Genetic compensation inpodocalyxin-likemutants during zebrafish liver development

Hepatic stellate cells (HSCs) are critical for normal liver development and regeneration.Podocalyxin-like (podxl)is highly expressed in zebrafish HSCs, but its role in liver development is not known. Here we report thatpodxlknockdown using CRISPR/Cas9 (“CRISPants”) significantly decreased HSC number in zebrafish larvae at different time points and in two independent HSC reporter lines, supporting a role forpodxlin HSC development. We generated fivepodxlmutants, including two mutants lacking the predictedpodxlpromoter region, and found that none of the mutants recapitulated the knockdown phenotype.PodxlCRISPR/Cas9 injection in mutants lacking thepodxlguide RNA cut site did not affect HSC number, supporting the hypothesis that the CRISPant phenotype was specific, requiring intactpodxl. PodxlmRNA levels in threepodxlmutants were similar to those of wildtype controls. RNA sequencing ofpodxlmutants and controls showed no significant change in transcript levels of genes with sequence similarity topodxl, but it revealed upregulation of a network of extracellular matrix genes inpodxlmutants. These results support a role forpodxlin zebrafish liver development and suggest that upregulation of a group of functionally related genes represents the main mechanism of compensation forpodxlgenomic loss.