Tissue distribution of cysteine string protein/DNAJC5 in C. elegans analysed by CRISPR/Cas9-mediated tagging of endogenous DNJ-14

AbstractCysteine string protein (CSP) is a member of the DnaJ/Hsp40 family of molecular chaperones. CSP is enriched in neurons, where it mainly localises to synaptic vesicles. Mutations in CSP-encoding genes in flies, worms, mice and humans result in neuronal dysfunction, neurodegeneration and reduced lifespan. Most attention has therefore focused on CSP ’s neuronal functions, although CSP is also expressed in non-neuronal cells. Here, we used genome editing to fluorescently tag theCaenorhabditis elegans CSP orthologue,dnj-14, to identify which tissues preferentially express CSP and hence may contribute to the observed mutant phenotypes. Replacement ofdnj-14 with wrmScarlet caused a strong chemotaxis defect, as seen with otherdnj-14 null mutants. In contrast, inserting the reporter in-frame to create a DNJ-14-wrmScarlet fusion protein had no effect on chemotaxis, indicating that C-terminal tagging does not impair DNJ-14 function. WrmScarlet fluorescence appeared most obvious in the intestine, head/pharynx, spermathecae and vulva/uterus in the reporter strains, suggesting that DNJ-14 is preferentially expressed in these tissues. Crossing the DNJ-14-wrmScarlet strain with GFP marker strains confirmed the intestinal and pharyngeal expression, but only a partial overlap with neuronal GFP was observed. DNJ-14-wrmScarlet fluorescence in the intestine was increased in response to starvation, which may be relevant to mammalian CSP α’s role in microautophagy. DNJ-14’s enrichmen...
Source: Cell and Tissue Research - Category: Cytology Source Type: research