The essential chaperone DNAJC17 activates HSP70 to coordinate RNA splicing and G2-M progression

Molecular chaperones including the heat-shock protein 70-kilodalton (HSP70) family and the J-domain containing protein (JDP) co-chaperones maintain homeostatic balance in eukaryotic cells through regulation of the proteome. The expansive JDP family helps direct specific HSP70 functions, and yet loss of single JDP-encoding genes is widely tolerated by mammalian cells, suggesting a high degree of redundancy. Using a genetic screen of DJPs in human cancer cell lines, we found the RNA recognition motif (RRM) containing DNAJC17 to be pan-essential. Knockdown of DNAJC17 leads to relatively few conserved changes in the abundance of individual mRNAs, but instead deranges gene expression through exon skipping, primarily of genes involved in cell cycle progression. Concordant with cell viability experiments, the C-terminal portions of DNAJC17 are dispensable for restoring splicing and G2-M progression. Our findings identify essential cellular JDPs and suggest that diversification in JDP structure extends the HSP70-JDP system to control divergent processes such as RNA splicing. Two pediatric cancer cell lines, Rh30 and ES8, were transduced with dCas9-KRAB and then with sgRNA targeting the 5'UTR of DNAJC17 or non-targeting control. As an additional control, we included Rh30 and ES8 cells transduced with a cDNA stably expressing a DNAJC17 lacking the C-terminal RRM (DRRM) that underwent knockdown of endogenous DNAJC17. Cells were selected in puromycin for 72 hours following sgRNA transduction prior to lysis and RNA extraction. Each experimental condition was tested in triplicate.