Rho-kinase regulates myosin II activation in MDCK cells during recovery after ATP depletion

Sutton, Timothy A.; Mang, Henry; Atkinson, Simon J.

doi:10.1152/ajprenal.2001.281.5.f810

Cited by 31 publications

(35 citation statements)

References 42 publications

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“…These data were supported by observations that the MLCK inhibitor ML-7 did not inhibit epithelial remodeling or MLC-Ser19 phosphorylation. They also suggest that MLC serves as a substrate for Rho kinase during epithelial tubule formation, a model consistent with that previously demonstrated for MDCK and other cells (3,52,54).…”

Section: Discussionsupporting

confidence: 87%

Modulation of epithelial tubule formation by Rho kinase

Eisen

Ratcliffe

Ojakian

2004

American Journal of Physiology-Cell Physiology

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We have developed a model system for studying integrin regulation of mammalian epithelial tubule formation. Application of collagen gel overlays to MadinDarby canine kidney (MDCK) cells induced coordinated disassembly of junctional complexes that was accompanied by lamellipodia formation and cell rearrangement (termed epithelial remodeling). In this study, we present evidence that the Rho signal transduction pathway regulates epithelial remodeling and tubule formation. Incubation of MDCK cells with collagen gel overlays facilitated formation of migrating lamellipodia with membrane-associated actin. Inhibitors of myosin II and actin prevented lamellipodia formation, which suggests that actomyosin function was involved in regulation of epithelial remodeling. To determine this, changes in myosin II distribution, function, and phosphorylation were studied during epithelial tubule biogenesis. Myosin II colocalized with actin at the leading edge of lamellipodia thereby providing evidence that myosin is important in epithelial remodeling. This possibility is supported by observations that inhibition of Rho kinase, a regulator of myosin II function, alters formation of lamellipodia and results in attenuated epithelial tubule development. These data and those demonstrating myosin regulatory light-chain phosphorylation at the leading edge of lamellipodia strongly suggest that Rho kinase and myosin II are important modulators of epithelial remodeling. They support a hypothesis that the Rho signal transduction pathway plays a significant role in regulation of epithelial tubule formation. signaling pathway; polarity EPITHELIA ARE PHYSIOLOGICAL barriers to movement of solutes and fluids in multicellular organisms. To provide these barriers, epithelia are organized into sheets or tubules with individual cells held together by intercellular junctions (15,19). Morphological and biochemical studies have demonstrated that epithelial cells are polarized and have structurally and functionally distinct apical and basolateral membranes (62). Epithelial biogenesis is currently an area of intense study in mammalian cells at both the subcellular and cellular levels (39, 62). Formation and migration of epithelial sheets in embryos are critical events during development (19,31,39) and work toward identification of the factors involved has benefited from genetic analysis of invertebrate model systems (32).An important approach to studying epithelial development has been observation of cellular dynamics and epithelial tubule development in collagen gels. This takes advantage of observations that extracellular matrix (ECM) plays an important role in epithelial biogenesis (7, 39). The Madin-Darby canine kidney (MDCK) cell line has been used in model systems to study ECM and regulation of epithelial tubule formation. Epithelial cysts formed by culturing MDCK cells in suspension have polarized distributions of membrane proteins (59). Resuspension of these cysts in collagen gel induces extensive membrane remodeling and subsequent epithelial polar...

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Section: Discussionsupporting

confidence: 87%

Modulation of epithelial tubule formation by Rho kinase

Eisen

Ratcliffe

Ojakian

2004

American Journal of Physiology-Cell Physiology

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“…The prevailing idea is that Ins(1,4,5)P 3 R-operated Ca 2+ release leads to phosphorylation of the myosin light chain (MLC) by the Ca 2+ /calmodulin-dependent MLC kinase (MLCK), thereby increasing myosin II ATPase activity and filament formation. In MDCK cells, MLCK activation induces purse-string contraction of the perijunctional actomyosin belt (Hecht et al, 1996), but does not seem to be essential for central stress-fiber assembly and dynamics (Sutton et al, 2001). Thus, as previously shown for cultured human fibroblasts (Totsukawa et al, 2000), MLCK activity is probably subjected to spatial regulation in polarized MDCK cells, and compartimentalization of Ins(1,4,5)P 3 Rs in the vicinity of tight junctions, through its interaction with myosin IIA, could contribute to this process.…”

Section: Discussionmentioning

confidence: 99%

The N-terminal domain of the type 1 Ins(1,4,5)P3 receptor stably expressed in MDCK cells interacts with myosin IIA and alters epithelial cell morphology

Hours

Méry

2010

Journal of Cell Science

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Summary Cytosolic Ca2+ controls a wide range of cellular events. The versatility of this second messenger depends on its ability to form diverse spatial and temporal patterns, including waves and oscillations. Ca 2+ -signaling patterns are thought to be determined in part by the subcellular distribution of inositol (1,4,5)-trisphosphate receptors [Ins(1,4,5)P 3 Rs] but little is currently known about how the localization of the Ins(1,4,5)P 3 R itself is regulated. Here, we report that the recruitment of GFP-tagged Ins(1,4,5)P 3 Rs in the vicinity of tight junctions in Madin-Darby canine kidney (MDCK) cells requires the N-terminal domain. Stable expression of this domain in polarized MDCK cells induced a flattened morphology, affected cytokinesis, accelerated cell migration in response to monolayer wounding and interfered with the cortical targeting of myosin IIA. In addition, downregulation of myosin IIA in polarized MDCK cells was found to mimic the effects of stable expression of the N-terminal part of Ins(1,4,5)P 3 R on cell shape and to alter localization of endogenous Ins(1,4,5)P 3 Rs. Taken together, these results support a model in which the recruitment of Ins(1,4,5)P 3 Rs at the apex of the lateral membrane in polarized MDCK cells, involves myosin IIA and might be important for the regulation of cortical actin dynamics.

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“…Actin cytoskeletal disruption is a hallmark of the cell response to hypoxia (25,49). Regulation of actin organization is mediated by members of the Rho GTPase family (14,50).…”

mentioning

confidence: 99%