Kellee Siegfried

Kryptolebias marmoratus (Kmar), a teleost fish of the order Cyprinodontiformes, has a suite of unique phenotypes and behaviors not observed in other fishes. Many of these phenotypes are discrete and highly plastic—varying over time within an individual, and in some cases reversible. Kmar and its interfertile sister species, K. hermaphroditus , are the only known self‐fertile vertebrates. This unusual sexual mode has the potential to provide unique insights into the regulation of vertebrate sexual development, and also lends itself to genetics. Kmar is easily adapted to the lab and requires little maintenance. However, its internal fertilization and small clutch size limits its experimental use. To support Kmar as a genetic model, we compared alternative husbandry techniques to maximize recovery of early cleavage‐stage embryos. We find that frequent egg collection enhances yield, and that protease treatment promotes the greatest hatching success. We completed a forward mutagenesis screen and recovered several mutant lines that serve as important tools for genetics in this model. Several will serve as useful viable recessive markers for marking crosses. Importantly, the mutant kissylips lays embryos at twice the rate of wild‐type. Combining frequent egg collection with the kissylips mutant background allows for a substantial enhancement of early embryo yield. These improvements were sufficient to allow experimental analysis of early development and the successful mono‐ and bi‐allelic targeted knockout of an endogenous tyrosinase gene with CRISPR/Cas9 nucleases. Collectively, these tools will facilitate modern developmental genetics in this fascinating fish, leading to future insights into the regulation of plasticity.

Epigenetic reprogramming, mediated by genomic alterations and dysregulation of histone reader and writer proteins, plays a critical role in driving prostate cancer progression and treatment resistance. However, the specific function and regulation of EHMT1 (also known as GLP) and EHMT2 (also known as G9A), well-known histone 3 lysine 9 methyltransferases, in prostate cancer progression remain poorly understood. Through comprehensive investigations, we discovered that both EHMT1 and EHMT2 proteins have the ability to activate oncogenic transcription programs in prostate cancer cells. Silencing EHMT1/2 or targeting their enzymatic activity with small-molecule inhibitors can markedly decrease prostate cancer cell proliferation and metastasis in vitro and in vivo. In-depth analysis of posttranslational modifications of EHMT1 protein revealed the presence of methylation at lysine 450 and 451 residues in multiple prostate cancer models. Notably, we found that lysine 450 can be demethylated by LSD1. Strikingly, concurrent demethylation of both lysine residues resulted in a rapid and profound expansion of EHMT1’s chromatin binding capacity, enabling EHMT1 to reprogram the transcription networks in prostate cancer cells and activate oncogenic signaling pathways. Overall, our studies provide valuable molecular insights into the activity and function of EHMT proteins during prostate cancer progression. Moreover, we propose that the dual-lysine demethylation of EHMT1 acts as a critical molecular switch, triggering the induction of oncogenic transcriptional reprogramming in prostate cancer cells. These findings highlight the potential of targeting EHMT1/2 and their demethylation processes as promising therapeutic strategies for combating prostate cancer progression and overcoming treatment resistance. Significance: In this study, we demonstrate that EHMT1 and EHMT2 proteins drive prostate cancer development by transcriptionally activating multiple oncogenic pathways. Mechanistically, the chromatin binding of EHMT1 is significantly expanded through demethylation of both lysine 450 and 451 residues, which can serve as a critical molecular switch to induce oncogenic transcriptional reprogramming in prostate cancer cells.