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TechNote - For those about to ROCK (we detach you)

Rho-kinase (ROCK) belongs to the AGC family (protein kinase A, protein kinase G and protein kinase C) of serine/threonine kinases and is an effector of the small GTPase, RhoA. The downstream substrates of ROCK modulate actin cytoskeleton organization, stress fiber formation and smooth muscle cell contraction. The roles of ROCK isoforms (ROCK1 and ROCK2) are hypothesized to be somewhat redundant because of a high percentage of homology in the kinase domain and similar downstream targets. However, there is growing evidence of discrepancies between ROCK1 and ROCK2 in a stressful environment. In their publications in Cell Death and Disease, Dr. Shi and colleagues from the Riley Heart Research Center at Indiana University demonstrated dissimilar roles for ROCK1 and ROCK2 in the regulation of cell detachment. Using Enzo's pan-ROCK inhibitor, Y-27632, in wild-type and ROCK-deficient mouse embryonic fibroblasts (MEFs) treated with doxorubicin, they demonstrated that absence of ROCK1, but not ROCK2, reduced cell detachment, favored cell viability and maintained cellular tridimensional organization. Cell detachment from extracellular matrix usually correlates with induction of apoptosis. Interestingly, ROCK1-negative MEFs that have detached in a caspase-independent manner following treatment with doxorubicin and/or Y-27632 were still susceptible to apoptosis. Furthermore, blocking of actomyosin contraction using Enzo’s Blebbistatin mimicked the effects of ROCK1 deficiency in wild-type MEFs after treatment with doxorubicin. Altogether, these results suggest distinct functions for ROCK isoforms in stress-induced stress fiber disassembly and cell detachment and re-emphasize the need for researchers to use isoform-selective compounds to have a complete overview of cellular events at the signaling level.

Enzo Life Sciences offers biologists a comprehensive range of chemicals including inhibitors of ROCK, as well as enzymatic assays and live cell analysis tools, some of which are described below:

 

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