Date of Award

8-2024

Document Type

Campus Access Thesis

Degree Name

Master of Science (MS)

Department

Biotechnology and Biomedical Science

First Advisor

Labib Rouhana

Second Advisor

Linda Huang

Third Advisor

Kellee Siegfried-Harris

Abstract

Germline development requires molecular mechanisms that are highly conserved across the animal kingdom. Post-transcriptional regulation of mRNA is one of the prevalent features of germline development, and is carried in part by the Cytoplasmic Polyadenylation Element Binding Protein (CPEBs) family. CPEBs bind to specific mRNAs via sequence elements in the 3’- untranslated region (3’UTR) of target mRNAs and regulate their translation. Smed-CPEB2 is a CPEB that is expressed in the testes and central nervous system of the planarian flatworm Schmidtea mediterranea. Smed-CPEB2 is required for spermatogenesis and for overall sexual maturation, at least in part through regulation of neuropeptide y-8 levels. However, the precise mechanisms by which Smed-CPEB2 regulates target mRNAs are still unknown. To gain insight into CPEB2-mediated control of spermatogenesis, 63 potential protein partners were identified through co-immunoprecipitation and mass spectrometry. I hypothesize that some of these candidate partners facilitate Smed-CPEB2 function and are therefore essential for sperm development. Systemic gene-specific RNA interference (RNAi) was induced for each of the 63 putative partners by double-stranded RNA feeding, and the knockdown of eight of these genes resulted in failure to complete spermatogenesis. Knockdown of eight other genes resulted in severe somatic phenotypes, while knockdown of six other genes resulted in mild phenotypes with potential indirect effects in spermatogenesis. Phenotypic analyses using stage-specific markers revealed defects at early stages of spermatogenesis upon CPSF6 RNAi, and defects at late stages for the majority of the candidate partners required for spermatogenesis. None of the genes tested phenocopy CPEB2 RNAi. These results reflect the complex post-transcriptional regulation network that occurs throughout the progression of spermatogenesis.

Comments

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Additional Files

Fanghemei_Zhang_Signatory_Page.docx.pdf (196 kB)

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