Identification of a mechanochemical checkpoint and negative feedback loop regulating branching morphogenesis.
Cleft formation is the initial step in submandibular salivary gland (SMG) branching morphogenesis, and may result from localized actomyosin-mediated cellular contraction. Since ROCK regulates cytoskeletal contraction, we investigated the effects of ROCK inhibition on mouse SMG ex vivo organ cultures. Pharmacological inhibitors of ROCK, isoform-specific ROCK I but not ROCK II siRNAs, as well as inhibitors of myosin II activity stalled clefts at initiation. This finding implies the existence of a mechanochemical checkpoint regulating the transition of initiated clefts into progression-competent clefts. Downstream of the checkpoint, clefts are rendered competent through localized assembly of fibronectin promoted by ROCK I/myosin II. Cleft progression is primarily mediated by ROCK I/myosin II-stimulated cell proliferation with a contribution from cellular contraction. Furthermore, we demonstrate that FN assembly itself promotes epithelial proliferation and cleft progression in a ROCK-dependent manner. ROCK also stimulates a proliferation-independent negative feedback loop to prevent further cleft initiations. These results reveal that cleft initiation and progression are two physically and biochemically distinct processes.
Graduate program in Molecular, Cellular, Developmental, and Neural Biology, Department of Biological Sciences, University at Albany, State University of New York, Albany, NY 12222, USA.
William P
WP
Daley
Kathryn M
KM
Gulfo
Sharon J
SJ
Sequeira
Melinda
M
Larsen
D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D002460:Cell Line; D049109:Cell Proliferation; D007150:Immunohistochemistry; D051379:Mice; D009024:Morphogenesis; D034741:RNA, Small Interfering; D013363:Submandibular Gland / Q000196:embryology*; D054460:rho-Associated Kinases / Q000235:genetics / Q000378:metabolism
D034741:RNA, Small Interfering; D054460:rho-Associated Kinases