Fragoso-Luna, A. | bioRxiv Science

bioRxiv2022

A multiparametric anti-aging CRISPR screen uncovers a role for BAF in protein translation

Progeria syndromes are very rare, incurable premature aging conditions recapitulating most aging features. Here, we report the first whole genome, multiparametric CRISPR anti-aging screen, identifying 43 new genes that can reverse multiple aging phenotypes in progeria. The screen was implemented in fibroblasts from Nestor- Guillermo Progeria Syndrome (NGPS) patients, carrying a homozygous p.Ala12Thr mutation in barrier-to-autointegration factor (BAF A12T). The hits were enriched for genes involved in protein translation, protein and RNA transport and osteoclast formation. We further confirmed that BAF A12T drives increased protein translation and translational errors that could directly contribute to premature aging in patients. This work has highlighted the power of multiparametric whole genome synthetic rescue screens to identify new anti-aging genes and uncover novel biology behind progeria-associated cellular dysfunction. One-Sentence SummaryA whole genome multiparametric screen in progeria identifies new pathways that can reverse cellular aging phenotypes.

molecular biology↗

bioRxiv2021

Expanded FLP toolbox for spatiotemporal protein degradation and transcriptomic profiling in C. elegans

Control of gene expression in specific tissues and/or at certain stages of development allows the study and manipulation of gene function with high precision. Site-specific genome recombination by the Flippase (FLP) and Cre enzymes has proven particularly relevant. Joint efforts of many research groups have led to the creation of efficient FLP and Cre drivers to regulate gene expression in a variety of tissues in Caenorhabditis elegans. Here, we extend this toolkit by the addition of FLP lines that drive recombination specifically in distal tip cells, the somatic gonad, coelomocytes and the epithelial P lineage. In some cases, recombination-mediated gene knockouts do not completely deplete protein levels due to persistence of long-lived proteins. To overcome this, we developed a spatiotemporally regulated degradation system for GFP fusion proteins (GFPdeg) based on FLP-mediated recombination. Using two stable nuclear pore proteins, MEL-28/ELYS and NPP-2/NUP85 as examples, we report the benefit of combining tissue-specific gene knockout and protein degradation to achieve complete protein depletion. We also demonstrate that FLP-mediated recombination can be utilized to identify transcriptomes in a C. elegans tissue of interest. We have adapted RNA polymerase DamID (RAPID) for the FLP toolbox and by focusing on a well-characterized tissue, the hypodermis, we show that the vast majority of genes identified by RAPID are known to be expressed in this tissue. These tools allow combining FLP activity for simultaneous gene inactivation and transcriptomic profiling, thus enabling the inquiry of gene function in various complex biological processes.

genetics↗