The intracellular movement and positioning of organelles and vesicles is mediated by the cytoskeleton and molecular motors. Small GTPases like Rab and Arf proteins are main regulators of intracellular transport by connecting membranes to cytoskeleton motors or adaptors. However, it is becoming clear that interactions between these small GTPases and the cytoskeleton are important not only for the regulation of membrane transport. In this review, we will cover our current understanding of the mechanisms underlying the connection between Rab and Arf GTPases and the cytoskeleton, with special emphasis on the double role of these interactions, not only in membrane trafficking but also in membrane and cytoskeleton remodeling. Furthermore, we will highlight the most recent findings about the fine control mechanisms of crosstalk between different members of Rab, Arf, and Rho families of small GTPases in the regulation of cytoskeleton organization.

Lysosomal signaling facilitates the migration of immune cells by releasing calcium to activate the actin-based motor myosin II at the cell rear. However, how the actomyosin cytoskeleton physically associates to lysosomes is unknown. We have previously identified myosin II as a direct interactor of Rab7b, a small GTPase that mediates the transport from late endosomes/lysosomes to the TGN. Here, we show that Rab7b regulates the migration of dendritic cells (DCs) in 1- and 3-dimensional environments. DCs are immune sentinels that transport antigens from peripheral tissues to lymph nodes to activate T lymphocytes and initiate adaptive immune responses. We found that lack of Rab7b reduces myosin II light chain phosphorylation and the activation of the transcription factor EB (TFEB), which controls lysosomal signaling and is required for fast DC migration. Furthermore, we demonstrate that Rab7b interacts with the lysosomal calcium channel TRPML1, enabling the local activation of myosin II at the cell rear. Altogether, our findings identify Rab7b as the missing physical link between lysosomes and the actomyosin cytoskeleton, allowing control of immune cell migration through lysosomal signaling.

Rab proteins are master regulators of intracellular membrane trafficking but they also contribute to cell division, signaling, polarization and migration. The majority of the works describing the mechanisms used by Rab proteins to regulate cell motility involve intracellular transport of key molecules important for migration. Interestingly, a few studies indicate that Rabs can modulate the activity of Rho GTPases, important regulators for the cytoskeleton rearrangements, but the mechanisms behind this crosstalk are still poorly understood. In this work, we identify Rab6 as a negative regulator of cell migration in vitro and in vivo. We show that loss of Rab6 promotes formation of actin protrusions and influences actomyosin dynamics by upregulating Cdc42 activity and downregulating myosin II phosphorylation. We further provide the molecular mechanism behind this regulation demonstrating that Rab6 interacts with both Cdc42 and Trio, a GEF for Cdc42. In sum, our results uncover a mechanism used by Rab proteins to ensure spatial regulation of Rho GTPase activity for coordination of cytoskeleton rearrangements required in migrating cells.U2OS, HeLa and H1299 cells were grown in Dulbecco´s modified Eagle´s medium (DMEM; Lonza, BioWhittaker). RPE-1 cells were grown DMEM F-12 (Lonza, BioWhittaker). Both DMEM and DMEM F-12 were supplemented with 10 % fetal calf serum (FCS), 2 mM L-glutamine, 100 U/ml penicillin and µg/ml streptomycin. Constructs and antibodies pEGFP-C1 was purchased from BD Biosciences, Clontech. pEGFP-C1 Rab6A wt, pEGFP-C1 Rab6A Q72L and pEGFP-C1 Rab6A T27N were a gift from Marci Scidmore (Addgene plasmid #49469, #49483 and #49484, respectively), [31]. pcDNA3-EGFP-Cdc42-Q61L and pcDNA3-EGFP-Cdc42-T17N were a gift from Gary Bokoch (Addgene plasmid #12986 and #12976, respectively), [32]. pTriEx-mCherry-cdc42 Q61L and the biosensor constructs, pTriEX Cdc42 wt and pTriEX Cdc42 G12V, were a gift from Luis Hodgson (Albert Einstein College of Medicine, NY, USA). mCherry-Cdc42-C-10 was a gift from Michael Davidson (Addgene plasmid # 55014). p CMV-LifeAct-RFP was purchased from Ibidi. pEGFP-C3 Cdc42 wt was a gift from Keith Burridge,