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.
Daley
William P
WP
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.
Gulfo
Kathryn M
KM
Sequeira
Sharon J
SJ
Larsen
Melinda
M
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