Poster abstracts

Poster number 92 submitted by Bohye Jeong

Translational regulation of photoreceptor-specific mRNA by Musashi 1 and Musashi 2 proteins for photoreceptor maintenance

Bohye Jeong (Department of Molecular and Biochemistry Medicine, West Virginia University), Fatimah Matalkah (Receptor Biology Laboratory, Toxicology and Molecular Biology Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, CDC, Morgantown, WV), Visvanathan Ramamurthy (Department of Molecular and Biochemistry Medicine, West Virginia University; Department of Ophthalmology, West Virginia University School of Medicine, Center for Neuroscience, West Virginia University), Peter Stoilov (Department of Molecular and Biochemistry Medicine, West Virginia University)

The Musashi (MSI) RNA binding protein family consists of two paralogs, Musashi 1 (MSI1) and Musashi 2 (MSI2) that are highly conserved across species. We and others have shown that these two proteins are essential for the photoreceptor's survival and morphogenesis and maintenance. Double knockout of the Msi1 and Msi2 in developing or mature photoreceptor cells leads to progressive vision loss accompanied with photoreceptor degeneration. MSI1-CLIP seq (crosslinking and immunoprecipitation) in mouse retina showed binding to introns and 3’-UTRs. The transcripts whose 3’-UTRs are bound by MSI1 were enriched for genes involved in phototransduction, suggesting that Musashi is involved in the differentiation and maintenance of photoreceptor cells. Our proteomics analysis of Musashi double knockout retina demonstrates that Musashi predominantly promotes protein expression of its targets. The upregulation of protein expression likely is achieved through stimulation of mRNA translation as it is not accompanied with increase in mRNA levels both in retina and when model Gnat1 transcripts were tested in 3T3 cells. However, MSI binding to the target transcript alone is not sufficient to confer regulation. Therefore, we hypothesize that there are additional factors that cooperate with the MSI proteins to regulate translation. Currently, we are investigating potential interactions with the RNA interference machinery in vivo and in transient transfections that use the rod transducin beta subunit (Gnb1) transcript as a model. Our most recent results using chimeric AGO2-eCLIP show that MSI stabilizes binding of RISC to the target 3’-UTRs. Paradoxically the increased binding of RISC to the 3’-UTRs results in elevated levels of protein expression. Gnb1 is bound by MSI and this binding stabilizes the binding of RISC complexes carrying the photoreceptor-specific miR-183 microRNA. We hope that our ongoing studies on the regulation of model Gnb1 transcripts in 3T3 cells will shed light on the mechanism by which Musashi and RISC cooperate to promote protein expression.

1. Sundar, J., Matalkah, F., Jeong, B., Stoilov, P. & Ramamurthy, V. The Musashi proteins MSI1 and MSI2 are required for photoreceptor morphogenesis and vision in mice. J Biol Chem 296, 100048 (2021).
2. Xiang, L. et al. miR-183/96 plays a pivotal regulatory role in mouse photoreceptor maturation and maintenance. Proc Natl Acad Sci U S A 114, 6376–6381 (2017).
3. Sanuki, R. et al. miR-124a is required for hippocampal axogenesis and retinal cone survival through Lhx2 suppression. Nat Neurosci 14, 1125–1134 (2011).
4. MacNicol, M. C., Cragle, C. E. & MacNicol, A. M. Context-dependent regulation of Musashi-mediated mRNA translation and cell cycle regulation. Cell Cycle Georget. Tex 10, 39–44 (2011).

Keywords: Photoreceptor, Translational regulation, RNA-binding protein