The newly organized ASBMB Student Chapter at Emory University aims to provide networking and career‐development opportunities coupled with access to research and science outreach. To broaden our ...impact, we have established a collaboration with the graduate student led organization, Emory Science Advocacy Network (EScAN). EScAN provides a forum to discuss current issues facing the biomedical field and determine how to support robust federal funding of biomedical research. The two organizations together are preparing for the annual EScAN‐sponsored Letter Writing Campaign. We aim to achieve record‐high participation this year of both undergraduate and graduate students from diverse educational and demographic backgrounds. Increases in political awareness and advocacy such as these will enhance the potential for productive legislative action. We will also participate in local (Atlanta) and national (Washington, D.C.) Hill Day in September 2019. In preparation for these events, we have organized focused workshops, including “Communicating Science to Non‐Scientists,” to ensure an effective interface between congress people and young scientists. Aligned with the ASBMB Student Chapter mission statement, our efforts will provide networking and career‐development opportunities for undergraduate students. This collaboration is supported and facilitated by a volunteer graduate student advisor, serving as a liaison between EScAN and the ASBMB Student Chapter. We will assess the outcomes of this collaboration by tracking the number of undergraduate students that participate in the Letter Writing Campaign as an initial metric of engagement. We also are working together with a sociologist to develop pre‐ and post‐surveys to assess the level of understanding of funding for biomedical science and commitment to advocacy for the ASBMB Student Chapter undergraduates. Ultimately, we seek to build a synergistic network of undergraduate and graduate students who are knowledgeable and engaged in scientific outreach and advocacy.
Support or Funding Information
R01 GM130147 01, R25 GM125598 01, R01 MH107305 04
This is from the Experimental Biology 2019 Meeting. There is no full text article associated with this published in The FASEB Journal.
The RNA Exosome and Genetic Disease Corbett, Anita H.; Leung, Sara W.; Sterrett, Maria C. ...
The FASEB journal,
April 2020, 2020-04-00, Letnik:
34, Številka:
S1
Journal Article
Recenzirano
Odprti dostop
The RNA exosome is an evolutionarily conserved, riboexonuclease complex that processes/degrades numerous classes of non‐coding RNA. The 10‐subunit core exosome forms an ring‐like structure composed ...of an upper ring of three S1/KH cap subunits (yeast Rrp4/40/Csl4‐human EXOSC2/3/1), a lower ring of six PH‐like subunits (yeast Rrp41/42/43/45/46/Mtr3‐human EXOSC4/7/8/9/5/6), and a 3′‐5′ riboexonuclease subunit, Rrp44/DIS3, at the base. The nuclear exosome contains an eleventh, cap‐associated riboexonuclease subunit, Rrp6/EXOSC10. RNAs are targeted directly to Rrp6 or threaded through the central channel of the exosome to Rrp44 for processing/degradation. Recently, mutations in four exosome subunit genes, EXOSC2/RRP4, EXOSC3/RRP40, EXOSC8/RRP43, and EXOSC9/RRP45 have been linked human diseases. Mutations in EXOSC2 cause a novel syndrome characterized by retinitis pigmentosa, hearing loss, and mild intellectual disability. In contrast, mutations in EXOSC3 and EXOSC8 cause pontocerebellar hypoplasia type 1b and 1c, respectively ‐ autosomal recessive diseases characterized by cerebellar hypoplasia and neuronal degeneration with early mortality ‐ and mutations in EXOSC9 cause cerebellar atrophy. We also recently collaborated to identify novel mutations in the EXOSC5 gene. To gain insight into the functional consequences of the mutations in these genes encoding structural subunits of the RNA exosome that have been identified in patients, we generated the corresponding mutations in the S. cerevisiae genes and examined their function in budding yeast. We find that rrp variants cause differential effects on cell growth and RNA exosome function, which could shed light on why impairments in different exosome subunits cause distinct and tissue specific phenotypes. We have complemented these functional studies with RNA‐Seq analysis to identify specific RNA targets that are differentially affected by the changes in RNA exosome subunits that model human disease as well as a high copy suppressor screen. To extend our functional studies of EXOSC2 and EXOSC3 mutations to mammalian cells, we have performed a proteomic analysis of variant and wildtype EXOSC2 and EXOSC3 proteins expressed in neuronal cells. These data support a model where differential interactions with RNA exosome cofactors could underlie disease‐specific phenotypes.
Support or Funding Information
NIH grant 5 R01 GM130147‐02 to van Hoof and Corbett
The RNA exosome complex is a key component of RNA processing and quality control that both degrades and processes many classes of RNA. This complex is highly conserved among eukaryotes and was first ...identified and studied in budding yeast (S. cerevisiae). Mutations in the human EXOSC2 gene, which encodes a cap subunit of the RNA exosome, have been linked to a novel syndrome characterized by retinitis pigmentosa, progressive hearing loss, premature aging, short stature, mild intellectual disability and distinctive gestalt. While the amino acid substitutions in EXOSC2 that cause this syndrome are known, how these amino acid changes impact RNA exosome function is not. The goal of my project is to analyze the functional consequences of retinitis pigmentosa‐linked amino acid substitutions modeled in the budding yeast ortholog of EXOSC2, Rrp4.
