Biological processes rely on the precise spatiotemporal regulation of gene expression. It is known that at the level of transcription, gene regulation is controlled by complex physicochemical interactions that occur between: transcription factors, the mediator complex, RNA polymerase, and cis-regulatory DNA elements such as enhancers and promoters; however the exact molecular mechanisms orchestrating transcriptional activation are not well understood. To gain insight regarding these mechanisms, ChIP-seq datasets were processed using MACS2 and quantitatively analyzed using deepTools' computeMatrix, to explore, genome-wide, the positional occupancy of the mediator complex, Pol II, TAF2, and the master transcription factor, Esrrb. Our results show that: 1) Esrrb preferentially binds to enhancers, instead of promoters; and 2) that the mediator complex tends to colocalize at enhancers bound by Esrrb. These findings challenge the commonly accepted model that mediator is generally docked on gene promoters and instead suggests mediator is delivered to gene promoters from activator-bound enhancers. The knowledge produced from this study is useful in understanding transcriptional dysregulation as an etiology of diseases, especially cancer, and can inform the generation of novel therapies aimed at targeting gene regulation.

Menses-derived Stem Cells (MenSCs) are uterine cells that are released during menstruation. MenSCs are highly proliferative and multipotent, enabling them to differentiate into different cell types within the uterus, and undergo environmental stress during the shredding of the endometrium through loss of vascularization. In order to maintain their proliferative potential during periods of stress, MenSCs may have cell-type specific nutrient signaling pathway responses that could be altered in disease. In this study, we used MenSCs extracted from volunteers who either experience severe menstrual pain or who have no pain to genetically identify, characterize and analyze MenSCs to better understand regeneration of the endometrium and to explore their potential involvement in gynecological disorders, such as endometriosis and dysmenorrhea. OCT-4, NANOG, and SOX-2 were chosen as stem cell markers and VEGFA, IL-6, MMP-2, and MMP-9 were selected as potential biomarkers of capacity for proliferation and involvement in disease. To explore altered signaling in response to nutrient starvation, we used antibodies for activated mTOR and Akt, as well as expression of p53. We found that MenSCs express NANOG, but do not express OCT-4. (SOX-2 had a low level of expression.) VEGFA, IL-6 and MMP-2 were expressed in all cell lines regardless of pain status. We found altered expression of activated mTOR in response to nutrient starvation by pain status. These results indicate that MenSCs have the potential to be involved in dysmenorrhea and their use as a model of uterine diseases may aid in the development of therapeutics for common reproductive disorders.

Recently, it has been discovered that RNA plays a role beyond transmitting genetic information, and can assume complex enzyme-like catalytic properties and regulatory function in cells. The structures of such RNAs, commonly referred to as non-coding RNA, are unknown, which is reflected by the lack of high resolution structures in the Protein Data Bank (PDB). Cryo-EM, or single particle electron microscopy, offers a potential methodology to address this lack of RNA-only structures. This technique has been primarily focused on studying proteins, but has shown promise when applied to larger RNA biomolecules. To study and elucidate the structure of various RNA, we seek to use large symmetric protein scaffolds to rigidly display single molecule target RNA. This approach leverages symmetry to enhance the signal-to-noise of our targets while simultaneously reducing the flexibility intrinsic to RNA biomolecules and increasing their relative size, making high resolution maps obtainable. The discovery of successful protein cage designs may lead to the creation of a visualization tool that allows for a greater appreciation of RNA diversity and advancement of the clinical revolution promised by RNA therapeutics.

A melanoma tumor can alter its environment to increase its growth. Cancer associated fibroblasts (CAF) have the ability to increase the growth of cancers, including melanoma. CAFs provide nutrients and even promote chemotherapy resistance. Melanomas can induce fibroblasts to activate autophagy in the tumor microenvironment (TME) in hopes of improving said environment to ultimately increase its size. Factors in the TME can be altered in Cre recombinase mice. In this study, Ubc.Atg mice had autophagy inactivated systematically and Fsp1.Atg7 in fibroblasts. This, in combination with melanomas injected into mice, led to noteworthy results. Melanoma weight in Ubc mice decreased by an average of 68.9%, and by 70.9% in Fsp1 mice. Using immunohistochemistry, Ubc and Fsp1 mice showed reduced expression of proliferation (KI-67) and blood vessel markers (CD31). In addition, there was an increase in the expression of apoptosis markers (Caspase-3), and tumor infiltrating T-cells (CD8). Moreover, fibroblasts in Fsp1 models showed higher levels of apoptosis and decreased levels in the TME. Furthermore, autophagy inactivation in Fsp1 models led to an increased immune response. The increased levels of tumor infiltrating CD8 T-cells in this immune response could explain how melanoma size decreased in Ubc and Fsp1 models. Since the inactivation of autophagy led to a TME with conditions that were less favorable for tumor growth and were associated with a decrease in melanoma size this could be a pathway to new possible therapies.