Microinjection of A-kinase or elevation of cAMP in fibroblasts caused a transient, reversible loss of actin microfilament bundles, likely via phosphorylation and inhibition of MLCK.
Does A-kinase regulate microfilament integrity through phosphorylation and inhibition of MLCK activity in nonmuscle cells?
A-kinase regulates microfilament integrity in nonmuscle cells by phosphorylating and inhibiting MLCK activity.
Microinjection of the catalytic subunit of cAMP-dependent protein kinase (A-kinase) into living fibroblasts or the treatment of these cells with agents that elevate the intracellular cAMP level caused marked alterations in cell morphology including a rounded phenotype and a complete loss of actin microfilament bundles. These effects were transient and fully reversible. Two-dimensional gel electrophoresis was used to analyze the changes in phosphoproteins from cells injected with A-kinase. These experiments showed that accompanying the disassembly of actin microfilaments, phosphorylation of myosin light chain kinase (MLCK) increased and concomitantly, the phosphorylation of myosin P-light chain decreased. Moreover, inhibiting MLCK activity via microinjection of affinity-purified antibodies specific to native MLCK caused a complete loss of microfilament bundle integrity and a decrease in myosin P-light chain phosphorylation, similar to that seen after injection of A-kinase. These data support the idea that A-kinase may regulate microfilament integrity through the phosphorylation and inhibition of MLCK activity in nonmuscle cells.
Lamb et al. (Wed,) conducted a other in Living nonmuscle cells (fibroblasts). Microinjection of cAMP-dependent protein kinase (A-kinase) or cAMP-elevating agents was evaluated on Cell morphology and actin microfilament bundle integrity. Microinjection of A-kinase or elevation of cAMP in fibroblasts caused a transient, reversible loss of actin microfilament bundles, likely via phosphorylation and inhibition of MLCK.