SummaryGuanylyl cyclases mediate a number of physiological processes, including smooth muscle function and axonal guidance. Here, we report a novel role for Drosophila receptor-type guanylyl cyclase at 76C, Gyc76C, in development of the embryonic somatic muscle. In embryos lacking function of Gyc76C or the downstream cGMP-dependent protein kinase (cGK), DG1, patterning of the somatic body wall muscles was abnormal with ventral and lateral muscle groups showing the most severe defects. In contrast, specification and elongation of the dorsal oblique and dorsal acute muscles of gyc76C mutant embryos was normal, and instead, these muscles showed defects in proper formation of the myotendinous junctions (MTJs). During MTJ formation in gyc76C and pkg21D mutant embryos, the βPS integrin subunit failed to localize to the MTJs and instead was found in discrete puncta within the myotubes. Tissue-specific rescue experiments showed that gyc76C function is required in the muscle for proper patterning and βPS integrin localization at the MTJ. These studies provide the first evidence for a requirement for Gyc76C and DG1 in Drosophila somatic muscle development, and suggest a role in transport and/or retention of integrin receptor subunits at the developing MTJs.

The Drosophila embryonic salivary gland is an epithelial organ formed by the coordinated invagination and migration of primordial cells. To identify genes that regulate gland migration we performed a deficiency screen of the third chromosome. Here, we report on the analysis of the beta 2 tubulin isoform (β2t) that maps at 85D15. We show that in β2t mutant embryos, salivary glands did not complete their posterior migration and that migration of fusion competent myoblasts (FCMs) and longitudinal visceral muscle founder (LVMF) cells between the gland and CVM was delayed. We also demonstrate that gland migration defects correlate with reduced βPS and αPS2 integrin expression in the surrounding mesoderm and that β2t genetically interacts with genes encoding integrin αPS1 and αPS2 subunits. Our studies reveal for the first time that β2t is expressed in embryogenesis and that β2t plays an important role in salivary gland and myoblast migration, possibly through proper regulation of integrin adhesion proteins.

During collective migration of the Drosophila embryonic salivary gland, cells rearrange to form a tube of a distinct shape and size. Here, we report a novel role for the Drosophila KASH (Klarsicht-Anc-Syne Homology) domain protein Klarsicht (Klar) in the regulation of microtubule (MT) stability and integrin receptor localization during salivary gland migration. In wild-type salivary glands, MTs became progressively stabilized as gland migration progressed. In embryos specifically lacking the KASH domain containing isoforms of Klar, salivary gland cells failed to rearrange and migrate, and these defects were accompanied by decreased MT stability and altered integrin receptor localization. In muscles and photoreceptors, KASH isoforms of Klar work together with Klaroid (Koi), a SUN domain protein, to position nuclei; however, loss of Koi had no effect on salivary gland migration, suggesting that Klar controls gland migration through novel interactors. The disrupted cell rearrangement and integrin localization observed in klar mutants could be mimicked by overexpressing Spastin (Spas), a MT severing protein, in otherwise wild-type salivary glands. In turn, promoting MT stability by reducing spas gene dosage in klar mutant embryos rescued the integrin localization, cell rearrangement and gland migration defects. Klar genetically interacts with the Rho1 small GTPase in salivary gland migration and is required for the subcellular localization of Rho1. We also show that Klar binds tubulin directly in vitro. Our studies provide the first evidence that a KASH-domain protein regulates the MT cytoskeleton and integrin localization during collective cell migration.

SummaryThe Drosophila embryonic salivary gland is formed by the invagination and collective migration of cells. Here, we report on a novel developmental role for receptor-type guanylyl cyclase at 76C, Gyc76C, in morphogenesis of the salivary gland. We demonstrate that Gyc76C and downstream cGMP-dependent protein kinase 1 (DG1) function in the gland and surrounding mesoderm to control invagination, collective migration and lumen shape. Loss of gyc76C resulted in glands that failed to invaginate, complete posterior migration and had branched lumens. Salivary gland migration defects of gyc76C mutant embryos were rescued by expression of wild-type gyc76C specifically in the gland or surrounding mesoderm, whereas invagination defects were rescued primarily by expression in the gland. In migrating salivary glands of gyc76C mutant embryos, integrin subunits localized normally to gland–mesoderm contact sites but talin localization in the surrounding circular visceral mesoderm and fat body was altered. The extracellular matrix protein, laminin, also failed to accumulate around the migrating salivary gland of gyc76C mutant embryos, and gyc76C and laminin genetically interacted in gland migration. Our studies suggest that gyc76C controls salivary gland invagination, collective migration and lumen shape, in part by regulating the localization of talin and the laminin matrix.