Juvenile hormones direct primordial germ cell migration to the embryonic gonad.

Germ cells are essential to sexual reproduction. Across the animal kingdom, extracellular signaling isoprenoids, such as retinoic acids (RAs) in vertebrates and juvenile hormones (JHs) in invertebrates, facilitate multiple processes in reproduction. Here we investigated the role of these potent signaling molecules in embryonic germ cell development, using JHs in Drosophila melanogaster as a model system. In contrast to their established endocrine roles during larval and adult germline development, we found that JH signaling acts locally during embryonic development. Using an in vivo biosensor, we observed active JH signaling first within and near primordial germ cells (PGCs) as they migrate to the developing gonad. Through in vivo and in vitro assays, we determined that JHs are both necessary and sufficient for PGC migration. Analysis into the mechanisms of this newly uncovered paracrine JH function revealed that PGC migration was compromised when JHs were decreased or increased, suggesting that specific titers or spatiotemporal JH dynamics are required for robust PGC colonization of the gonad. Compromised PGC migration can impair fertility and cause germ cell tumors in many species, including humans. In mammals, retinoids have many roles in development and reproduction. We found that like JHs in Drosophila, RA was sufficient to impact mouse PGC migration in vitro. Together, our study reveals a previously unanticipated role of isoprenoids as local effectors of pre-gonadal PGC development and suggests a broadly shared mechanism in PGC migration.

Hedgehog is required for CySC self-renewal but does not contribute to the GSC niche in the Drosophila testis.

The Drosophila testis harbors two types of stem cells: germ line stem cells (GSCs) and cyst stem cells (CySCs). Both stem cell types share a physical niche called the hub, located at the apical tip of the testis. The niche produces the JAK/STAT ligand Unpaired (Upd) and BMPs to maintain CySCs and GSCs, respectively. However, GSCs also require BMPs produced by CySCs, and as such CySCs are part of the niche for GSCs. Here we describe a role for another secreted ligand, Hedgehog (Hh), produced by niche cells, in the self-renewal of CySCs. Hh signaling cell-autonomously regulates CySC number and maintenance. The Hh and JAK/STAT pathways act independently and non-redundantly in CySC self-renewal. Finally, Hh signaling does not contribute to the niche function of CySCs, as Hh-sustained CySCs are unable to maintain GSCs in the absence of Stat92E. Therefore, the extended niche function of CySCs is solely attributable to JAK/STAT pathway function.

Drosophila adducin regulates Dlg phosphorylation and targeting of Dlg to the synapse and epithelial membrane.

Adducin is a cytoskeletal protein having regulatory roles that involve actin filaments, functions that are inhibited by phosphorylation of adducin by protein kinase C. Adducin is hyperphosphorylated in nervous system tissue in patients with the neurodegenerative disease amyotrophic lateral sclerosis, and mice lacking ß-adducin have impaired synaptic plasticity and learning. We have found that Drosophila adducin, encoded by hu-li tai shao (hts), is localized to the post-synaptic larval neuromuscular junction (NMJ) in a complex with the scaffolding protein Discs large (Dlg), a regulator of synaptic plasticity during growth of the NMJ. hts mutant NMJs are underdeveloped, whereas over-expression of Hts promotes Dlg phosphorylation, delocalizes Dlg away from the NMJ, and causes NMJ overgrowth. Dlg is a component of septate junctions at the lateral membrane of epithelial cells, and we show that Hts regulates Dlg localization in the amnioserosa, an embryonic epithelium, and that embryos doubly mutant for hts and dlg exhibit defects in epithelial morphogenesis. The phosphorylation of Dlg by the kinases PAR-1 and CaMKII has been shown to disrupt Dlg targeting to the NMJ and we present evidence that Hts regulates Dlg targeting to the NMJ in muscle and the lateral membrane of epithelial cells by controlling the protein levels of PAR-1 and CaMKII, and consequently the extent of Dlg phosphorylation.

Multicellular rosette formation links planar cell polarity to tissue morphogenesis.

Elongation of the body axis is accompanied by the assembly of a polarized cytoarchitecture that provides the basis for directional cell behavior. We find that planar polarity in the Drosophila embryo is established through a sequential enrichment of actin-myosin cables and adherens junction proteins in complementary surface domains. F-actin accumulation at AP interfaces represents the first break in planar symmetry and occurs independently of proper junctional protein distribution at DV interfaces. Polarized cells engage in a novel program of locally coordinated behavior to generate multicellular rosette structures that form and resolve in a directional fashion. Actin-myosin structures align across multiple cells during rosette formation, and adherens junction proteins assemble in a stepwise fashion during rosette resolution. Patterning genes essential for axis elongation selectively affect the frequency and directionality of rosette formation. We propose that the generation of higher-order rosette structures links local cell interactions to global tissue reorganization during morphogenesis.

Drac1 and Crumbs participate in amnioserosa morphogenesis during dorsal closure in Drosophila.

Dorsal closure of the Drosophila embryo involves morphological changes in two epithelia, the epidermis and the amnioserosa, and is a popular system for studying the regulation of epithelial morphogenesis. We previously implicated the small GTPase Drac1 in the assembly of an actomyosin contractile apparatus, contributing to cell shape change in the epidermis during dorsal closure. We now present evidence that Drac1 and Crumbs, a determinant of epithelial polarity, are involved in setting up an actomyosin contractile apparatus that drives amnioserosa morphogenesis by inducing apical cell constriction. Expression of constitutively active Drac1 causes excessive constriction of amnioserosa cells and contraction of the tissue, whereas expression of dominant-negative Drac1 impairs amnioserosa morphogenesis. These Drac1 transgenes may be acting through their effects on the amnioserosa cytoskeleton, as constitutively active Drac1 causes increased staining for F-actin and myosin, whereas dominant-negative Drac1 reduces F-actin levels. Overexpression of Crumbs causes premature cell constriction in the amnioserosa, and dorsal closure defects are seen in embryos homozygous for hypomorphic crumbs alleles. The ability of constitutively active Drac1 to cause contraction of the amnioserosa is impaired in a crumbs mutant background. We propose that amnioserosa morphogenesis is a useful system for studying the regulation of epithelial morphogenesis by Drac1.