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Jesse Sundar, Peter Stoilov, Visvanathan Ramamurthy; Alternative splicing of mRNA regulated by Musashi is crucial for photoreceptor development and function.. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3422.
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© ARVO (1962-2015); The Authors (2016-present)
The mechanisms that drive the production of photoreceptor-specific protein isoforms and their roles in photoreceptor function are poorly understood. Our previous studies suggest that photoreceptor-specific protein isoforms are needed for development of the outer segment (OS). In addition, our data implicated a role for Musashi, a family of RNA-binding proteins, in promoting the inclusion of photoreceptor-specific exons. Therefore, we hypothesized that the Musashi proteins are required for the morphogenesis and function of photoreceptor cells.
We generated retina-specific knockout mice in which either Musashi-1 (Msi1), Musashi-2 (Msi2), or both genes were ablated. After validating these models by western blot, we analyzed their photoreceptor function by electroretinography (ERG) and their morphology by immunocytochemistry. The splicing of photoreceptor-specific exons in mature transcripts was determined by reverse transcriptase PCR. Statistical analyses were performed using the two-tailed student’s t-test.
Photoreceptor function was absent in the Msi1/Msi2 double knockout mice at postnatal day 16, which was the earliest time point that we tested (n=3) (Fig. 1). The loss of function correlated with disrupted photoreceptor OS development in double knockout mice (n=3). Lastly, the inclusion of 13 out of 14 photoreceptor specific exons that we tested was blocked in the double knockout mice. In contrast to the double knockout, photoreceptor function and morphology was mostly preserved at postnatal day 16 in the single Msi1 or Msi2 knockouts. The Msi1 but not Msi2 single knockout had a moderate effect on exon inclusion for some of the alternative exons tested.
Our results show that the Musashi proteins are required for normal photoreceptor development and likely control the photoreceptor-specific splicing program. We also observed significant functional redundancy of the two Musashi genes in photoreceptor cells.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
Figure 1: Scotopic and photopic ERGs showing absence of the light-dependent photoreceptor response in the Msi1/Msi2 double knockout mouse compared to littermate control.
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