The two variants I have analyzed, rrp4‐G58V and rrp4‐G226D, correspond to patient mutations G30V and G198D, respectively. I first assessed growth of the mutant strains compared to wildtype yeast cells, which revealed that rrp4‐G226D mutant cells exhibit a growth defect at 37°C, whereas the rrp4‐G58V mutant cells grow normally. To assess whether these amino acid substitutions affect Rrp4 protein levels, I used immunoblotting. Results of this analysis reveal that the rrp4‐G58V and rrp4‐G226D proteins are expressed, but at somewhat reduced level compared to wildtype Rrp4. In the future, I will continue characterization of the mutants by using biochemical approaches to study the assembly of the RNA exosome complex and genetic analysis of rrp4 mutant interactions with RNA exosome cofactors.
Support or Funding Information
EMORY INITIATIVE TO MAXIMIZE STUDENT DEVELOPMENT
Emory Initiative to Maximize Student Development: R25 GM125598
Neurodevelopmental Role of an RNA Binding Protein Required for Cognitive Function: R01 MH107305
This is from the Experimental Biology 2019 Meeting. There is no full text article associated with this published in The FASEB Journal.
RNA exosomopathies, a growing family of diseases, are linked to missense mutations in genes encoding structural subunits of the evolutionarily conserved, 10-subunit exoribonuclease complex, the RNA ...exosome. This complex consists of a three-subunit cap, a six-subunit, barrel-shaped core, and a catalytic base subunit. While a number of mutations in RNA exosome genes cause pontocerebellar hypoplasia, mutations in the cap subunit gene
cause an apparently distinct clinical presentation that has been defined as a novel syndrome SHRF (
hort stature,
earing loss,
etinitis pigmentosa, and distinctive
acies). We generated the first in vivo model of the SHRF pathogenic amino acid substitutions using budding yeast by modeling pathogenic
missense mutations (p.Gly30Val and p.Gly198Asp) in the orthologous
gene
The resulting
mutant cells show defects in cell growth and RNA exosome function. Consistent with altered RNA exosome function, we detect significant transcriptomic changes in both coding and noncoding RNAs in
cells that model
p.Gly198Asp, suggesting defects in nuclear surveillance. Biochemical and genetic analyses suggest that the Rrp4 G226D variant subunit shows impaired interactions with key RNA exosome cofactors that modulate the function of the complex. These results provide the first in vivo evidence that pathogenic missense mutations present in
impair the function of the RNA exosome. This study also sets the stage to compare exosomopathy models to understand how defects in RNA exosome function underlie distinct pathologies.
Abstract
The RNA exosome is an essential ribonuclease complex required for processing and/or degradation of both coding and non-coding RNAs. We identified five patients with biallelic variants in ...EXOSC5, which encodes a structural subunit of the RNA exosome. The clinical features of these patients include failure to thrive, short stature, feeding difficulties, developmental delays that affect motor skills, hypotonia and esotropia. Brain MRI revealed cerebellar hypoplasia and ventriculomegaly. While we ascertained five patients, three patients with distinct variants of EXOSC5 were studied in detail. The first patient had a deletion involving exons 5–6 of EXOSC5 and a missense variant, p.Thr114Ile, that were inherited in trans, the second patient was homozygous for p.Leu206His and the third patient had paternal isodisomy for chromosome 19 and was homozygous for p.Met148Thr. The additional two patients ascertained are siblings who had an early frameshift mutation in EXOSC5 and the p.Thr114Ile missense variant that were inherited in trans. We employed three complementary approaches to explore the requirement for EXOSC5 in brain development and assess consequences of pathogenic EXOSC5 variants. Loss of function for exosc5 in zebrafish results in shortened and curved tails/bodies, reduced eye/head size and edema. We modeled pathogenic EXOSC5 variants in both budding yeast and mammalian cells. Some of these variants cause defects in RNA exosome function as well as altered interactions with other RNA exosome subunits. These findings expand the number of genes encoding RNA exosome subunits linked to human disease while also suggesting that disease mechanism varies depending on the specific pathogenic variant.
Noonan syndrome with multiple lentigines (NSML) is a rare autosomal dominant disorder that presents with cardio-cutaneous-craniofacial defects. Hypertrophic cardiomyopathy (HCM) represents the major ...life-threatening presentation in NSML. Mutations in the PTPN11 gene that encodes for the protein tyrosine phosphatase (PTP), SHP2, represents the predominant cause of HCM in NSML. NSML-associated PTPN11 mutations render SHP2 catalytically inactive with an "open" conformation. NSML-associated PTPN11 mutations cause hypertyrosyl phosphorylation of the transmembrane glycoprotein, protein zero-related (PZR), resulting in increased SHP2 binding. Here we show that NSML mice harboring a tyrosyl phosphorylation-defective mutant of PZR (NSML/PZRY242F) that is defective for SHP2 binding fail to develop HCM. Enhanced AKT/S6 kinase signaling in heart lysates of NSML mice was reversed in NSML/PZRY242F mice, demonstrating that PZR/SHP2 interactions promote aberrant AKT/S6 kinase activity in NSML. Enhanced PZR tyrosyl phosphorylation in the hearts of NSML mice was found to drive myocardial fibrosis by engaging an Src/NF-κB pathway, resulting in increased activation of IL-6. Increased expression of IL-6 in the hearts of NSML mice was reversed in NSML/PZRY242F mice, and PZRY242F mutant fibroblasts were defective for IL-6 secretion and STAT3-mediated fibrogenesis. These results demonstrate that NSML-associated PTPN11 mutations that induce PZR hypertyrosyl phosphorylation trigger pathophysiological signaling that promotes HCM and cardiac fibrosis.
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