Poster abstracts
Poster number 31 submitted by Pooja Gangras
EJC core protein Rbm8a is critical for muscular and neural development in zebrafish
Pooja S. Gangras (Department of Molecular Genetics), Thomas L. Gallagher (Department of Molecular Genetics), Kiel T. Tietz (Department of Molecular Genetics), Natalie C. Deans (Department of Molecular Genetics)
Abstract:
Nonsense Mediated mRNA Decay (NMD) is a post-transcriptional mechanism that degrades aberrant mRNAs and regulates ~30% of the eukaryotic transcriptome to execute key cellular processes. ‘Physiological’ NMD targets can arise by regulated alternative splicing events that introduce a premature termination codon or contain ‘NMD-inducing features’ such as 3’ UTR introns. NMD targets are recognized by the Upf proteins during translation termination often with aid of a downstream EJC (Exon Junction Complex), a protein complex deposited 24 nucleotides upstream of most mRNA exon-exon junctions during pre-mRNA splicing. EJC function is essential for human health; mutations in EJC core components have been linked to intellectual disability, brain malformation and limb defects. To study EJC function in a whole animal model, we have generated zebrafish mutants in the EJC core proteins Rbm8a, Eif4a3 and Magoh. Homozygous rbm8a mutants survive well beyond gastrulation, likely due to maternal deposition of Rbm8a protein and rbm8a transcript; once maternal Rbm8a stores are depleted, phenotypic abnormalities appear, with neural and muscle tissues being particularly susceptible. I have shown that rbm8a mutant embryos appear morphologically normal but are completely paralyzed at 19 hpf (hours post fertilization), when movement normally begins. At the same stage, apoptotic cells are present throughout the rbm8a mutant brain and spinal cord. By 26 hpf, rbm8a mutant embryos have defects in myofibril organization and motor neuron outgrowth. Preliminary RNA-seq analysis shows significant downregulation of fast muscle myosin genes in rbm8a mutants compared to wild-type siblings. To distinguish direct versus indirect effects, I am identifying in vivo EJC footprints to reveal its preferentially bound RNAs. My goal is to determine the cellular and molecular mechanisms governed by the EJC during zebrafish development to ultimately help understand the role of EJC in NMD and in animal development.
Keywords: Exon Junction Complex, zebrafish, development