Exploring novel regulatory mechanisms underlying enhancer activation and cell fate transition

Enhancer malfunction leads to various developmental disorders including thalassemia, Cornelia de Lange syndrome, and Kabuki syndrome. The major enhancer regulator Mll4 belongs to the family of the Su(var)3-9, Enhancer-of-zeste, Trithorax (SET) domain containing histone methyltransferases named COMplex of Proteins ASsociated with Set1 (COMPASS), which places methylation marks at lysine 4 of histone H3 (H3K4). Mll4 is essential for mammalian development and its heterozygous loss-of-function (LOF) mutations lead to various human diseases including Kabuki syndrome and cancer. The principal investigator (PI) recently demonstrated that Mll4 is the major enzyme depositing mono-methylation at H3K4 (H3K4me1), an enhancer-decorating epigenetic mark whose function remains elusive. Moreover, the PI has defined catalytic activity dependent and independent functions of Mll4 in enhancer modulation and stem cell differentiation, and has unveiled an antagonism between enhancer-binding epigenetic machineries in transcriptional control that could underlie the developmental consequences of the heterozygous LOF mutations of MLL4 in human disease. Based on these findings, the PI hypothesizes that Mll4 cooperates with key transcription factors and epigenetic modifiers to modulate enhancer activity, chromatin structure, and transcriptional outputs during stem cell maintenance and differentiation. The studies proposed here aim to elucidate the mechanisms of enhancer regulation to provide insights for developing novel therapies targeting diseases driven by LOF of epigenetic modifiers. Specifically, the PI will 1) determine the role of H3K4me1 regulation on enhancer function and cellular differentiation; 2) delineate how Mll3 and Mll4 regulate the genome architecture in pluripotent stem cells; 3) identify co-factors functioning with Mll4 in regulating gene expression and cell fate transitions. To achieve the career goal of defining the epigenetic mechanisms underlying mammalian development and disease, the PI plans to receive further training from a panel of experienced co-mentors and collaborators in addition to his primary mentor. During the mentored phase of this application, the PI will acquire techniques in biochemistry, proteomics, bioinformatics, stem cell biology, and genome-wide screening. Moreover, the PI will participate in grant writing workshops and career development courses to strengthen skills in writing and leadership, and will attend intramural and extramural scientific activities to extend his scientific network. With the acquired training, the PI will be wellprepared for the task of delineating the machineries and mechanisms in modulating gene expression, chromatin structure, and cell fate determination in the independent R00 phase. In summary, the K99/R00 award, together with the experiences that the PI has garnered as a graduate student and postdoc, the guidance from the mentors and collaborators, and the superb research environment at Northwestern University’s Feinberg School of Medicine will ensure a successful transition for the PI to continue his independent scientific career in the field of stem cell biology and epigenetics.