Regulation of molecular motor proteins.

Motor proteins in the kinesin, dynein, and myosin superfamilies are tightly regulated to perform multiple functions in the cell requiring force generation. Although motor proteins within families are diverse in sequence and structure, there are general mechanisms by which they are regulated. We first discuss the regulation of the subset of kinesin family members for which such information exists, and then address general mechanisms of kinesin family regulation. We review what is known about the regulation of axonemal and cytoplasmic dyneins. Recent work on cytoplasmic dynein has revealed the existence of multiple isoforms for each dynein chain, making the study of dynein regulation more complicated than previously realized. Finally, we discuss the regulation of myosins known to be involved in membrane trafficking. Myosins and kinesins may be evolutionarily related, and there are common themes of regulation between these two classes of motors.

Regulation of organelle movement in melanophores by protein kinase A (PKA), protein kinase C (PKC), and protein phosphatase 2A (PP2A).

We used melanophores, cells specialized for regulated organelle transport, to study signaling pathways involved in the regulation of transport. We transfected immortalized Xenopus melanophores with plasmids encoding epitope-tagged inhibitors of protein phosphatases and protein kinases or control plasmids encoding inactive analogues of these inhibitors. Expression of a recombinant inhibitor of protein kinase A (PKA) results in spontaneous pigment aggregation. alpha-Melanocyte-stimulating hormone (MSH), a stimulus which increases intracellular cAMP, cannot disperse pigment in these cells. However, melanosomes in these cells can be partially dispersed by PMA, an activator of protein kinase C (PKC). When a recombinant inhibitor of PKC is expressed in melanophores, PMA-induced pigment dispersion is inhibited, but not dispersion induced by MSH. We conclude that PKA and PKC activate two different pathways for melanosome dispersion. When melanophores express the small t antigen of SV-40 virus, a specific inhibitor of protein phosphatase 2A (PP2A), aggregation is completely prevented. Conversely, overexpression of PP2A inhibits pigment dispersion by MSH. Inhibitors of protein phosphatase 1 and protein phosphatase 2B (PP2B) do not affect pigment movement. Therefore, melanosome aggregation is mediated by PP2A.

Antibodies directed against microtubule proteins from Drosophila melanogaster cross react with similar proteins in the rat brain.

Monoclonal antibodies (Mabs) were used to delineate the localization of three proteins in rat cerebral cortex, hippocampus and cerebellum. The proteins were identified by Mabs directed against Drosophila melanogaster microtubule proteins (MTP). We have provisionally designated these proteins as Drosophila microtubule-associated proteins (DMAPs). The corresponding monoclonal antibodies are designated Mab DMAP-45, -55 and -66 indicating the molecular weights of each protein. All three Mabs cross-react with proteins of similar molecular weights in the rat brain. Correspondingly, these rat proteins are designated DMAPRs. DMAP-45 binds microtubules in an ATP-dependent manner. The molecular weight and subcellular localization of DMAP-45R differs significantly from previously described mammalian brain MAPs suggesting that it represents a novel MAP. Biochemical evidence suggests it may be an actin-related protein. DMAP-55R co-purifies stoichiometrically with rat brain microtubules and appears to be a previously undescribed isoform of tubulin. DMAP-66, which co-purifies stoichiometrically with Drosophila microtubules, does not do so in the rat brain. Immunohistochemistry performed with all three Mabs revealed a general pattern of staining of cell somata and dendrites in the cortex, hippocampus and cerebellum. Mab DMAP-55 also stained axons. In cerebral cortex all three Mabs preferentially, but not exclusively, stained layer V neuronal somata and dendrites. In hippocampus, Mabs DMAP-45 and -66 stained cell somata and dendrites in all hippocampal subfields, particularly the subiculum and CA3, whereas Mab DMAP-55 was most prevalent in mossy fibers. All three Mabs stain Purkinje cells in cerebellum with additional staining of cerebellar basket cells and Golgi cells observed with Mab DMAP-66.