubMed/NCBI
29	Liu W, Qian C and Francke U: Silent mutation induces exon skipping of fibrillin-1 gene in Marfan syndrome. Nat Genet. 16:328–329. 1997. View Article : Google Scholar : PubMed/NCBI
30	Huang C-H, Reid M, Daniels G and Blumenfeld OO: Alteration of splice site selection by an exon mutation in the human glycophorin A gene. J Biol Chem. 268:25902–25908. 1993.PubMed/NCBI
31	Teraoka SN, Telatar M, Becker-Catania S, et al: Splicing defects in the ataxia-telangiectasia gene ATM: underlying mutations and consequences. Am J Hum Genet. 64:1617–1631. 1999. View Article : Google Scholar : PubMed/NCBI
32	Eskesen ST, Eskesen FN and Ruvinsky A: Natural selection affects frequencies of AG and GT dinucleotides at the 5′ and 3′ ends of exons. Genetics. 167:543–550. 2004.PubMed/NCBI
33	Frischmeyer PA and Dietz HC: Nonsense-mediated mRNA decay in health and disease. Hum Mol Genet. 8:1893–1900. 1999. View Article : Google Scholar : PubMed/NCBI
34	Holbrook JA, Neu-Yilik G, Hentze MW and Kulozik AE: Nonsense-mediated decay approaches the clinic. Nat Genet. 36:801–808. 2004. View Article : Google Scholar : PubMed/NCBI
35	Silva AL and Romao L: The mammalian nonsense-mediated mRNA decay pathway: to decay or not to decay! Which players make the decision? FEBS Lett. 583:499–505. 2009.
36	Hamosh A, Rosenstein BJ and Cutting GR: CFTR nonsense mutations G542X and W1282X associated with severe reduction of CFTR mRNA in nasal epithelial cells. Hum Mol Genet. 1:542–544. 1992. View Article : Google Scholar : PubMed/NCBI
37	Gossage DL, Norby-Slycord CJ, Hershfield MS and Markert ML: A homozygous 5 base-pair deletion in exon 10 of the adenosine deaminase (ADA) gene in a child with severe combined immunodeficiency and very low levels of ADA mRNA and protein. Hum Mol Genet. 2:1493–1494. 1993. View Article : Google Scholar : PubMed/NCBI
38	Rajavel KS and Neufeld EF: Nonsense-mediated decay of human HEXA mRNA. Mol Cel Biol. 21:5512–5519. 2001. View Article : Google Scholar : PubMed/NCBI
39	Perrin-Vidoz L, Sinilnikova OM, Stoppa-Lyonnet D, Lenoir GM and Mazoyer S: The nonsense-mediated mRNA decay pathway triggers degradation of most BRCA1 mRNAs bearing premature truncation codons. Hum Mol Genet. 11:2805–2814. 2002. View Article : Google Scholar : PubMed/NCBI
40	Inacio A, Silva AL, Pinto J, et al: Nonsense mutations in close proximity to the initiation codon fail to trigger full nonsense-mediated mRNA decay. J Biol Chem. 279:32170–32180. 2004. View Article : Google Scholar : PubMed/NCBI
41	Nagy E and Maquat LE: A rule for termination-codon position within intron-containing genes: when nonsense affects RNA abundance. Trends Biochem Sci. 23:198–199. 1998. View Article : Google Scholar : PubMed/NCBI
42	Ong KR, Woodward ER, Killick P, Lim C, Macdonald F and Maher ER: Genotype-phenotype correlations in von Hippel-Lindau disease. Hum Mutat. 28:143–149. 2007. View Article : Google Scholar : PubMed/NCBI
43	Micale L, Muscarella LA, Marzulli M, et al: VHL frameshift mutation as target of nonsense-mediated mRNA decay in Drosophila melanogaster and human HEK293 cell line. J Biomed Biotechnol. 2009:8607612009. View Article : Google Scholar : PubMed/NCBI
44	Herman JG, Latif F, Weng Y, et al: Silencing of the VHL tumour-suppressor gene by DNA methylation in renal carcinoma. Proc Natl Acad Sci USA. 91:9700–9704. 1994. View Article : Google Scholar : PubMed/NCBI
45	Zbar B, Kishida T, Chen F, et al: Germline mutations in the von Hippel-Lindau disease (VHL) gene in families from North America, Europe and Japan. Hum Mutat. 8:348–357. 1996. View Article : Google Scholar : PubMed/NCBI
46	Highsmith WE, Burch LH, Zhou Z, et al: A novel mutation in the cystic fibrosis gene in patients with pulmonary disease but normal sweat chloride concentrations. New Engl J Med. 331:974–980. 1994. View Article : Google Scholar : PubMed/NCBI
47	Christie PT, Harding B, Nesbit MA, Whyte MP and Thakker RV: X-linked hypophosphatemia attributable to pseudoexons of the PHEX gene. J Clin Endocrinol Metab. 86:3840–3844. 2001. View Article : Google Scholar : PubMed/NCBI
48	Whaley JM, Naglich J, Gelbert L, et al: Germ-line mutations in the von Hippel-Lindau tumor-suppressor gene are similar to somatic von Hippel-Lindau aberrations in sporadic renal cell carcinoma. Am J Hum Genet. 55:1092–1102. 1994.PubMed/NCBI
49	Maxwell PH, Wiesener MS, Chang GW, et al: The tumour suppressor protein VHL targets hypoxia-inducible factors for oxygen-dependent proteolysis. Nature. 399:271–275. 1999. View Article : Google Scholar : PubMed/NCBI
50	Ikediobi ON, Davies H, Bignell G, et al: Mutation analysis of 24 known cancer genes in the NCI-60 cell line set. Mol Cancer Ther. 5:2606–2612. 2006. View Article : Google Scholar : PubMed/NCBI