-Molecular basis of the malignant, stem cell and memory lymphocyte phenotypes
-Mechanisms of gene regulation by Oct transcription factors
The major long-term goal of our research is to understand the transcriptional underpinnings of immune responses, stem cell function and the malignant state. Sequence-specific DNA binding transcription factors sculpt gene expression patterns by interacting with particular DNA sequences in our genomes. We use biochemical, genetic and genomic approaches to determine mechanisms and functions of these factors.
Control of developmentally inducible gene expression in differentiating embryonic stem cells
The embryonic stem cell master transcription regulator Oct4 is a potent controller of pluripotency. We identified a transcriptional mechanism hat Oct4 uses uniquely at its core pluripotency targets. This mechanism involves the Jmjd1c histone demethylase and FACT histone chaperone. Another category of Oct4 targets are developmental-specific genes that are silent but held in a configuration that allows them to be readily activated, or stably repressed, later in development. We are studying the role of Oct4 paralogs (e.g., Oct1) in this process. These proteins have similar in vitro DNA binding specificity as Oct4 and recognize many of the same target genes. We have found that pluripotent cells with inducible Oct1 knockout behave normally until differentiation, where they fail to induce developmentally appropriate genes, and aberrantly express developmentally inappropriate genes. Upon ESCs differentiation, Oct1 occupies Oct4 target genes, sometimes transiently. These results support a “handoff mode”, whereby activities related to Oct4 take over gene poising and gene silencing duties from Oct4 as cells differentiate and Oct4 is lost. We are determining whether this occurs genome-wide, and identifying the cofactors used in this process.
Upstream control of Oct1 protein stability and cellular metabolism by BRCA1
Oct1is widely expressed, but more highly expressed in somatic and cancer stem cell compartments, where it promotes glycolytic metabolic phenotypes. Interestingly mRNA levels remain unchanged. We have found that Oct1 protein is destabilized by BRCA1/BARD1 via its N-terminal E3 ligase activity. Consistently, we have found that BRCA1 promotes oxidative metabolic profiles. In primary breast cancer specimens, Oct1 protein levels tend to increase with tumor grade while BRCA1 declines. We are studying the biochemical and metabolic circuitry of this process, the effect of BRCA1 mutants and variants in Oct1 stability, and whether Oct1 promotes breast cancer tumorigenicity through metabolic mechanisms.
Gene regulatory basis of CD4 T cell memory
Oct1 controls target gene expression through related mechanisms that enforce either a repressed transcription state, or a silent but "poised" state. Poised gene expression states are indicative not only of stem cells, but also of memory lymphocytes. Interestingly, Oct1 controls CD4 T memory lymphocyte formation and activity. Oct1 is widely expressed but one of its cofactors, OCA-B, is induced in CD4 T cells upon activation. We found that T cell-specific OCA-B loss confers the same phenotype. We are studying whether OCA-B expression can be used to prospectively identify CD4 memory cells during ongoing immune responses, the ability of membrane-soluble peptide inhibitors to block this process, the role of these activities in CD8 T cells, and the ability of these proteins to coordinately regulate distant target genes by spatially localizing them together.
Contribution of Oct1 to tissue regeneration and malignancy
Oct protein is elevated in multiple tumor types. It promotes oncogenic transformation and its loss protects mice in several tumor models. In normal cells Oct1 is a somatic and cancer stem cell determinant. It promotes these functions through multiple pathways including control of cellular metabolism. We have found that Oct1 is dispensable for reconstitution of the blood system by hematopoietic stem cells, but is required in the case of serial or competitive transplants. In the gut, we have found that Oct1 is dispensable for normal homeostasis but required for regeneration. We are working to understand the molecular basis of these phenomena, and determining the role of Oct1 and cofactors in mouse models of leukemia and colon cancer.