Consolidated BRCA1/2 Variant Interpretation by MH BRCA Correlates with Predicted PARP Inhibitor Efficacy Association by MH Guide.

BRCA1/2 variants are prognostic biomarkers for hereditary breast and/or ovarian cancer (HBOC) syndrome and predictive biomarkers for PARP inhibition. In this study, we benchmarked the classification of BRCA1/2 variants from patients with HBOC-related cancer using MH BRCA, a novel computational technology that combines the ACMG guidelines with expert-curated variant annotations. Evaluation of BRCA1/2 variants (n = 1040) taken from four HBOC studies showed strong concordance within the pathogenic (98.1%) subset. Comparison of MH BRCA's ACMG classification to ClinVar submitter content from ENIGMA, the international consortium of investigators on the clinical significance of BRCA1/2 variants, the ARUP laboratories, a clinical testing lab of the University of UTAH, and the German Cancer Consortium showed 99.98% concordance (4975 out of 4976 variants) in the pathogenic subset. In our patient cohort, refinement of patients with variants of unknown significance reduced the uncertainty of cancer-predisposing syndromes by 64.7% and identified three cases with potential family risk to HBOC due to a likely pathogenic variant BRCA1 p.V1653L (NM_007294.3:c.4957G > T; rs80357261). To assess whether classification results predict PARP inhibitor efficacy, contextualization with functional impact information on DNA repair activity were performed, using MH Guide. We found a strong correlation between treatment efficacy association and MH BRCA classifications. Importantly, low efficacy to PARP inhibition was predicted in 3.95% of pathogenic variants from four examined HBOC studies and our patient cohort, indicating the clinical relevance of the consolidated variant interpretation.

Insight into the assembly properties and functional organisation of the magnetotactic bacterial actin-like homolog, MamK.

Magnetotactic bacteria (MTB) synthesize magnetosomes, which are intracellular vesicles comprising a magnetic particle. A series of magnetosomes arrange themselves in chains to form a magnetic dipole that enables the cell to orient itself along the Earth's magnetic field. MamK, an actin-like homolog of MreB has been identified as a central component in this organisation. Gene deletion, fluorescence microscopy and in vitro studies have yielded mechanistic differences in the filament assembly of MamK with other bacterial cytoskeletal proteins within the cell. With little or no information on the structural and behavioural characteristics of MamK outside the cell, the mamK gene from Magnetospirillium gryphiswaldense was cloned and expressed to better understand the differences in the cytoskeletal properties with its bacterial homologues MreB and acitin. Despite the low sequence identity shared between MamK and MreB (22%) and actin (18%), the behaviour of MamK monitored by light scattering broadly mirrored that of its bacterial cousin MreB primarily in terms of its pH, salt, divalent metal-ion and temperature dependency. The broad size variability of MamK filaments revealed by light scattering studies was supported by transmission electron microscopy (TEM) imaging. Filament morphology however, indicated that MamK conformed to linearly orientated filaments that appeared to be distinctly dissimilar compared to MreB suggesting functional differences between these homologues. The presence of a nucleotide binding domain common to actin-like proteins was demonstrated by its ability to function both as an ATPase and GTPase. Circular dichroism and structural homology modelling showed that MamK adopts a protein fold that is consistent with the 'classical' actin family architecture but with notable structural differences within the smaller domains, the active site region and the overall surface electrostatic potential.