ABSTRACT
The regulatory proteins of Ascaris suum striated skeletal muscle were partially purified and characterized. A tropomyosin isoform (Mr 41K) and three troponin subunits identified as troponin T (Mr 37.5K), troponin I (Mr 25.5K) and troponin C (Mr 18.5K) were purified. Three myosin light chains (Mr 25K, 19K, and 17K) were isolated from washed Ascaris actomyosin; the 19K subunit was phosphorylated in vitro. A calcium/calmodulin-dependent myosin light chain kinase activity was identified in the muscle. In contrast to previously reported data suggesting that Ascaris obliquely striated muscle contraction is regulated by a myosin-mediated mechanism, these data indicate that all of the proteins required for actin-mediated, calcium-dependent muscle contraction are present in this tissue.
Subject(s)
Ascaris/physiology , Calcium/pharmacology , Muscle Contraction/drug effects , Animals , Electrophoresis, Polyacrylamide Gel , Muscle Proteins/isolation & purification , Muscles/drug effects , Muscles/physiology , Myofibrils/ultrastructure , Myosin-Light-Chain Kinase , Myosins/metabolism , Phosphorylation , Protein Kinases/metabolism , Tropomyosin/metabolism , Troponin/metabolismABSTRACT
The kinetics for activation of the cyclic adenosine 3':5'-monophosphate (cyclic AMP)-dependent protein kinase (PKA) and thymidine incorporation into DNA was investigated in epinephrine- and prostaglandin E1 (PGE1)-treated murine P1798 lymphosarcoma cells. A positive correlation between the duration and extent of PKA activation and accumulation of cyclic AMP and inhibition of thymidine incorporation into DNA was observed with both hormones. Epinephrine and PGE1 elevated intracellular cyclic AMP 34- and 14-fold, respectively. All hormone concentrations which increased cyclic AMP accumulation also promoted inhibition of thymidine incorporation into DNA. In addition, dibutyryl cyclic AMP (50 microM) inhibited thymidine incorporation. No difference in the kinetics for activation of PKA was observed when cells were treated with microM epinephrine or PGE1. With both agents, 50% PKA activation was observed when intracellular cyclic AMP concentrations were elevated 6.5-fold, or to 9 pmol/10(6) cells. In the presence of microM epinephrine, the cyclic AMP concentration was approximately 3-fold greater than that required for maximal PKA activation. In this case, the duration of the activation time for PKA was also 3- to 4-fold longer than that observed with 0.1 microM epinephrine. The data are consistent with a mechanism wherein both epinephrine and PGE1 suppress DNA synthesis by a cyclic AMP-mediated cascade of protein phosphorylation. No evidence for independent cyclic AMP or PKA pools which respond independently to either epinephrine or PGE1 could be detected.
