Insulin signaling modulates border cell movement in Drosophila oogenesis.
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ABSTRACT:
SUBMITTER: Sharma A
PROVIDER: S-EPMC6078333 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Intercellular communication mediated by gap junction (GJ) proteins is indispensable during embryogenesis, tissue regeneration and wound healing. Here we report functional analysis of a gap junction protein, Innexin 2 (Inx2), in cell type specification during Drosophila oogenesis. Our data reveal a novel involvement of Inx2 in the specification of Border Cells (BCs), a migratory cell type, whose identity is determined by the cell autonomous STAT activity. We show that Inx2 influences BC fate specification by modulating STAT activity via Domeless receptor endocytosis. Furthermore, detailed experimental analysis has uncovered that Inx2 also regulates a calcium flux that transmits across the follicle cells. We propose that Inx2 mediated calcium flux in the follicle cells stimulates endocytosis by altering Dynamin (Shibire) distribution which is in turn critical for careful calibration of STAT activation and, thus for BC specification. Together our data provide unprecedented molecular insights into how gap junction proteins can regulate cell-type specification.
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Project description:Amino-acid starvation leads to an inhibition of cellular proliferation and the induction of programmed cell death (PCD) in the Drosophila ovary. Disruption of insulin signaling has been shown to inhibit the progression of oogenesis, but it is unclear whether this phenotype mimics starvation. Here, we investigate whether the insulin-mediated phosphoinositide kinase-3 pathway regulates PCD in mid oogenesis. We reasoned that under well-fed conditions, disruption of positive components of the insulin signaling pathway within the germline would mimic starvation and produce degenerating egg chambers. Surprisingly, mutants did not mimic starvation but instead produced many abnormal egg chambers in which the somatic follicle cells disappeared and the germline persisted. These abnormal egg chambers did not show an induction of caspases and lysosomes like that observed in wild-type (WT) degenerating egg chambers. Egg chambers from insulin signaling mutants were resistant to starvation-induced PCD, indicating that a complete block in insulin-signaling prevents the proper response to starvation. However, target of rapamycin (Tor) mutants did show a phenotype that mimicked WT starvation-induced PCD, indicating an insulin independent regulation of PCD via Tor signaling. These results suggest that inhibition of the insulin signaling pathway is not sufficient to regulate starvation-induced PCD in mid oogenesis. Furthermore, starvation-induced PCD is regulated by Tor signaling converging with the canonical insulin signaling pathway.
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Project description:Reproduction is a process that is extremely sensitive to changes in nutritional status. The nutritional control of oogenesis via insulin signaling has been reported; however, the mechanism underlying its sensitivity and tissue specificity has not been elucidated. Here, we determined that Drosophila Makorin RING finger protein 1 gene (Mkrn1) functions in the metabolic regulation of oogenesis. Mkrn1 was endogenously expressed at high levels in ovaries and Mkrn1 knockout resulted in female sterility. Mkrn1-null egg chambers were previtellogenic without egg production. FLP-FRT mosaic analysis revealed that Mkrn1 is essential in germline cells, but not follicle cells, for ovarian function. As well, AKT phosphorylation via insulin signaling was greatly reduced in the germline cells, but not the follicle cells, of the mutant clones in the ovaries. Furthermore, protein-rich diet elevated Mkrn1 protein levels, without increased mRNA levels. The p-AKT and p-S6K levels, downstream targets of insulin/Tor signaling, were significantly increased by a nutrient-rich diet in wild-type ovaries whereas those were low in Mkrn1exS compared to wild-type ovaries. Taken together, our results suggest that nutrient availability upregulates the Mkrn1 protein, which acts as a positive regulator of insulin signaling to confer sensitivity and tissue specificity in the ovaries for proper oogenesis based on nutritional status.
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Project description:BackgroundCalpains are calcium regulated intracellular cysteine proteases implicated in a variety of physiological functions and pathological conditions. The Drosophila melanogaster genome contains only two genes, CalpA and CalpB coding for canonical, active calpain enzymes. The movement of the border cells in Drosophila egg chambers is a well characterized model of the eukaryotic cell migration. Using this genetically pliable model we can investigate the physiological role of calpains in cell motility.ResultsWe demonstrate at the whole organism level that CalpB is implicated in cell migration, while the structurally related CalpA paralog can not fulfill the same function. The downregulation of the CalpB gene by mutations or RNA interference results in a delayed migration of the border cells in Drosophila egg chambers. This phenotype is significantly enhanced when the focal adhesion complex genes encoding for α-PS2 integrin ( if), β-PS integrin (mys) and talin (rhea) are silenced. The reduction of CalpB activity diminishes the release of integrins from the rear end of the border cells. The delayed migration and the reduced integrin release phenotypes can be suppressed by expressing wild-type talin-head in the border cells but not talin-head(R367A), a mutant form which is not able to bind β-PS integrin. CalpB can cleave talin in vitro, and the two proteins coimmunoprecipitate from Drosophila extracts.ConclusionsThe physiological function of CalpB in border cell motility has been demonstrated in vivo. The genetic interaction between the CalpB and the if, mys, as well as rhea genes, the involvement of active talin head-domains in the process, and the fact that CalpB and talin interact with each other collectively suggest that the limited proteolytic cleavage of talin is one of the possible mechanisms through which CalpB regulates cell migration.
