Understanding SHRF, an RNA exosome-linked disease with multi-organ involvement

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Project Summary Short stature, hearing loss, retinitis pigmentosa, and distinctive facies (SHRF) is a rare autosomal recessive disorder characterized by short stature, brachydactyly, dysmorphic facial features, hearing loss, and visual impairment. Patients also exhibit mild intellectual disability. SHRF is caused by homozygous or compound heterozygous mutation in the EXOSC2 gene, which encodes a subunit of the RNA exosome complex, a conserved multi-subunit ribonucleolytic complex that controls the 3´ to 5´ processing and degradation of various RNAs in all eukaryotic cells. Nine subunits (EXOSC1-9) form a catalytically inert core that serves as a scaffold for two ribonuclease subunits (EXOSC10 and Dis3). EXOSC2 and EXOSC3 are S1 and KH domain containing RNA-binding proteins that form the exosome cap structure. Intriguingly, despite the sequence similarity and similar positions in the exosome structure occupied by EXOSC2 and EXOSC3, mutations in these subunits result in distinct diseases, with mutations in EXOSC2 causing SHRF and mutations in EXOSC3 causing Pontocerebellar Hypoplasia type 1b (PCH1b), a rare autosomal recessive neonatal/fetal neurodegenerative disease characterized by hypoplasia and atrophy of the cerebellar cortex, dentate nuclei, pontine nuclei and inferior olives. That mutation in core subunits of a seemingly universally required RNA exosome complex can result in distinct diseases reflects inherent complexity in the organization, function, and regulation of this fundamental machinery of post-transcriptional gene regulation. But our understanding of the mechanistic basis underlying these processes is very limited. We hypothesize that RNA exosome subunits are assembled into different subcomplexes with different RNA substrate engagements, and that these subcomplexes may function in a tissue or cell type-specific manner. To test this hypothesis, we propose to employ the recently developed proximity labeling using the engineered enzyme ascorbate peroxidase 2 (APEX2) to systematically identify proteins and RNAs in the immediate proximity of EXOSC2 in mammalian cell culture models, including iPSC-derived neuronal and muscle models, and in vivo Drosophila models. The functional involvement of newly identified factors in exosome biology and SHRF pathogenesis will be tested in Drosophila models. Successful execution of this project will not only lead to new knowledge on the composition, regulation, and tissue-specific requirement of the RNA exosome complex, but also shed light on the pathogenesis of RNA exosome-linked diseases, from SHRF, PCH, SMA and pulmonary fibrosis to cancer, diseases affecting multiple body systems. It is therefore expected that findings from this study will be applicable to the missions of multiple NIH Institutes or Centers (ICs), one of the stated Research Objectives of this R21 funding opportunity.