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Updated: Jul 21, 2026
Identifying the Effects of BRCA1 Mutations on Homologous Recombination using Cells that Express Endogenous Wild-type BRCA1
Published on: February 17, 2011
Genetic, functional, and histopathological evaluation of two C-terminal BRCA1 missense variants
Background:
The 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.
Objective:
To 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 Results:
Data 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.
Conclusions:
A 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.
Insights
The BRCA1 G1706A variant is benign, while the A1708E variant shows pathogenic effects like mislocalization and centrosome amplification. Comprehensive analysis is crucial for classifying BRCA variants.
Area of Science:
- Genetics
- Molecular Biology
- Cancer Research
Background:
- The majority of BRCA1 missense variants are unclassified due to unknown effects on protein function and pathogenicity.
- BRCA1 has diverse cellular roles, making single functional assays insufficient for comprehensive mutation assessment.
Discussion:
- The A1708E variant disrupts BRCA1 function, causing cytoplasmic mislocalization and centrosome amplification.
- The G1706A variant exhibits a phenotype similar to wild-type BRCA1.
- Tumor pathology correlates with variant function, with 1708E carriers showing BRCA1-associated features and 1706A carriers showing few.
Key Insights:
- Multifactorial analysis classifies G1706A as benign (odds 1:142).
- Functional assays suggest A1708E is pathogenic (odds 262:1), but it remains unclassified due to not meeting the 1000:1 threshold.
- Subcellular mislocalization is a potential mechanism for A1708E pathogenicity.
Outlook:
- Comprehensive genetic and functional analyses are vital for definitive BRCA variant classification.
- This integrated approach can be applied to characterize other unclassified variants in BRCA1 and BRCA2.
