Implications of a RAD54L polymorphism (2290C/T) in human meningiomas as a risk factor and/or a genetic marker
© Leone et al; licensee BioMed Central Ltd. 2003
Received: 31 October 2002
Accepted: 4 March 2003
Published: 4 March 2003
RAD54L (OMIM 603615, Locus Link 8438) has been proposed as a candidate oncosupressor in tumours bearing a non-random deletion of 1p32, such as breast or colon carcinomas, lymphomas and meningiomas. In a search for RAD54L mutations in 29 menigiomas with allelic deletions in 1p, the only genetic change observed was a silent C/T transition at nucleotide 2290 in exon 18. In this communication the possible association of the 2290C/T polymorphism with the risk of meningiomas was examined. In addition, the usefulness of this polymorphism as a genetic marker within the meningioma consensus deletion region in 1p32 was also verified. The present study comprises 287 blood control samples and 70 meningiomas from Spain and Ecuador. Matched blood samples were only available from Spanish patients.
The frequency of the rare allele-T and heterozygotes for the 2290C/T polymorphism in the blood of Spanish meningioma patients and in the Ecuadorian meningioma tumours was higher than in the control population (P < 0.05). Four other rare variants (2290C/G, 2299C/G, 2313G/A, 2344A/G) were found within 50 bp at the 3' end of RAD54L. Frequent loss of heterozygosity for the 2290C/T SNP in meningiomas allowed to further narrow the 1p32 consensus region of deletion in meningiomas to either 2.08 Mbp – within D1S2713 (44.35 Mbp) and RAD54L (46.43 Mbp) – or to 1.47 Mbp – within RAD54L and D1S2134 (47.90 Mbp) – according to recent gene mapping results.
The statistical analysis of genotypes at the 2290C/T polymorphism suggest an association between the rare T allele and the development of meningeal tumours. This polymorphism can be used as a genetic marker inside the consensus deletion region at 1p32 in meningiomas.
Meningiomas are slow-growing tumours derived from the arachnoid membrane surrounding the central nervous system. They are one of the commonest intracranial tumours, accounting for 20% of all brain tumours, with an overall incidence of 2.3/100,000 and a 2:1 female-to-male ratio . The frequent finding of asymptomatic meningiomas in computerised tomography and magnetic resonance studies  will probably raise these prevalence figures. A close relationship exists between meningiomas and neurofibromatosis type II (NF2). Meningiomas have been found in more than 50% of patients with this common hereditary disorder and sporadic meningiomas show frequent mutations in the NF2 oncosupressor gene [3–5]. Besides NF2, other candidate genes are suspected to be involved in the multistep development of meningiomas; among these genes are those presumably inactivated by deletion/mutation in 1p32, a region of frequent loss of heterozygosity (LOH) in sporadic and hereditary meningiomas [4, 6–9]. Mapped at 1p32 and with likely functions in mitotic and meiotic recombination, RAD54L (OMIM 603615, Locus Link 8438), a member of the SNF2/SWI2 family of DNA-dependent ATPases, has been proposed as a candidate oncosupressor in breast tumours . The finding of mutations in a small fraction of breast carcinomas (1 out of 95 tumours), colon carcinomas (1 out of 13 tumours) and lymphomas (1 out of 24) supports the involvement of RAD54L in tumorigenesis .
In a previous single strand conformation polymorphism (SSCP) analysis of 29 meningiomas with 1 p deletions we failed to detect any mutation in the RAD54 L gene, but found instead a silent C/T polymorphism (Ala730Ala) that was identified by direct sequencing of PCR-amplified exons . This polymorphism has been independently identified (NCB SNP cluster ID: rs1048771) in chromosome 1 contig NT_004386 using high output methods for SNP detection. Blast analysis has unequivocally linked the variation to nucleotide 2290 of RAD54L mRNA (Gen Bank accession X97795.1) or to nucleotide position 2851 in the NCBI RefSeq (accession NM_003579.2); however, lack of information regarding population genotype and allele frequency has precluded its validation as an SNP marker. In this communication the possible association of 2290C/T polymorphism with the risk of meningiomas was examined. In addition, the usefulness of this polymorphism as a genetic marker within the meningioma consensus deletion region in 1p32  was also ascertained.
