Sepers, Jorian J. et al. (2021) International Worm Meeting "ERM-1 phosphorylation and NRFL-1 redundantly control lumen formation in the C. elegans intestine"

Kroll, Jason R., Boxem, Mike, Ramalho, Joao J., Sepers, Jorian J., Smidt, Ruben

[International Worm Meeting, 2021]

Reorganization of the plasma membrane and underlying actin cytoskeleton into specialized cortical domains is essential for the function of most polarized cells in animals. ERM and NHERF proteins are conserved regulators of cortical specialization, that function together as membrane-actin linker and organizers of molecular hubs. ERM protein activity involves a conformational switch from an inactive cytoplasmic form into an active membrane- and actin-bound form, which is thought to be mediated by phosphorylation of a conserved C-terminal threonine residue. NHERF proteins can bind to ERM proteins and act as scaffolding proteins by revealing multiple PDZ domains. However, data on the role of the interaction between ERM and NHERF proteins is scarce. Here, we study the relationship between the ERM and NHERF orthologs ERM-1 and NRFL-1, respectively, and their contribution to intestine formation in vivo. Loss of ERM-1 causes defects in tubulogenesis resulting in an early larval arrest, while ERM-1 phosphorylation defective mutants have milder defects and are viable. Using CRISPR/Cas9-generated nrfl-1 alleles we demonstrate that NRFL-1 localizes at the intestinal microvilli, and that this localization is depended on an interaction with ERM-1. However, nrfl-1 loss of function mutants are viable and develop normal intestines. Interestingly, nrfl-1 mutants combined with erm-1 phospho-mutants show severe intestinal defects and an early larval arrest, which closely resembles the ERM-1 mutant phenotype. This data indicate that ERM-1 activity involves phosphorylation of the C-terminal threonine residue and recruitment of NRFL-1. NRFL-1 loss did not affect localization, stability or phosphorylation of ERM-1, suggesting that NRFL-1 does not regulate ERM-1 activity. Therefore, the most likely NRFL-1 contribution to intestinal development is by recruiting and anchoring proteins to the apical cortex through its two PDZ domains. Collectively, our data shows that NRFL-1 is important for the morphology of the C. elegans intestine in an ERM-1 depended manner.