title: PKC activation increases Ca²⁺ sensitivity of permeabilized lymphatic muscle via myosin light chain 20 phosphorylation-dependent and -independent mechanisms.
abstract: The contractile activity of muscle cells lining the walls of collecting lymphatics is responsible for generating and regulating flow within the lymphatic system. Activation of PKC signaling contributes to the regulation of smooth muscle contraction by enhancing sensitivity of the contractile apparatus to Ca(2+). It is currently unknown whether PKC signaling contributes to the regulation of lymphatic muscle contraction. We hypothesized that the activation of PKC signaling would increase the sensitivity of the lymphatic myofilament to Ca(2+). To test this hypothesis, we determined the effects of PKC activation with phorbol esters [PMA or phorbol dibutyrate (PDBu)] on the contractile behavior of α-toxin-permeabilized rat mesenteric and cervical lymphatics or the thoracic duct. The addition of PMA or PDBu induced a significant increase in the contractile force of submaximally activated α-toxin-permeabilized lymphatic muscle independent of a change in intracellular Ca(2+) concentration, and the Ca(2+)-force relationship of lymphatic muscle was significantly left shifted, indicating greater myofilament Ca(2+) sensitivity. Phorbol esters increased the maximal rate of force development, whereas the rate of relaxation was reduced. Western blot and immunohistochemistry data indicated that the initial rapid increase in tension development after stimulation by PDBu was associated with myosin light chain (MLC)20 phosphorylation; however, the later, steady-state Ca(2+) sensitization of permeabilized lymphatic muscle was not associated with increased phosphorylation of MLC20 at Ser(19), 17-kDa C-kinase-potentiated protein phosphatase-1 inhibitor at Thr(38), or caldesmon at Ser(789). Thus, these data indicate that PKC-dependent Ca(2+) sensitization of lymphatic muscle may involve MLC20 phosphorylation-dependent and -independent mechanism(s).
authors.affiliation: Department of Medical Physiology, College of Medicine, Texas A&M Health Science Center, College Station, Texas; and.
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Dougherty
authors.forename: Zhanna V
authors.initials: ZV
authors.lastname: Nepiyushchikh
authors.forename: Sanjukta
authors.initials: S
authors.lastname: Chakraborty
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Davis
authors.forename: David C
authors.initials: DC
authors.lastname: Zawieja
authors.forename: Mariappan
authors.initials: M
authors.lastname: Muthuchamy
mesh_terms: D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects; D002148:Calmodulin-Binding Proteins / Q000378:metabolism; D002463:Cell Membrane Permeability; D004789:Enzyme Activation; D020536:Enzyme Activators / Q000494:pharmacology; D006460:Hemolysin Proteins / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects; D009124:Muscle Proteins / Q000378:metabolism; D009126:Muscle Relaxation; D053580:Muscle Strength; D009130:Muscle, Smooth / Q000187:drug effects / Q000201:enzymology*; D018994:Myosin Light Chains / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012694:Serine; D013897:Thoracic Duct / Q000187:drug effects / Q000201:enzymology*
chemical_list: D001427:Bacterial Toxins; D002148:Calmodulin-Binding Proteins; D020536:Enzyme Activators; D006460:Hemolysin Proteins; D009124:Muscle Proteins; D018994:Myosin Light Chains; D010750:Phosphoproteins; C491519:Ppp1r14a protein, rat; C021091:staphylococcal alpha-toxin; D012694:Serine; D011493:Protein Kinase C
keywords: calcium sensitivity; lymphatic muscle; myosin light chain 20 phosphorylation; protein kinase C activation