The long-term goal of the proposed work is to gain a better understanding of how gene expression is regulated during mammalian cell development. This proposal focuses on the study of post-transcriptional control of gene expression, whereby mRNAs made from a single gene can be modified in different ways to change gene output. In the nucleus, this includes altering the protein-coding and non-coding sequences of mRNA. In the cytoplasm, this includes regulatory events that dictate which mRNAs are preferentially degraded or translated to make proteins. Together, these regulatory processes specify the identity and abundance of proteins present in each cell, and consequently, cell properties. In order for cells to progress through different stages of development, its mRNAs undergo extensive stage-specific regulation. Defects in mRNA regulation are the direct cause of many human diseases, thus an understanding of how mRNAs are regulated is essential. In the proposed study, we will comprehensively characterize mRNA regulatory events that drive cells through different stages of male germ cell development. This cellular program depends on undefined mRNA regulatory programs. Our plan is to isolate postnatal mouse germ cells at different steps in this pathway and use transcriptome-wide tools to 1) identify changes in mRNA expression and translation, 2) understand the functional significance of these changes, and 3) determine how these changes are controlled at the molecular level. We will use an approach established in our laboratory that combines dual fluorescence cell labeling in transgenic mice and fluorescence activated cell sorting to isolate cells at different stages of development. Combining this approach with deep sequencing, biochemical, and bioinformatic tools, we will reveal multiple layers of mRNA-based gene control during germ cell development. Altogether, we will gain important insights into the functions and mechanisms of mRNA regulation in mammalian cell development in unprecedented molecular and cellular detail. As a result, this study will lead to an improved understanding of how gene expression is controlled through mRNA regulation during germ cell development. The data has the potential to provide important new insights into human reproductive biology and fertility. This study will also reveal molecular mechanisms controlling mammalian cell differentiation and proliferation, and therefore will be of direct relevance to human development and disease.
Key to understanding how different cells develop is an improved understanding of how their genes are regulated. This study will investigate how genes are regulated during the development of male germline stem cells ? which allow sperm production throughout life. These specialized cells and cancer cells have many similar characteristics with respect to their growth and proliferation. The information obtained in this study will improve our understanding of the fundamentals of how genes are controlled, while also providing insights into human reproductive biology and human diseases where cell proliferation programs are perturbed. !
|Guenther, Ulf-Peter; Weinberg, David E; Zubradt, Meghan M et al. (2018) The helicase Ded1p controls use of near-cognate translation initiation codons in 5' UTRs. Nature 559:130-134|
|Zagore, Leah L; Sweet, Thomas J; Hannigan, Molly M et al. (2018) DAZL Regulates Germ Cell Survival through a Network of PolyA-Proximal mRNA Interactions. Cell Rep 25:1225-1240.e6|
|Hannigan, Molly M; Zagore, Leah L; Licatalosi, Donny D (2017) Ptbp2 Controls an Alternative Splicing Network Required for Cell Communication during Spermatogenesis. Cell Rep 19:2598-2612|
|Licatalosi, Donny D (2016) Roles of RNA-binding Proteins and Post-transcriptional Regulation in Driving Male Germ Cell Development in the Mouse. Adv Exp Med Biol 907:123-51|
|Zagore, Leah L; Grabinski, Sarah E; Sweet, Thomas J et al. (2015) RNA Binding Protein Ptbp2 Is Essential for Male Germ Cell Development. Mol Cell Biol 35:4030-42|
|Sweet, Thomas J; Licatalosi, Donny D (2014) 3' end formation and regulation of eukaryotic mRNAs. Methods Mol Biol 1125:3-12|