Voltage-gated calcium channels (VGCCs) play critical roles in cardiac and skeletal muscle contractions, hormone and neurotransmitter release, as well as slower processes such as cell proliferation, differentiation, migration and death. Mutations in VGCCs lead to numerous cardiac, muscle and neurological disease, and their physiological function is tightly regulated by kinases, phosphatases, G-proteins, calmodulin and many other proteins. Fifteen years ago, RGK proteins were discovered as the most potent endogenous regulators of VGCCs. They are a family of monomeric GTPases (Rad, Rem, Rem2, and Gem/Kir), in the superfamily of Ras GTPases, and they have two known functions: regulation of cytoskeletal dynamics including dendritic arborization and inhibition of VGCCs. Here we review the mechanisms and molecular determinants of RGK-mediated VGCC inhibition, the physiological impact of this inhibition, and recent evidence linking the two known RGK functions. calcium, channel modulation, trafficking, cardiac physiology, neurobiology, beta subunit Citation:
The cytoskeleton is a major focus of physical studies to understand organization inside cells given its primary role in cell motility, cell division, and cell mechanics. Recently, protein condensation has been shown to be another major intracellular organizational strategy. Here, we report that the microtubule crosslinking proteins, MAP65-1 and PRC1, can form phase separated condensates at physiological salt and temperature without additional crowding agents in vitro. The size of the droplets depends on the concentration of protein. MAP65 condensates are liquid at first and can gelate over time. We show that these condensates can nucleate and grow microtubule bundles that form asters, regardless of the viscoelasticity of the condensate. The droplet size directly controls the number of projections in the microtubule asters, demonstrating that the MAP65 concentration can control the organization of microtubules. When gel-like droplets nucleate and grow asters from a shell of tubulin at the surface, the microtubules are able to re-fluidize the MAP65 condensate, returning the MAP65 molecules to solution. This work implies that there is an interplay between condensate formation from microtubule-associated proteins, microtubule organization, and condensate dissolution that could be important for the dynamics of intracellular organization.
Microtubule organization in cells is essential for the internal structure and coordination of events of intracellular transport, mitosis, and cell motility. For many cell types, microtubule organization is dominated by centrosomal nucleation that use gamma-tubulin to template filaments. Yet, some cell types lack centrosomes or centrioles, such as plant cells. Instead, microtubules nucleate from regions with high concentrations of microtubule binding and nucleating proteins. A mechanism that can drive high local concentrations of nucleators is liquid-liquid phase separation of proteins with intrinsically disordered regions. Here, we report that the plant microtubule nucleator and crosslinking protein, MAP65-1, can form phase separated condensates at physiological salt and temperature without extra crowding agents. These condensates are liquid at first and can mature to gel-like phases over time and with different environmental conditions. We show that these condensates can nucleate and grow microtubule bundles that form asters, regardless of the viscoelasticity of the condensate. When gel-like droplets nucleate and grow asters from a shell of tubulin at the surface, the microtubules are able to re-fluidize the MAP65 condensate. Condensate-induced cytoskeletal formation could be a universal mechanism for organization of the microtubule and actin cytoskeletons in all cell types, especially cells without centrosomes.
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