Compressive stress induces dephosphorylation of the myosin regulatory light chain via RhoA phosphorylation by the adenylyl cyclase/protein kinase A signaling pathway.
Mechanical stress that arises due to deformation of the extracellular matrix (ECM) either stretches or compresses cells. The cellular response to stretching has been actively studied. For example, stretching induces phosphorylation of the myosin regulatory light chain (MRLC) via the RhoA/RhoA-associated protein kinase (ROCK) pathway, resulting in increased cellular tension. In contrast, the effects of compressive stress on cellular functions are not fully resolved. The mechanisms for sensing and differentially responding to stretching and compressive stress are not known. To address these questions, we investigated whether phosphorylation levels of MRLC were affected by compressive stress. Contrary to the response in stretching cells, MRLC was dephosphorylated 5 min after cells were subjected to compressive stress. Compressive loading induced activation of myosin phosphatase mediated via the dephosphorylation of myosin phosphatase targeting subunit 1 (Thr853). Because myosin phosphatase targeting subunit 1 (Thr853) is phosphorylated only by ROCK, compressive loading may have induced inactivation of ROCK. However, GTP-bound RhoA (active form) increased in response to compressive stress. The compression-induced activation of RhoA and inactivation of its effector ROCK are contradictory. This inconsistency was due to phosphorylation of RhoA (Ser188) that reduced affinity of RhoA to ROCK. Treatment with the inhibitor of protein kinase A that phosphorylates RhoA (Ser188) induced suppression of compression-stimulated MRLC dephosphorylation. Incidentally, stretching induced phosphorylation of MRLC, but did not affect phosphorylation levels of RhoA (Ser188). Together, our results suggest that RhoA phosphorylation is an important process for MRLC dephosphorylation by compressive loading, and for distinguishing between stretching and compressing cells.
Transdisciplinary Life Science Course, Faculty of Advanced Life Science, Hokkaido University, Sapporo, Japan.
Kenji
K
Takemoto
Transdisciplinary Life Science Course, Faculty of Advanced Life Science, Hokkaido University, Sapporo, Japan; Research Center for Cooperative Projects, Hokkaido University Graduate School of Medicine, Sapporo, Japan.
Seiichiro
S
Ishihara
Transdisciplinary Life Science Course, Faculty of Advanced Life Science, Hokkaido University, Sapporo, Japan.
Takeomi
T
Mizutani
Transdisciplinary Life Science Course, Faculty of Advanced Life Science, Hokkaido University, Sapporo, Japan.
Kazushige
K
Kawabata
Transdisciplinary Life Science Course, Faculty of Advanced Life Science, Hokkaido University, Sapporo, Japan; Research Center for Cooperative Projects, Hokkaido University Graduate School of Medicine, Sapporo, Japan.
Hisashi
H
Haga
D008841:Actin Cytoskeleton / Q000378:metabolism; D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D019245:Compressive Strength / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D008387:Marine Toxins; D051379:Mice; D018994:Myosin Light Chains / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D043263:Myosin-Light-Chain Phosphatase; D010080:Oxazoles / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D054460:rho-Associated Kinases / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D008387:Marine Toxins; D018994:Myosin Light Chains; D010080:Oxazoles; D034741:RNA, Small Interfering; C059041:calyculin A; D054460:rho-Associated Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D009219:Myosin-Light-Chain Kinase; D043263:Myosin-Light-Chain Phosphatase; C510645:Ppp1r12a protein, mouse; D020742:rhoA GTP-Binding Protein; D000262:Adenylyl Cyclases