Dataset Information

Probing structure-function relationships in missense variants in the carboxy-terminal region of BRCA1.

ABSTRACT: Germline inactivating variants in BRCA1 lead to a significantly increased risk of breast and ovarian cancers in carriers. While the functional effect of many variants can be inferred from the DNA sequence, determining the effect of missense variants present a significant challenge. A series of biochemical and cell biological assays have been successfully used to explore the impact of these variants on the function of BRCA1, which contribute to assessing their likelihood of pathogenicity. It has been determined that variants that co-localize with structural or functional motifs are more likely to disrupt the stability and function of BRCA1. Here we assess the functional impact of 37 variants chosen to probe the functional impact of variants in phosphorylation sites and in the BRCT domains. In addition, we perform a meta-analysis of 170 unique variants tested by the transcription activation assays in the carboxy-terminal domain of BRCA1 using a recently developed computation model to provide assessment for functional impact and their likelihood of pathogenicity.

SUBMITTER: Carvalho RS

PROVIDER: S-EPMC4028255 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Probing structure-function relationships in missense variants in the carboxy-terminal region of BRCA1.

Carvalho Renato S RS Abreu Renata B V RB Velkova Aneliya A Marsillac Sylvia S Rodarte Renato S RS Suarez-Kurtz Guilherme G Iversen Edwin S ES Monteiro Alvaro N A AN Carvalho Marcelo A MA

PloS one 20140520 5

Germline inactivating variants in BRCA1 lead to a significantly increased risk of breast and ovarian cancers in carriers. While the functional effect of many variants can be inferred from the DNA sequence, determining the effect of missense variants present a significant challenge. A series of biochemical and cell biological assays have been successfully used to explore the impact of these variants on the function of BRCA1, which contribute to assessing their likelihood of pathogenicity. It has ...[more]

PMID: 24845084

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Project description:BackgroundThe vast majority of BRCA1 missense sequence variants remain uncharacterized for their possible effect on protein expression and function, and therefore are unclassified in terms of their pathogenicity. BRCA1 plays diverse cellular roles and it is unlikely that any single functional assay will accurately reflect the total cellular implications of missense mutations in this gene.ObjectiveTo elucidate the effect of two BRCA1 variants, 5236G>C (G1706A) and 5242C>A (A1708E) on BRCA1 function, and to survey the relative usefulness of several assays to direct the characterisation of other unclassified variants in BRCA genes.Methods and resultsData from a range of bioinformatic, genetic, and histopathological analyses, and in vitro functional assays indicated that the 1708E variant was associated with the disruption of different cellular functions of BRCA1. In transient transfection experiments in T47D and 293T cells, the 1708E product was mislocalised to the cytoplasm and induced centrosome amplification in 293T cells. The 1708E variant also failed to transactivate transcription of reporter constructs in mammalian transcriptional transactivation assays. In contrast, the 1706A variant displayed a phenotype comparable to wildtype BRCA1 in these assays. Consistent with functional data, tumours from 1708E carriers showed typical BRCA1 pathology, while tumour material from 1706A carriers displayed few histopathological features associated with BRCA1 related tumours.ConclusionsA comprehensive range of genetic, bioinformatic, and functional analyses have been combined for the characterisation of BRCA1 unclassified sequence variants. Consistent with the functional analyses, the combined odds of causality calculated for the 1706A variant after multifactorial likelihood analysis (1:142) indicates a definitive classification of this variant as "benign". In contrast, functional assays of the 1708E variant indicate that it is pathogenic, possibly through subcellular mislocalisation. However, the combined odds of 262:1 in favour of causality of this variant does not meet the minimal ratio of 1000:1 for classification as pathogenic, and A1708E remains formally designated as unclassified. Our findings highlight the importance of comprehensive genetic information, together with detailed functional analysis for the definitive categorisation of unclassified sequence variants. This combination of analyses may have direct application to the characterisation of other unclassified variants in BRCA1 and BRCA2.

Dataset Information

Probing structure-function relationships in missense variants in the carboxy-terminal region of BRCA1.

Publications

Probing structure-function relationships in missense variants in the carboxy-terminal region of BRCA1.

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