Myosin phosphatase targeting subunit 1 affects cell migration by regulating myosin phosphorylation and actin assembly.
Myosin II plays important roles in many contractile-like cell functions, including cell migration, adhesion, and retraction. Myosin II is activated by regulatory light chain (RLC) phosphorylation whereas RLC dephosphorylation by myosin light chain phosphatase containing a myosin phosphatase targeting subunit (MYPT1) leads to myosin inactivation. HeLa cells contain MYPT1 in addition to a newly identified human variant 2 containing an internal deletion. RLC dephosphorylation, cell migration, and adhesion were inhibited when either or both MYPT1 isoforms were knocked down by RNA interference. RLC was highly phosphorylated (60%) when both isoforms were suppressed by siRNA treatment relative to control cells (10%) with serum-starvation and ROCK inhibition. Prominent stress fibers and focal adhesions were associated with the enhanced RLC phosphorylation. The reintroduction of MYPT1 or variant 2 in siRNA-treated cells decreased stress fibers and focal adhesions. MYPT1 knockdown also led to an increase of F-actin relative to G-actin in HeLa cells. The myosin inhibitor blebbistatin did not inhibit this effect, indicating MYPT1 likely affects actin assembly independent of RLC phosphorylation. Proper expression of MYPT1 or variant 2 is critical for RLC phosphorylation and actin assembly, thus maintaining normal cellular functions by simultaneously controlling cytoskeletal architecture and actomyosin activation.
Xia
Donglan
D
Department of Physiology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-9040, USA. Donglan.Xia@utsouthwestern.edu
Stull
James T
JT
Kamm
Kristine E
KE
D000199:Actins / Q000378:metabolism*; D002465:Cell Movement / Q000502:physiology*; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D015536:Down-Regulation / Q000502:physiology; D022001:Focal Adhesions / Q000378:metabolism; D006367:HeLa Cells; D006576:Heterocyclic Compounds, 4 or More Rings / Q000494:pharmacology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D018994:Myosin Light Chains / Q000378:metabolism*; D024462:Myosin Type II / Q000378:metabolism*; D043263:Myosin-Light-Chain Phosphatase / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D009218:Myosins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010766:Phosphorylation; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D034622:RNA Interference; D022502:Stress Fibers / Q000378:metabolism; D054460:rho-Associated Kinases
D000199:Actins; D016895:Culture Media, Serum-Free; D006576:Heterocyclic Compounds, 4 or More Rings; D047908:Intracellular Signaling Peptides and Proteins; D018994:Myosin Light Chains; D020033:Protein Isoforms; C472645:blebbistatin; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D043263:Myosin-Light-Chain Phosphatase; C490372:PPP1R12A protein, human; D024462:Myosin Type II; D009218:Myosins