Twenty-nine Spanish tumours with deletions in 1p and matched blood samples were obtained from meningioma patients as previously described [5, 12]. Forty-one paraffin archival, randomly-chosen, samples of meningiomas (20 meningothelial, 11 transitional, 5 fibroblastic, 3 psamomatose and 2 angioblastic) were obtained from the pathology services of E. Espejo and C. Andrade Marín hospitals in Quito, Ecuador. Two of them were grade II and 39 grade I, according the WHO Classification. No matched blood samples from those patients were available. Spanish and Ecuadorian blood control were obtained from healthy transfusion blood donors.
Genomic DNA was prepared from frozen tissues by standard methods previously described [5, 12]. DNA from paraffin-embedded samples was extracted by standardised protocols . DNA from peripheral lymphocytes was extracted using a DNA-extraction kit from Stratagene, according to the manufacturer's protocol.
PCR based analysis of RAD54L polymorphism and 1p polymorphic markers
The allelic status of 2290 C/T polymorphism in tumour and blood samples was ascertained by PCR amplification of exon 18 followed by SSCP analysis as described . On some occasions confirmatory direct sequencing analysis of PCR products was carried out as described . The allelic status at microsatellite loci was analysed as described [5, 12].
Epidemiological variables and genotypes were evaluated using Tukey-Kramer comparison tests, Odds ratios and Fisher's two-sided exact test.
Results and discussion
Stratification of menigioma patients by sex, age, grade, and histology
P values a (Ecuador vs. Spain)
mean age (years)
P < 0.05
mean age (years)
P < 0.0001
P < 0.0001
Heterozygotes and T-allele frequencies in population controls and meningioma patients. Alleles were identified by exon 18 PCR amplification followed by SSCP as described .
Groups and number of patients or tumour samples
Group comparison for heterozygotes vs. homozygotesb
Group comparison for T-allele frequencyb
A) Control blood from Ecuador N = 149
B) Control blood from Spain N = 87
A-B) OR = 1.160 (0.583 – 2.309) P = 0.7313
A-B) OR = 0.878 (0.5175 – 1.734) P = 0.8776
C) Patients blood from Spaind N = 22
C-B) OR = 4.800 (1.758 – 13.103) P = 0.0036
C-B) OR = 3.421 (1.531 – 7.646) P = 0.0037
D) Meningioma tumours from Spaind,e N = 29
D-B) OR = 0.355 (0.076 – 1.660) P = 0.2327
D-B) OR = 2.357 (1.081 – 5.140) P = 0.0455
E) Meningioma tumours from Ecuadorf N = 41
E-A) OR = 11.285 (5.064 – 25.148) P < 0.0001
E-A) OR = 4.969 (2.773 – 8.905) P = 0.0001
The statistical association between the rare T allele in the 2290C/T polymorphism and the risk of meningioma development (Table 1) requires some attention. It is unlikely that the 2290C/T silent variation (Ala730Ala) could result in functionally different alleles, and there is no evidence for any plausible effect on mRNA transcription. Alternatively, it is possible that the T-allele is in linkage disequilibrium with another sequence of DNA directly implicated in tumour development. According to the draft chromosome 1 sequence (accession NT_004386, in Entrez Nucleotide from NCBI web), RAD54L and MUF1 share 69 bp of a convergent and inverted untranslated mRNA that contains the poly (A+) signal and the site for polyadenylation for both genes (Figure 2). This is a condition for dsRNA production that could initiate a chain reaction of gene silencing through degradation of homologous messenger RNA molecules [22, 23]. Whether the 2290T-allele is actually linked to a gene silencing mechanism through homologous antisense dsRNA of either the RAD54L mRNA or MUF1 mRNA would require dilucidation in further studies, which also have to consider the genomic instability observed at the 3' end of RAD54L.
The statistical analysis of genotype (2290C/T) distribution among meningioma patients and healthy controls suggests that the rare T allele is associated with the risk of meningioma development. In addition, this study also shows that the 2290C/T variation can succesfully be exploited as a polymorphic marker inside the consensus deletion at 1p32 in meningiomas.
