Project Grant F31HD123506

INTEGRATING NOVEL TRANSCRIPTIONAL AND ADHESIVE NETWORKS TO BUILD A FUNCTIONAL GONAD IN C. ELEGANS. - PROJECT SUMMARY: IN SEXUALLY REPRODUCING ORGANISMS, THE GERMLINE MUST BE PRECISELY REGULATED TO ENSURE THE PRODUCTION OF FUNCTIONAL HAPLOID GAMETES. CRITICAL DECISIONS SUCH AS GERM CELL QUIESCENCE, PROLIFERATION, AND DIFFERENTIATION MUST BE SPATIOTEMPORALLY CONTROLLED DURING DEVELOPMENT TO AVOID BOTH INFERTILITY AND GERMLINE TUMORS. THE SOMATIC GONAD IS KNOWN TO PROVIDE THIS LEVEL OF REGULATION BY FORMING INTIMATE CONNECTIONS TO THE GERMLINE. IN MANY ANIMAL MODEL SYSTEMS, THESE CONNECTIONS ARE ESTABLISHED DURING EARLY EMBRYOGENESIS WHEN SOMATIC GONADAL CELLS WRAP THEIR MEMBRANES AROUND PRIMORDIAL GERM CELLS (PGCS) TO FORM THE EMBRYONIC GONADAL NICHE. WHILE A MAJORITY OF GERMLINE BIOLOGY RESEARCH HAS FOCUSED ON THE SIGNALING PATHWAYS SOMATIC CELLS USE TO GUIDE PGC FATE DECISIONS, LITTLE IS KNOWN ABOUT THE MECHANISMS THAT ENSURE THESE TWO CELL POPULATIONS COALESCE. IN THIS PROJECT, I PROPOSE USING CAENORHABDITIS ELEGANS AS A MODEL SYSTEM TO ANSWER NOVEL QUESTIONS REGARDING EARLY GONAD ASSEMBLY. IN MOST MODELS, PGCS MIGRATE TO THE SOMATIC GONAD TO FORM THE GONAD PRIMORDIUM; HOWEVER, IN C. ELEGANS, THIS IS REVERSED. IN C. ELEGANS, TWO SOMATIC GONADAL CELLS MIGRATE AND ENWRAP TWO PGCS. THIS HIGHLIGHTS HOW PGCS DIRECT SOMATIC GONAD DEVELOPMENT AND PROVIDES AN OPPORTUNITY TO UNDERSTAND GERM-CELL-SPECIFIC CUES USED FOR WRAPPING AT A SINGLE-CELL RESOLUTION. TO INITIALLY VALIDATE THAT PGCS PROVIDE SIGNALS TO SOMATIC GONADAL CELLS, WE ASSESSED THE ABILITY FOR THE GONAD NICHE TO FORM IN A "GERM CELL-LESS" ANIMAL. AS EXPECTED, WHEN GERM CELLS ARE NOT SPECIFIED PROPERLY, SOMATIC CELLS FAIL TO ENWRAP ANY CELL TYPE. IN AN INITIAL GENETIC SCREEN, WE IDENTIFIED SELECT ADHESION RECEPTORS INVOLVED IN THE RECOGNITION AND ENWRAPMENT OF PGCS, SUGGESTING THAT ADHESION BETWEEN BOTH CELL TYPES FACILITATES WRAPPING. ADDITIONALLY, IN FURTHER PRELIMINARY STUDIES, WE HAVE OBSERVED THAT POSTERIOR PATTERNING MUTANTS CAUSE SEVERE DEFECTS IN THE ABILITY OF SOMATIC CELLS TO REMAIN WRAPPED AROUND PGCS, SHOWING THAT POSTERIOR CELLS MAY ACTIVATE SPECIFIC GENE REGULATORY NETWORKS THAT REINFORCE WRAPPING. THE PROPOSED WORK WILL BUILD UPON THESE FINDINGS BY UTILIZING A COMBINATION OF CELL-TYPE-SPECIFIC GENETIC MANIPULATIONS, IMAGING TOOLS, AND RNA-SEQ TO FURTHER ELUCIDATE HOW THE GONADAL NICHE IS ASSEMBLED. AIM 1 WILL INVESTIGATE THE ROLE OF DIFFERENTIAL ADHESION AND HOW ADHESION MEDIATES SOMATIC GONADAL CELL CYTOSKELETAL DYNAMICS IN THE INITIATION OF WRAPPING. IN AIM 2, I WILL CHARACTERIZE NOVEL TRANSCRIPTIONAL NETWORKS THAT REINFORCE CELL POLARITY AND WRAPPING AROUND PGCS. THESE EXPERIMENTS WILL PROVIDE INSIGHT INTO THE GENES AND REGULATORY NETWORKS NEEDED FOR NICHE ASSEMBLY, WHICH IS A PROCESS CRITICAL FOR THE DEVELOPMENT OF THE ADULT GONAD AND FERTILITY.