Work performed with differentiated contractile smooth muscle tissue over the last two decades has made clear that covalent modification of myosin by phosphorylation of the 20-kDa myosin light chains is a significant mode of regulation of contractile activity in smooth muscle, particularly in regard to the generation of phasic contractions and the initial development of tonic contractions. This regulatory mechanism appears to be of unique importance in smooth muscle compared with striated muscle. It is equally clear, however, that there is an important role for protein kinase C in the regulation of smooth muscle tone maintenance, particularly in vascular smooth muscle. Several possible signal transduction cascades involving protein kinase C are outlined. Increasing evidence suggests a link between protein kinase C and actin-based regulatory mechanisms. This review places emphasis on relating up-to-date biochemical facts to the physiological realities of the smooth muscle cell.

Controlled regulation of Rho GTPase activity is an essential component mediating growth factor–stimulated migration. We have previously shown that angiomotin (Amot), a membrane-associated scaffold protein, plays a critical role during vascular patterning and endothelial migration during embryogenesis. However, the signaling pathways by which Amot controls directional migration are not known. Here we have used peptide pull-down and yeast 2-hybrid (Y2H) screening to identify proteins that interact with the C-terminal PDZ-binding motifs of Amot and its related proteins AmotL1 and 2. We report that Amot and its related proteins bind to the RhoA GTPase exchange factor (RhoGEF) protein Syx. We show that Amot forms a ternary complex together with Patj (or its paralogue Mupp1) and Syx. Using FRET analysis, we provide evidence that Amot controls targeting of RhoA activity to lamellipodia in vitro. We also report that, similar to Amot, morpholino knockdown of Syx in zebrafish results in inhibition of migration of intersegmental arteries. Taken together, our results indicate that the directional migration of capillaries in the embryo is governed by the Amot:Patj/Mupp1:Syx signaling that controls local GTPase activity.

Proteoglycans participate in growth factor interaction with the cell surface through their heparan sulfate chains (HS), but it is not known if they are otherwise involved in growth factor signaling. It appears now that the syndecan-4 core protein, a transmembrane proteoglycan shown previously to bind phosphatidylinositol 4,5-bisphosphate (PIP₂) and activate PKCα, participates in mediating the effects of fibroblast growth factor (FGF)2 on cell function. Mutations in the cytoplasmic tail of syndecan-4 that either reduced its affinity to PIP₂ (PIP₂⁻) or disrupted its postsynaptic density 95, disk large, zona occludens-1 (PDZ)-dependent binding (PDZ⁻) produced a FGF2-specific dominant negative phenotype in endothelial cells as evidenced by the marked decline of their migration and proliferation rates and the impairment of their capacity to form tubes. In both cases, the molecular mechanism was determined to consist of a decrease in the syndecan-4–dependent activation of PKCα. This decrease was caused either by inhibition of FGF2-induced syndecan-4 dephosphorylation in the case of the PDZ⁻ mutation or by disruption of basolateral targeting of syndecan-4 and its associated PDZ-dependent complex in the case of the PIP₂⁻ mutation. These results suggest that PKCα activation and PDZ-mediated formation of a serine/threonine phosphatase-containing complex by syndecan-4 are downstream events of FGF2 signaling.

Tubular structures are a fundamental anatomic theme recurring in a wide range of animal species. In mammals, tubulogenesis underscores the development of several systems and organs, including the vascular system, the lungs, and the kidneys. All tubular systems are hierarchical, branching into segments of gradually diminishing diameter. There are only 2 cell types that form the lumen of tubular systems: either endothelial cells in the vascular system or epithelial cells in all other organs. The most important feature in determining the morphology of the tubular systems is the frequency and geometry of branching. Hence, deciphering the molecular mechanisms underlying the sprouting of new branches from preexisting ones is the key to understanding the formation of tubular systems. The morphological similarity between the various tubular systems is underscored by similarities between the signaling pathways which control their branching. A prominent feature common to these pathways is their duality—an agonist counterbalanced by an inhibitor. The formation of the tracheal system in Drosophila melanogaster is driven by fibroblast growth factor and inhibited by Sprouty/Notch. In vertebrates, the analogous pathways are fibroblast growth factor and transforming growth factor-β in epithelial tubular systems or vascular endothelial growth factor and Notch in the vascular system.

Myosin VI (myo6) is the only actin-based molecular motor that translocates along actin filaments toward the minus end. Myo6 participates in two steps of endocytic trafficking; it is recruited to both clathrin-coated pits and to ensuing uncoated endocytic vesicles (UCV). Although there is evidence suggesting that the PDZ adaptor protein GIPC/synectin is involved in the association of myo6 with UCV, the recruitment mechanism is unknown. We show that GIPC/synectin is required for both internalization of cell surface receptors and for coupling of myo6 to UCV. This coupling occurs via a mechanism wherein engagement of the GIPC/synectin PDZ domain by C termini of internalized receptors facilitates in trans myo6 binding to the GIPC/synectin C terminus located outside of the PDZ domain. Analysis of megalin, a prototypical GIPC/synectin-binding receptor, revealed that deletion of its PDZ-binding motif drastically reduced GIPC/synectin and myo6 recruitment to UCV. Furthermore, interaction with GIPC/synectin was required for megalin’s function, as megalin was mistargeted in the renal proximal tubules of GIPC/synectin-null mice and these mice exhibited proteinuria, a condition consistent with defective megalin trafficking.