RNAi Screen of RING/U-Box Domain Ubiquitin Ligases Identifies Critical Regulators of Tissue Regeneration in Planarians

Frontiers in Cell and Developmental Biology

Cell and Developmental Biology

Elizabeth M. Duncan¹  Sarai Alvarez Zepeda²  Ricardo M. Zayas²  Carolina Cano Macip²  Madison Balagtas²  Elizabeth Barajas²  Ionit Iberkleid²  John M. Allen³ 

[1] Deparment of Biology, University of Kentucky, Lexington, KY, United States;Department of Biology, San Diego State University, San Diego, CA, United States;Department of Biology, San Diego State University, San Diego, CA, United States;Deparment of Biology, University of Kentucky, Lexington, KY, United States;

关键词: planarian;    stem cells;    regeneration;    E3 ubiquitin ligases;    PRC1;    Prpf19;    RNF2;   

DOI  :  10.3389/fcell.2021.803419

 received in 2021-10-28, accepted in 2021-12-20,  发布年份 2022

来源: Frontiers

【 摘 要 】

Regenerative processes depend on the interpretation of signals to coordinate cell behaviors. The role of ubiquitin-mediated signaling is known to be important in many cellular and biological contexts, but its role in regeneration is not well understood. To investigate how ubiquitylation impacts tissue regeneration in vivo, we are studying planarians that are capable of regenerating after nearly any injury using a population of stem cells. Here we used RNAi to screen RING/U-box E3 ubiquitin ligases that are highly expressed in planarian stem cells and stem cell progeny. RNAi screening identified nine genes with functions in regeneration, including the spliceosomal factor prpf19 and histone modifier rnf2; based on their known roles in developmental processes, we further investigated these two genes. We found that prpf19 was required for animal survival but not for stem cell maintenance, suggesting a role in promoting cell differentiation. Because RNF2 is the catalytic subunit of the Polycomb Repressive Complex 1 (PRC1), we also examined other putative members of this complex (CBX and PHC). We observed a striking phenotype of regional tissue misspecification in cbx and phc RNAi planarians. To identify genes regulated by PRC1, we performed RNA-seq after knocking down rnf2 or phc. Although these proteins are predicted to function in the same complex, we found that the set of genes differentially expressed in rnf2 versus phc RNAi were largely non-overlapping. Using in situ hybridization, we showed that rnf2 regulates gene expression levels within a tissue type, whereas phc is necessary for the spatial restriction of gene expression, findings consistent with their respective in vivo phenotypes. This work not only uncovered roles for RING/U-box E3 ligases in stem cell regulation and regeneration, but also identified differential gene targets for two putative PRC1 factors required for maintaining cell-type-specific gene expression in planarians.

【 授权许可】

Unknown   
Copyright © 2022 Allen, Balagtas, Barajas, Cano Macip, Alvarez Zepeda, Iberkleid, Duncan and Zayas.