List of abbreviations
(polymerase chain reaction)
(single stranded chain polymorfism)
(loss of heterozygosity)
(single nucleotide polymorphism)
This work was supported by grants from the Spanish Ministry of Education and Science (Programa de Cooperación Científica con Iberoamérica). MM is a fellow from the Spanish Ministry of Health (Fondo de Investigación Sanitaria). We are indebted to Paola Nebreda and Javier Pérez for able technical assistance in processing tissue specimens and drawings, respectively. The careful reading of the manuscript by Brenda Ashley is also acknowledged.
- Bondy M, Lignon BL: Epidemiology and etiology of intracranial meningiomas: a review. J Neurooncol. 1996, 29: 197-205.View ArticlePubMedGoogle Scholar
- Kuratsu J, Kochi M, Ushio Y: Incidence and clinical features of asymptomatic meningiomas. J Neurosurg. 2000, 92: 766-770.View ArticlePubMedGoogle Scholar
- Harada T, Irving RM, Xuereb JH, Barton DE, Hardy DG, Moffat DA, Maher ER: Molecular genetic investigation of the neurofibromatosis type 2 tumor suppressor gene in sporadic meningioma. J Neurosurg. 1996, 84: 847-851.View ArticlePubMedGoogle Scholar
- Lamszus K, Vahldiek F, Mautner VF, Schichor C, Tonn J, Stavron D, Fillbrandt , Westphal M, Kluwe L: Allelic loses in neurofibromatosis 2-associated meningiomas. J Neuropathol Exp Neurol. 2000, 59: 504-512.View ArticlePubMedGoogle Scholar
- Leone PE, Bello MJ, de Campos JM, Vaquero J, Sarasa JL, Pestaña A, Rey JA: NF2 mutations and allelic status of 1p, 14q and 22q in sporadic meningiomas. Oncogene. 1999, 18: 2231-2239. 10.1038/sj.onc.1202531.View ArticlePubMedGoogle Scholar
- Sulman EP, Dumanski JP, White PS, Khao H, Maris JM, Mathiesen T, Bruder C, Cnaan A, Brodeur GM: Identification of a consistent region of allelic loss on 1p32 in meningiomas: correlation with increased morbidity. Cancer Res. 1998, 58: 3226-3230.PubMedGoogle Scholar
- Carlson KM, Bruder C, Nordenskjöld M, Dumanski JP: 1p and 3p deletions in meningiomas without detectable aberrations of chromosoma 22 identified by comparative genomic hybridization. Genes Chromosomes Cancer. 1997, 20: 419-424. 10.1002/(SICI)1098-2264(199712)20:4<419::AID-GCC15>3.0.CO;2-H.View ArticlePubMedGoogle Scholar
- Lindblom A, Ruttledge M, Collins VP, Nordenskjöld M, Dumanski JP: Chromosomal deletion in anaplastic meningiomas suggest multiple regions outside chromosome 22 as important in tumor progression. Int J Cancer. 1994, 56: 354-357.View ArticlePubMedGoogle Scholar
- Leuraud P, Marie Y, Robin E, Huguet S, He J, Mokhtari K, Cornu P, Hoang-Xuan K, Sanson M: Frequent loss of 1p32 region but no mutation of the p18 tumor suppressor gene in meningioma. J Neurooncol. 2000, 50: 207-213. 10.1023/A:1006400723490.View ArticlePubMedGoogle Scholar
- Rasio D, Murakum Y, Robbins D, Roth T, Silver A, Negrini M, Schmidt C, Burczak J, Fishel R, Croce CM: Characterization of the human homologue of RAD54: a gene located on chromosome 1p32 at a region of high loss of heterozygosity in breast tumors. Cancer Res. 1997, 57: 2378-2383.PubMedGoogle Scholar
- Matsuda M, Miyagawa K, Takahashi M, Fukuda T, Kataoka T, Asahara T, Inui H, Watatani M, Yasutomi M, Kamada N, Dohi K, Kamiya K: Mutations in the RAD54 recombination gene in primary cancers. Oncogene. 1999, 18: 3427-3430. 10.1038/sj.onc.1202692.View ArticlePubMedGoogle Scholar
