Control of mRNA Translation During Spermatogenesis

University Of Massachusetts Boston, Dorchester MA

Investigators

Abstract

Spermatogenesis, the development of sperm, is unusual because the patterns of gene regulation differ greatly from those in other cell types reflecting atypical evolutionary pressures on male germ cells. A striking example of the atypical patterns of gene expression in developing sperm cells is the prominence of translational control, a set of mechanisms which regulate translation of the sequence of bases in messenger RNAs into proteins. This research is designed to identify factors, proteins or RNAs, that bind to the spermatogenic cell-specific Smcp mRNA and regulate its translation. The sperm mitochondria-associated cysteine-rich protein (SMCP) functions in sperm motility and is a constituent of a keratinous structure in the outer membranes of sperm mitochondria. The Smcp mRNA undergoes dramatic developmental changes in translational activity in spermatogenic cells. Analysis in transgenic mice using the green fluorescent protein (GFP) reveals that translation of the Smcp mRNA is regulated by both UTRs and several mechanisms including repression in free-mRNPs, reductions in polysome size, post-initiation repression and positive control. The research will use three approaches to investigate the mechanisms of translational regulation of the Smcp mRNA. (1) Several new Smcp-Gfp transgenes will be studied. In one of these transgenes, a long conserved sequence in the Smcp 3'UTR will be randomized, and another transgene will contain both the Smcp 5' and 3' UTRs to determine whether interactions between the UTRs prolong repression and/or mediate positive control. Subsequent transgenes will be designed to identify mRNA-specific regulatory factors. (2) Non-isotopic in situ hybridization will be used to determine whether the various translational controls of the Smcp and Gfp-Smcp mRNAs are correlated with localization in the chromatoid body, a spermatogenic cell-specific RNA granule. (3) Factors that bind to regulatory elements in the Smcp 5'UTR or 3'UTR will be detected with RNA-EMSAs designed to detect complexes that contain microRNAs and/or proteins. The UMass Boston student body includes a high proportion of minorities, students for whom English is their second or third language, and students who are the first in their family to attend college. The project provides an opportunity to mentor minorities and introduce talented undergraduates to research in molecular biology.