Automated RNAi screening reveals MRG-1/MRG15 as a barrier for germline to neuron reprogramming in C. elegans

Abstract: Chromatin-regulating factors safeguard cell identities by suppressing ectopic gene expression thereby imposing barriers for transcription factor-induced reprogramming of cell fates. Identifying factors opposing the induction of ectopic cell fates is important to better understand developmental differentiation processes and may improve cellular reprogramming for therapeutic approaches. Reprogramming barriers have previously been studied mostly in tissue cultures. Hence, knowledge about chromatin regulators acting as reprogramming barriers in vivo needs improvement. We therefore made use of the nematode C. elegans as an in vivo gene discovery tool and interrogated its chromatin-regulating factors for being involved in protecting germ cells against reprogramming to neurons by performing automated RNAi screening. We identified the conserved chromodomain factor MRG-1 (MRG15 in humans) as a barrier for converting germ cells into neurons and characterized its protein-protein and genome interactions using IP-MS and ChIP-seq in wild-type animals versus mutants lacking the germline. We found that MRG-1 associates predominantly with genes carrying active histone modifications but it also cooperates with the histone H3K9 methyltransferase SET-26 in order to prevent conversion of germ cells into neurons. Conserved chromatin regulators such as MRG-1 potentially have similar functions in higher organisms as shown previously for the histone chaperone LIN-53 (CAF-1p48/RBBP7). LIN-53 was initially identified as a barrier of C. elegans germ cell reprogramming and was later shown to also block reprogramming in mammals. Such remarkable conservation of reprogramming barriers illustrates that understanding cell fate protection in C. elegans could help to improve the generation of new cell types for therapeutic approaches in humans. Overall design: Two c. elegans worm strains ( N2 and glp-4(bn2) ) were analysed in triplicates using ChIP-seq to identify binding regions of chromodomain factor MRG-1 (MRG15 in humans).