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E. E. Capowski, J. S. Meyer, R. L. Shearer, K. Wallace, L. S. Wright, D. M. Gamm; MITF Isoform Expression in Developing Human Retinal Cell Populations. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4007.
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© ARVO (1962-2015); The Authors (2016-present)
To assess the isoform expression of the basic helix loop helix leucine zipper transcription factor microphthalmia transcription factor (MITF) in prenatal human retinal tissues and differentiating human embryonic stem cells (hESC).
The expression of specific isoforms of the human MITF gene was determined by RT-PCR using isoform specific primers and by transient transfection of reporter constructs. These reporters contained different MITF promoter sequences controlling the expression of fluorescent marker genes. Tissue sources studied included dissected human fetal eyes, primary RPE cultures, fetal retinal progenitor cell (hRPC) cultures and human embryonic stem cells induced to differentiate along a retinal lineage.
MITF has a complex genomic structure consisting of multiple isoforms expressed from several different promoters. These isoforms are differentially expressed during rodent eye development (1). Human MITF-A was ubiquitously expressed in all eye tissues examined, like its murine counterpart. Human MITF-B was also expressed in eye tissue in contrast to mouse Mitf-B, which was not detected in adult or developing eye. Human MITF-H and MITF-D expression paralleled that seen in mouse with both present at the optic vesicle stage in differentiating hESC as well as in RPE cultures and RPE dissected from fetal eyes at 85 days of gestation. In addition, the human H isoform was expressed in adult neural retina and neural retina dissected from 85d fetal eyes while the human D isoform was not detected in neural retinal tissue. The human M isoform, which is melanocyte-specific in mouse, was undetectable in RPE, optic vesicle stage hESC or neural retina dissected from fetal eyes. It was expressed in combined RPE/choroid tissue dissected from 92d fetal eyes. Finally, the C isoform, which has not been detected in the developing murine eye, was expressed in hRPC cultures but not in human fetal RPE.
Multiple MITF promoters show distinct expression patterns in prenatal human retinal tissues and differentiating hESCs. These results provide a foundation for studying the complex roles of MITF isoforms during human retinal development.1) Bharti et al (2008) Development 135: 1169
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