Actin reorganization at the centrosomal area and the immune synapse regulates polarized secretory traffic of multivesicular bodies in T lymphocytes

Abstract

T-cell receptor stimulation induces the convergence of multivesicular bodies towards the microtubule-organizing center (MTOC) and the polarization of the MTOC to the immune synapse (IS). These events lead to exosome secretion at the IS. We describe here that upon IS formation centrosomal area F-actin decreased concomitantly with MTOC polarization to the IS. PKC{delta}-interfered T cell clones showed a sustained level of centrosomal area F-actin associated with defective MTOC polarization. We analysed the contribution of two actin cytoskeleton-regulatory proteins, FMNL1 and paxillin, to the regulation of cortical and centrosomal F-actin networks. FMNL1{beta} phosphorylation and F-actin reorganization at the IS were inhibited in PKC{delta}-interfered clones. F-actin depletion at the central region of the IS, a requirement for MTOC polarization, was associated with FMNL1{beta} phosphorylation at its C-terminal, autoregulatory region. Interfering all FMNL1 isoforms prevented MTOC polarization; nonetheless, FMNL1{beta} re-expression restored MTOC polarization in a centrosomal area F-actin reorganization-independent manner. Moreover, PKC{delta}-interfered clones exhibited decreased paxillin phosphorylation at the MTOC, which suggests an alternative actin cytoskeleton regulatory pathway. Our results infer that PKC{delta} regulates MTOC polarization and secretory traffic leading to exosome secretion in a coordinated manner by means of two distinct pathways, one involving FMNL1{beta} regulation and controlling F-actin reorganization at the IS, and the other, comprising paxillin phosphorylation potentially controlling centrosomal area F-actin reorganization.