- Mendiola M, Bello MJ, Alonso J, Leone PE, Vaquero J, Sarasa JL, Kusak ME, de Campos JM, Pestaña A, Rey JA: Search for mutations of the hRAD54 gene in sporadic meningiomas with deletion at 1p32. Molecular Carcinogenesis. 1999, 24: 300-304. 10.1002/(SICI)1098-2744(199904)24:4<300::AID-MC8>3.0.CO;2-G.View ArticlePubMedGoogle Scholar
- Herrington CS, McGee OD: Diagnostic Molecular Pathology: A Practical Approach. The Practical Approach Series. Oxford, IRL Press. 1992, 93-94.Google Scholar
- Kanaar R, Toelstra C, SMA , Swagemakers Essers J, Smit B, Franssen JH, Pastink A, Bezzubova OY, Buerstedde JM, Clever B, Heyer WD, Hoeijmakers HJ: Human and mouse homologs of the Saccharomyces cerevisiae RAD54 DNA repair gene: evidence for functional conservation. Curr Biol. 1996, 6: 828-838.View ArticlePubMedGoogle Scholar
- Kaul SC, Sugihara T, Yoshida A, Nomura H, Wahwa R: Gros1, a potential grwth suppresor on chromosome 1: its identity to basement membrane-associated proteoglycan, leprecan. Oncogene. 2000, 19: 3576-3583. 10.1038/sj.onc.1203696.View ArticlePubMedGoogle Scholar
- Blais A, Labrie Y, Pouliot F, Lachance Y, Labrie C: Structure of the gene encoding the human cyclin-dependent kinase inhibitor p18 and mutational analysis in breast cancer. Biochem Biophys Res Commun. 1998, 247: 146-153. 10.1006/bbrc.1998.8497.View ArticlePubMedGoogle Scholar
- Begley CG, Visvader J, Green AR, Aplan PD, Metcalf D, Kirsch IR, Gough NM: Molecular cloning and chromosomal localization of the murine homolog of the human helix-loop-helix gene SCL. Proc Natl Acad Sci USA. 1991, 88: 869-873.View ArticlePubMedPubMed CentralGoogle Scholar
- Aplan PD, Jones CA, Chervinsky DS, Zhao X, Ellsworth M, Wu C, McGuire EA, Gross KW: An scl gene product lacking the transactivation domain induces bony abnormalities and cooperates with LMO1 to generate T-cell malignancies in transgenic mice. EMBO J. 1997, 16: 2408-2419. 10.1093/emboj/16.9.2408.View ArticlePubMedPubMed CentralGoogle Scholar
- Smyth I, Narang MA, Evans T, Heimann C, Nakamura Y, Chenevix-Trench G, Pietsch T, Wicking C, Wainwright BJ: Isolation and characterization of human patched 2 (PTCH2), a putative tumour suppressor gene in basal cell carcinoma and medulloblastoma on chromosome 1p32. Hum Mol Genet. 1999, 8: 291-297. 10.1093/hmg/8.2.291.View ArticlePubMedGoogle Scholar
- Slupska MM, Baikalov C, Luther WM, Chiang JH, Wei P, Miller JH: Cloning and sequencing a human homolog (hMYH) of the Escherichia coli mut Y gene whose function is required for the repais of oxidative DNA damage. J Bacteriol. 1996, 178: 3885-3892.PubMedPubMed CentralGoogle Scholar
- Kamura T, Burian D, Yan Q, Schmidt SL, Lane WS, Querido E, Branton PE, Shilatifard A, Conaway RC, Conaway JW: Muf1, a novel Elongin BC-interacting leucine-rich repeat protein that can assemble with Cul5 and Rbx1 to reconstitute a ubiquitin ligase. J Biol Chem. 2001, 276: 29748-29753. 10.1074/jbc.M103093200.View ArticlePubMedGoogle Scholar
- Carthew RW: Gene silencing by double-stranded RNA. Curr Opin Cell Biology. 2001, 13: 244-248. 10.1016/S0955-0674(00)00204-010.1016/S0955-0674(00)00204-0.View ArticleGoogle Scholar
- Nishikura K: A short primer on RNAi: RNA-directed RNA polymerase acts as a key catalist. Cell. 2001, 107: 415-418.View ArticlePubMedGoogle Scholar
- The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2407/3/6/prepub
This article is published under license to BioMed Central Ltd. This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original URL.