Trans-Zonal Projections May Permit the Transfer of RNA-Protein Complexes Between the Cumulus Cells and the Oocyte.

Abstract

Oocyte maturation is mediated by communication events with the cumulus cells that promote oocyte quality for maturation, fertilization, and embryo development. Cumulus cells are in direct contact with the oolemma through cytoplasmic processes known as transzonal projections. The projections are anchored by zonula adherens and contain gap junctions. The gap junctions permit the transfer of cyclic nucleotides that mediate oocyte growth and maturation. In insects and amphibians there are mechanisms present for the transfer of larger materials through cellular connections that promote development, however, in mammals; only small molecule signaling has been studied. When an oocyte is fully-grown, it reaches a critical period where it stores RNA in protein bound complexes called ribonucleoparticles prior to becoming transcriptionally inactive. Stored RNA is used for cellular division and function during the period between transcriptional arrest and the transcriptional resumption that occurs after the maternal to embryonic transition. Previous work has shown that an unspecific RNA binding protein, the Fragile-X mental retardation protein, is abundant in both cumulus cells and the oocyte and is known for roles in translation repression and RNA shuttling. The depletion of the Fragile-X mental retardation protein in the ovary is associated with premature ovarian syndrome in humans. From this evidence it is hypothesized that large molecule transfer, in the form of ribonucleoproteins occurs between the oocyte and cumulus cell. Our initial investigation using light, confocal, and transmission electron microscopy in combination with fluorescent staining, autoradiography, and fluorescent RNA staining techniques has revealed two novel concepts. The first is that vesicles are present between the membranes where the ooctyes and transzonal projections articulate. The second finding is that RNA exists in the transzonal projections near to the oolemma. To test the importance of exocytosis on oocyte maturation two endocytosis inhibitors, monodansyl cadaverine and Dynasore, were added to in-vitro maturing oocytes that were pre-selected using brilliant cresyl blue staining. Both inhibitors negatively impacted the rate of ooctyes that extruded polar bodies (64% for monodansyl cadaverine and 61% for Dynasore) when compared to untreated controls (89%). These findings suggest that oocyte maturation is more dependent upon endocytic function that previously thought. The transfer of large molecules like proteins, or RNA bound to proteins, during this critical period may have significant consequences upon the quality of the oocyte. Understanding that the proteome and the transcriptome of the oocyte may be modified by cumulus cell transfer of RNA and proteins could have significant impact upon the way we view oocyte competence and ovarian health, and may permit us to improve in-vitro systems were we mature ooctyes for clinical and experimental applications.