Browsing by Author "Phillips, M. Joseph"

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    Loss of MITF expression during human embryonic stem cell differentiation disrupts retinal pigment epithelium development and optic vesicle cell proliferation
    (Oxford University Press, 2014-12-01) Capowski, Elizabeth E.; Simonett, Joseph M.; Clark, Eric M.; Wright, Lynda S.; Howden, Sara E.; Wallace, Kyle A.; Petelinsek, Anna M.; Pinilla, Isabel; Phillips, M. Joseph; Meyer, Jason S.; Schneider, Bernard L.; Thomson, James A.; Gamm, David M.; Department of Biology, School of Science
    Microphthalmia-associated transcription factor (MITF) is a master regulator of pigmented cell survival and differentiation with direct transcriptional links to cell cycle, apoptosis and pigmentation. In mouse, Mitf is expressed early and uniformly in optic vesicle (OV) cells as they evaginate from the developing neural tube, and null Mitf mutations result in microphthalmia and pigmentation defects. However, homozygous mutations in MITF have not been identified in humans; therefore, little is known about its role in human retinogenesis. We used a human embryonic stem cell (hESC) model that recapitulates numerous aspects of retinal development, including OV specification and formation of retinal pigment epithelium (RPE) and neural retina progenitor cells (NRPCs), to investigate the earliest roles of MITF. During hESC differentiation toward a retinal lineage, a subset of MITF isoforms was expressed in a sequence and tissue distribution similar to that observed in mice. In addition, we found that promoters for the MITF-A, -D and -H isoforms were directly targeted by Visual Systems Homeobox 2 (VSX2), a transcription factor involved in patterning the OV toward a NRPC fate. We then manipulated MITF RNA and protein levels at early developmental stages and observed decreased expression of eye field transcription factors, reduced early OV cell proliferation and disrupted RPE maturation. This work provides a foundation for investigating MITF and other highly complex, multi-purposed transcription factors in a dynamic human developmental model system.
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    Retinal Ganglion Cell Diversity and Subtype Specification from Human Pluripotent Stem Cells
    (Cell Press, 2018-04-10) Langer, Kirstin B.; Ohlemacher, Sarah K.; Phillips, M. Joseph; Fligor, Clarisse M.; Jiang, Peng; Gamm, David M.; Meyer, Jason S.; Biology, School of Science
    Retinal ganglion cells (RGCs) are the projection neurons of the retina and transmit visual information to postsynaptic targets in the brain. While this function is shared among nearly all RGCs, this class of cell is remarkably diverse, comprised of multiple subtypes. Previous efforts have identified numerous RGC subtypes in animal models, but less attention has been paid to human RGCs. Thus, efforts of this study examined the diversity of RGCs differentiated from human pluripotent stem cells (hPSCs) and characterized defined subtypes through the expression of subtype-specific markers. Further investigation of these subtypes was achieved using single-cell transcriptomics, confirming the combinatorial expression of molecular markers associated with these subtypes, and also provided insight into more subtype-specific markers. Thus, the results of this study describe the derivation of RGC subtypes from hPSCs and will support the future exploration of phenotypic and functional diversity within human RGCs.