Clickable, selective, and cell-permeable activity-based probe of human cathepsin B - Minimalistic approach for enhanced selectivity.

Human cathepsin B is a cysteine-dependent protease whose roles in both normal and diseased cellular states remain yet to be fully delineated. This is primarily due to overlapping substrate specificities and lack of unambiguously annotated physiological functions. In this work, a selective, cell-permeable, clickable and tagless small molecule cathepsin B probe, KDA-1, is developed and kinetically characterized. KDA-1 selectively targets active site Cys25 residue of cathepsin B for labeling and can detect active cellular cathepsin B in proteomes derived from live human MDA-MB-231 breast cancer cells and HEK293 cells. It is anticipated that KDA-1 probe will find suitable applications in functional proteomics involving human cathepsin B enzyme.

The impact of genetically controlled splicing on exon inclusion and protein structure.

Common variants affecting mRNA splicing are typically identified though splicing quantitative trait locus (sQTL) mapping and have been shown to be enriched for GWAS signals by a similar degree to eQTLs. However, the specific splicing changes induced by these variants have been difficult to characterize, making it more complicated to analyze the effect size and direction of sQTLs, and to determine downstream splicing effects on protein structure. In this study, we catalogue sQTLs using exon percent spliced in (PSI) scores as a quantitative phenotype. PSI is an interpretable metric for identifying exon skipping events and has some advantages over other methods for quantifying splicing from short read RNA sequencing. In our set of sQTL variants, we find evidence of selective effects based on splicing effect size and effect direction, as well as exon symmetry. Additionally, we utilize AlphaFold2 to predict changes in protein structure associated with sQTLs overlapping GWAS traits, highlighting a potential new use-case for this technology for interpreting genetic effects on traits and disorders.