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81.
Myosin light chain diphosphorylation is enhanced by growth promotion of cultured smooth muscle cells 总被引:1,自引:0,他引:1
Minoru Seto Katsuhiko Sakurada Kristine E. Kamm James T. Stull Yasuharu Sasaki 《Pflügers Archiv : European journal of physiology》1996,432(1):7-13
The characteristics of actively growing smooth muscle cells (a variant, SM-3) were compared with those of growth-arrested
cells with regard to response of myosin light chain (MLC) phosphorylation. Augmented MLC phosphorylation, in particular diphosphorylation,
was observed in actively growing cells when stimulated with 30 μM prostaglandin F2α (PGF2α). The maximum level of diphosphorylation in growing cells was significantly higher than that in growth-arrested cells. The
MLC diphosphorylation was sensitive to protein kinase C down-regulation by phorbol dibutylate and pretreatment by the protein
kinase inhibitors, staurosporine (30 nM) and isoquinoline sulphonamide HA1077 (20 μM). The actively growing cells contained
larger amounts of protein kinase C than growth-arrested cells. The phosphorylation sites of mono- and diphospho-MLC were determined
to be MLC kinase-dependent sites (Thr18, Ser19). The PGF2αconcentration/response curves of MLC diphosphorylation were shifted to the left and upwards in the presence of the protein
phosphatase inhibitor calyculin A. These results suggest that PGF2αstimulation of actively growing SM-3 cells augments MLC kinase-dependent MLC diphosphorylation. Protein kinase C is involved
indirectly in this reaction, possibly through MLC phosphatase-sensitive regulatory mechanisms.
Received: 19 October 1995/Received after revision: 16 January 1996/Accepted: 22 January 1996 相似文献
82.
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J J LiPuma M C Fisher S E Dasen J E Mortensen T L Stull 《The Journal of infectious diseases》1991,164(1):133-136
Eighty-three isolates of Pseudomonas capacia were recovered from respiratory secretions from 12 chronically colonized cystic fibrosis patients and examined by ribotype analysis. In 9 patients, the ribotype of the cultured P. cepacia remained unchanged throughout the entire period of observation, indicating chronic pulmonary colonization with a single strain. In each of the remaining 3 patients, two genetically distinct strains were detected among serial P. cepacia isolates. No significant change in clinical condition was correlated with the change in identity of the colonizing strain. In control experiments, the stability of strain ribotype was demonstrated among isolated that had been subcultured 100 times in vitro and among isolates recovered from chronically colonized mice. These data demonstrate the utility of ribotype analysis and indicate that most chronically colonized cystic fibrosis patients harbor a single strain of P. cepacia for prolonged periods. 相似文献
86.
Myosin light chain kinase from smooth muscle has been shown to be phosphorylated by cyclic AMP-dependent protein kinase, which leads to a decrease in the affinity of the kinase for Ca2+ . calmodulin and, hence, a decrease in enzymatic activity. This event has been proposed as a mechanism for the relaxation of smooth muscle in response to increased intracellular concentrations of cyclic AMP. The ratio of myosin light chain kinase activities measured in the presence of 4 microM or 100 microM Ca2+, at 1 microM calmodulin, permits evaluation of such a change in the calmodulin activation properties of myosin light chain kinase. This activity ratio was decreased by phosphorylation of either purified bovine tracheal smooth muscle myosin light chain kinase, or the endogenous myosin light chain kinase in a homogenate of tracheal smooth muscle, with the addition of the catalytic subunit of cyclic AMP-dependent protein kinase. The ratio was unchanged, however, by activation of the endogenous cyclic AMP-dependent protein kinase in homogenates of tracheal smooth muscle by the addition of cyclic AMP. Incubation of tracheal smooth muscle with isoproterenol, at a concentration sufficient to relax the muscle and to increase phosphorylase a formation, had no effect upon the activity ratio. Incubation of tracheal smooth muscle for 2 hr in the presence of carbachol resulted in a transient increase and then a decrease in myosin light chain phosphate content to control values with no decrease in isometric force. The addition of isoproterenol at 2 hr still resulted in relaxation. These findings are inconsistent with a role of myosin light chain kinase phosphorylation in mediating relaxation of tracheal smooth muscle by beta-adrenergic agonists. Cyclic AMP-dependent effects on cytoplasmic calcium concentrations may be more important in mediating relaxation. 相似文献
87.
Keith M. Olsen Pharm.D. Frank H. Ma M.D. Bruce H. Ackerman Pharm.D. Richard E. Stull Ph.D. 《Pharmacotherapy》1993,13(3):229-232
Study Objective . To evaluate two methods of gastrointestinal decontamination, low-volume whole bowel irrigation (WBI) and activated charcoal, for their ability to prevent absorption of salicylate. Design . Randomized, two-phase crossover study. Setting . A clinical research unit in a university-based teaching hospital. Patients . Six healthy, volunteer men. Interventions . Subjects were assigned to receive 3000 ml WBI or syrup of ipecac 30 ml followed by activated charcoal 50 g in sorbitol, and were crossed over to the other treatment phase after 1 week. All treatments began 30 minutes after ingestion of 3.25 g aspirin. Urine was collected over 24 hours for analysis of total urinary excretion of salicylate. Serial blood samples were collected for salicylate determination and were subjected to pharmacokinetic analysis. Measurements and Main Results . Mean ± SD recovery of salicylate were WBI 48.6 ± 5.4% and ipecac-charcoal 37.0 ± 2.6% from urine (p<0.01). Conclusion . Ipecac-charcoal produced a significantly lower salicylate absorption (peak concentration, AUC) than WBI (p<0.01) and thus was superior to low-volume WBI. 相似文献
88.
89.
Charge replacement near the phosphorylatable serine of
the myosin regulatory light chain mimics aspects of
phosphorylation. 总被引:9,自引:0,他引:9 下载免费PDF全文
H L Sweeney Z Yang G Zhi J T Stull K M Trybus 《Proceedings of the National Academy of Sciences of the United States of America》1994,91(4):1490-1494
Phosphorylation of the myosin regulatory lightchains (RLCs) activates contraction in smooth muscle and modulates forceproduction in striated muscle. RLC phosphorylation changes the net charge in acritical region of the N terminus and thereby may alter interactions between theRLC and myosin heavy chain. A series of N-terminal charge mutations in the humansmooth muscle RLC has been engineered, and the mutants have been evaluated fortheir ability to mimic the phosphorylated form of the RLC when reconstitutedinto scallop striated muscle bundles or into isolated smooth muscle myosin.Changing the net charge in the region from Arg-13 to Ser-19 potentiates force inscallop striated muscle and maintains smooth muscle myosin in an unfoldedfilamentous state without affecting ATPase activity or motility of smooth musclemyosin. Thus, the effect of RLC phosphorylation in striated muscle and itsability to regulate the folded-to-extended conformational transition in smoothmuscle may be due to a simple reduction of net charge at the N terminus of thelight chain. The ability of phosphorylation to regulate smooth musclemyosin's ATPase activity and motility involves a more complexmechanism. 相似文献