Fam65b phosphorylation relieves tonic RhoA inhibition during T cell migration

We previously identified Fam65b as an atypical inhibitor of the small G protein RhoA. Using a conditional model of a Fam65b-deficient mouse, we first show that Fam65b restricts spontaneous RhoAactivation in resting T lymphocytes and regulates intranodal T cell migrationin vivo. We next aimed at understanding, at the molecular level, how the brake that Fam65b exerts on RhoAcan be relieved upon signaling to allow RhoAactivation. Here, we show that chemokine stimulation phosphorylates Fam65b in T lymphocytes. This post-translational modification decreases the affinity of Fam65b for RhoAand favors Fam65b shuttling from the plasma membrane to the cytosol. Functionally, we show that the degree of Fam65b phosphorylation controls some cytoskeletal alterations downstream active RhoAsuch as actin polymerization, as well as T cell migrationin vitro. Altogether, our results show that Fam65b expression and phosphorylation can finely tune the amount of active RhoAin order to favor optimal T lymphocyte motility.