Infertility is a growing problem that affects millions of people in Western countries. An important reproductive process that is understudied in internally fertilizing animals is the process by which sperm find the oocyte. Increasing evidence suggest that sperm respond to chemotactic cues within the oviduct to locate recently ovulated oocytes. Prostaglandins (PGs) are fat-derived signaling molecules that are implicated in sperm chemotaxis and other reproductive processes. The TGF? superfamily of peptide ligands is critical for human ovarian follicle development and fertilization, but the underlying mechanisms are not well understood. In the nematode C. elegans, the TGF? ligand DAF-7 couples environmental conditions to female fertility through the modulation of F-series prostaglandin (CePGF) metabolism in the ovary. Oocytes secrete a mixture of CePGF isomers that function as sperm chemoattractants. CePGFs are synthesized by a novel, TGF? dependent mechanism that may be conserved in mammals. The objective of this project is to investigate the mechanism by which DAF-7/TGF? regulates CePGF levels and sperm chemotaxis. Preliminary data demonstrate that the DAF-3 co-SMAD transcription factor represses CePGF synthesis or enhances CePGF breakdown. The central hypothesis is that DAF-3 acts in transcriptionally active oocyte precursors to regulate expression of genes critical for CePGF metabolism. Two independent aims are proposed to test this hypothesis.
AIM 1 will identify cellular and subcellular sites of DAF-3 action to promote sperm guidance. CRISPR/Cas9 technology will be used to determine endogenous DAF-3 expression. Transgenic approaches will be used to identify the key cell types in which DAF-3 function is necessary and sufficient to regulate sperm chemotaxis.
AIM 2 will identify direct or indirect DAF-3 transcriptional targets that are critical for sperm guidance and CePGF metabolism. Preliminary data using RNA sequencing and RNA-mediated interference suggest that multiple DAF-3 target genes act in the germ line to regulate CePGF metabolism. Genetic and mass spectrometry approaches will be used to test this model. The results obtained from this project will provide novel insight into the mechanism(s) by which TGF? signaling regulates female fertility, as well as the function(s) and regulation of a new class of PGs found in C. elegans and mammals.
TGFb is critical for ovarian follicle development and fertilization, but the mechanisms are not well understood. This study seeks to uncover the role of TGFb signaling in the metabolism of a specific class of prostaglandins that are important in guiding C. elegans sperm to the oocytes. Understanding this mechanism may shed light on a novel prostaglandin synthesis pathway and may provide therapeutic avenues for cases of unexplained infertility.