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Project description:Protein components of the invertebrate occluding junction-known as the septate junction (SJ)-are required for morphogenetic developmental events during embryogenesis in Drosophila melanogaster. In order to determine whether SJ proteins are similarly required for morphogenesis during other developmental stages, we investigated the localization and requirement of four representative SJ proteins during oogenesis: Contactin, Macroglobulin complement-related, Neurexin IV, and Coracle. A number of morphogenetic processes occur during oogenesis, including egg elongation, formation of dorsal appendages, and border cell (BC) migration. We found that all four SJ proteins are expressed in egg chambers throughout oogenesis, with the highest and the most sustained levels in the follicular epithelium (FE). In the FE, SJ proteins localize along the lateral membrane during early and mid-oogenesis, but become enriched in an apical-lateral domain (the presumptive SJ) by stage 11. SJ protein relocalization requires the expression of other SJ proteins, as well as Rab5 and Rab11 like SJ biogenesis in the embryo. Knocking down the expression of these SJ proteins in follicle cells throughout oogenesis results in egg elongation defects and abnormal dorsal appendages. Similarly, reducing the expression of SJ genes in the BC cluster results in BC migration defects. Together, these results demonstrate an essential requirement for SJ genes in morphogenesis during oogenesis, and suggest that SJ proteins may have conserved functions in epithelial morphogenesis across developmental stages.
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Project description:The bone morphogenetic protein (BMP) signaling pathway is a conserved regulator of cellular and developmental processes in animals. The mechanisms underlying BMP signaling activation differ among tissues and mostly reflect changes in the expression of pathway components. BMP signaling is one of the major pathways responsible for the patterning of the Drosophila eggshell, a complex structure derived from a layer of follicle cells (FCs) surrounding the developing oocyte. Activation of BMP signaling in the FCs is dynamic. Initially, signaling is along the anterior-posterior (A/P) axis; later, signaling acquires dorsal-ventral (D/V) polarity. These dynamics are regulated by changes in the expression pattern of the type I BMP receptor thickveins (tkv). We recently found that signaling dynamics and TKV patterning are highly correlated in the FCs of multiple Drosophila species. In addition, we showed that signaling patterns are spatially different among species. Here, we use a mathematical model to simulate the dynamics and differences of BMP signaling in numerous species. This model predicts that qualitative and quantitative changes in receptor expression can lead to differences in the spatial pattern of BMP signaling. We tested these predications experimentally in three different Drosophila species and through genetic perturbations of BMP signaling in D. melanogaster. On the basis of our results, we concluded that changes in tkv patterning can account for the experimentally observed differences in the patterns of BMP signaling in multiple Drosophila species.
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Project description:Collective cell migration is a complex process that happens during normal development of many multicellular organisms, as well as during oncological transformations. In Drosophila oogenesis, a small set of follicle cells originally located at the anterior tip of each egg chamber become motile and migrate as a cluster through nurse cells toward the oocyte. These specialized cells are referred to as border cells (BCs) and provide a simple and convenient model system to study collective cell migration. The process is known to be complexly regulated at different levels and the product of the slow border cells (slbo) gene, the C/EBP transcription factor, is one of the key elements in this process. However, little is known about the regulation of slbo expression. On the other hand, the ubiquitously expressed transcription factor GAGA, which is encoded by the Trithorax-like (Trl) gene was previously demonstrated to be important for Drosophila oogenesis. Here, we found that Trl mutations cause substantial defects in BC migration. Partially, these defects are explained by the reduced level of slbo expression in BCs. Additionally, a strong genetic interaction between Trl and slbo mutants, along with the presence of putative GAGA binding sites within the slbo promoter and enhancer, suggests the direct regulation of this gene by GAGA. This idea is supported by the reduction in the slbo-Gal4-driven GFP expression within BC clusters in Trl mutant background. However, the inability of slbo overexpression to compensate defects in BC migration caused by Trl mutations suggests that there are other GAGA target genes contributing to this process. Taken together, the results define GAGA as another important regulator of BC migration in Drosophila oogenesis.
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