Endometriosis is a painful, chronic condition caused by endometrial tissues growing outside the uterus. Adenomyosis occurs when endometrial glands and stroma invade the myometrium — the smooth muscle layer of the uterus responsible for uterine contractions — causing the uterine wall to thicken. Uterine fibroids are benign monoclonal neoplasms of the myometrium and represent the most common tumors in women worldwide. These reproductive disorders are associated with chronic pain, infertility, and affect a disproportionate number of women: endometriosis alone impacts over 190 million globally. Despite their prevalence, these conditions remain under-diagnosed and underfunded, with a historic reliance on invasive procedures and histopathologic examination for diagnosis. The Coller Lab has previously investigated the methyltransferase SUV4-20H2, which generates the histone mark H4K20me3 associated with cellular quiescence. Knockdown of SUV4-20H2 reduces H4K20me3, resulting in more open chromatin and increases the fraction of proliferating cells. Notably, SUV4-20H2 knockout models exhibit interesting gonadal phenotypes, suggesting a role in reproductive structure development and fertility. As quiescence and cell cycle regulation are essential in germ cells and hormone-responsive tissues, we hypothesize that SUV4-20H2 is essential for fertility. Our preliminary data suggests that Suv4-20h2 has a significant role in the estrous cycle, uterine function, and development of the myometrial tissue.

The pattern of epigenetic modifications at the histone and DNA level critically dictates chromatin availability and transcriptional activity. While the effects of histone modifications and DNA methylation have been characterized separately, emerging evidence suggests they mechanistically crosstalk to coordinate transcriptional control. Recently, the Jacobsen Lab identified that the activating histone H3 lysine 4 trimethylation (H3K4me3) mark is associated with DNA methylation removal in Arabidopsis thaliana. Targeting the H3K4me3 methyltransferase SET DOMAIN GROUP 2 (SDG2) with an artificial zinc finger (ZF) potently erased DNA methylation and activated reporter gene expression. Nevertheless, the molecular intermediates linking H3K4me3 to DNA demethylation remain elusive. In this study, using ChIP-Seq, we found that Arabidopsis DNA demethylase REPRESSOR OF SILENCING 1 (ROS1) was enriched at the ZF-binding sites in the SDG2-ZF lines, suggesting ROS1 is recruited downstream of H3K4me3 to mediate DNA demethylation. IP-MS revealed further interaction partners that hinted at their role in this pathway. These protein-protein interactions were validated through Yeast Two-Hybrid and Co-IP, pinpointing key amino acid residues required for the interaction. Together, these findings unveil the intermediates linking SDG2-mediated H3K4me3 with DNA demethylation for transcriptional activation. On a broader scale, a mechanistic understanding of their epigenetic crosstalk can inform the design of more efficient epigenome engineering tools.

Electronic cigarettes (E-cigs) are marketed as a safer alternative to traditional cigarettes, but the effect of inhalation of their liquid components such as vegetable glycerin (VG), propylene glycol (PG), nicotine, and different additives on the airway is not well understood. This study investigates the effects of e-cig exposure on primary human airway epithelial cells grown in air-liquid interface (ALI). Here, we specifically focus on the lung’s innate defense mechanism, mucociliary clearance, in which secreted mucus traps inhaled pathogens and particles, which is then cleared by unidirectional beating of cilia on ciliated cells. We developed a novel exposure system to expose ALI cultures to e-cig smoke. By examining the effects of carrier compounds that are common to all e-cigs (unflavored e-cigs), we found that ALI airway cultures exposed to PG, VG, and 5% nicotine unflavored e-cig liquid show a reduction in cilia beat frequency. Moreover, using transmission electron microscopy, we identified defects in the ciliary ultrastructure induced by unflavored e-cigs. When examining induced molecular changes, we uncovered changes in phosphorylation of proteins involved in cadherin and actin binding and the rho GTPase signaling pathway, which are all involved in cytoskeletal remodeling that influences cilia. Altogether, our findings suggest that exposure to all e-cigarette types impairs mucociliary clearance, likely through disruption of cytoskeletal organization that comprises ciliary ultrastructure and function.

Bacterial pneumonia is a deadly lung infection that is characterized by alveolar inflammation. Current therapies are antibiotics and oxygen supplementation, but no molecular targets exist that improve patient outcomes. We recently showed that activation of BK K+ channels protects lung endothelial cells from lipopolysaccharide (LPS)-induced inflammation but whether such anti-inflammatory effects can be translated into a live animal pneumonia model remains unknown. Therefore, we designed a mouse model of LPS-induced bacterial pneumonia and hypothesized that pharmacological BK channel activation with NS1619 protect mouse lungs from inflammatory cell infiltration and mediator secretion. Mice in the treatment arm received two i.t. injections of NS1619, one at the time of infection and one 24 hrs later. Lung injury endpoints were measured 48 hrs post-infection and showed protective and anti-inflammatory effects of NS1619, including a significantly reduced total cell and neutrophil count in bronchoalveolar lavage fluid (BALF), and decreased CCL2 levels, a key neutrophil chemoattractant. We also found a reduction in oxidative stress as evidenced by lower reactive oxygen species levels, including hydrogen peroxide, and increased superoxide dismutase (SOD) and catalase levels. Other markers of neutrophil activation such as myeloperoxidase (MPO) and neutrophil elastase were not affected by NS1619. These findings suggests that BK channel activation may represent a promising novel therapeutic approach against bacterial pneumonia.

Strongylocentrotus purpuratus has many poorly-defined genes that are fundamentally important to the functionality of multiple developmental processes. The determination of the identity of these genes, as well as their functionality and the ability to clone them, paves the way for testing of these genes in Strongylocentrotus purpuratus and potential applications in understanding the functionality of these genes in humans. The focus of this project was to determine the identity of an unknown Strongylocentrotus purpuratus gene utilizing comparative analysis with existing nucleotide database entries, clone it utilizing a plasmid vector, and to confirm the identity of the clones. Through BLAST, InterPro, and phylogenetic analysis, the unknown gene was identified as estrogen-related receptor gamma (esrrg), a gene involved in the regulation of transcription and linked to cancer. Searching for the gene in EchinoBase confirmed the identity of this gene. Plasmid vectors were then ligated with esrrg to utilize for transformation of E. coli cells. Through colony PCR and subsequent gel electrophoresis, transformed bacteria were confirmed to contain intact copies of esrrg. Future experiments could focus on determination of the functionality of esrrg utilizing knockdown and gain-of-function experiments in different tissues.