Preimplantation is a short yet critical time period for the embryo that includes dramatic changes in gene expression and developmental potential. During early cleavage, a combination of maternal and zygotic factors program the embryonic genome. These dynamic events result in critical differentiation steps including the first lineage specification (ICM and TE), which is required for implantation.
The first half of this thesis investigates the role of Smyd3 as a possible maternal-effect gene, which was identified through an mRNA-based expression assay. Using a Smyd3 conditional allele to remove the function of the protein, we were able to confirm successful allelic recombinations but were unable to confirm the absence of the protein. Due to a lack of a phenotype, two conclusions were made: 1) If we did disrupt Smyd3 function, the lack of a phenotype indicates that Smyd3 is not a maternal-effect gene, and is not required for embryonic viability or adult fertility. 2) It is possible that newly-annotated transcripts are responsible for the necessity of Smyd3, and that the conditional allele used did not disrupt the function of Smyd3.
The second half of this thesis uses a reverse genetic RNAi screen that identified Wdr74 as being required for the critical first steps of mammalian development. Knockdown of Wdr74 results in embryos that develop normally until the morula stage but fail to form blastocysts or properly specify the inner cell mass and trophectoderm. In Wdr74-deficient embryos, we find activated Trp53-dependent apoptosis as well as a global reduction of RNA polymerase I, II and III transcripts. In Wdr74-deficient embryos, blocking Trp53 function rescues blastocyst formation and lineage differentiation. These results indicate that Wdr74 is required for RNA transcription, processing and/or stability during preimplantation development and is an essential gene in the mouse.
| Identifer | oai:union.ndltd.org:UMASS/oai:scholarworks.umass.edu:theses-1810 |
| Date | 01 January 2011 |
| Creators | Walentuk, Melanie A |
| Publisher | ScholarWorks@UMass Amherst |
| Source Sets | University of Massachusetts, Amherst |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Masters Theses 1911 - February 2014 |
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