Role of maternal mRNA degradation in whole-body cellular phenotypes caused by DNA polymerase α deficiency Alex Lin Georgia Thomas Khai C. Ang Damian B. van Rossum Victor A. Canfield Keith C. Cheng 10.6084/m9.figshare.12145740.v1 https://tagc2020.figshare.com/articles/poster/Role_of_maternal_mRNA_degradation_in_whole-body_cellular_phenotypes_caused_by_DNA_polymerase_deficiency/12145740 <p>A histology-based zebrafish forward genetic screen for mutations causing cytological abnormalities yielded a mutant, <i>huli hutu </i>(<i>hht</i>), whose pleiotropic phenotype includes atypical nuclei in gastrointestinal cells and nuclear fragmentation in the retina and central nervous system. We have used a new, 3D form of histology based on micro-CT, X-ray histotomography, to reveal diminished heart volume and shape, and dysplastic cartilage and skin. We were also able to definitively establish the absence of a swim bladder and pneumatic duct, which is challenging to determine usig histology. Cell death shown by karyorrhexis was associated with DNA damage identified by ɤ-H2AX stainng in the brain, eyes, and spinal cord. These phenotypes were caused by a frameshift-based premature stop codon at the 38<sup>th</sup> amino acid position of the 600-amino acid Pola2 protein, or B subunit of DNA polymerase α (Pol α). An extended 120-168 hpf survival of the <i>hht </i>fish stands in striking contrast to the immediate cell cycle arrest of the corresponding mutants in yeast and <i>Arabidopsis. </i>We found that the prolonged lifespan of these mutants can be explained by the presence of wild-type maternal <i>pola2 </i>mRNA in the fertilized embryos. The gradual disappearance of wild-type mRNA in homozygous mutant embryos appears to cause diminished DNA synthesis, increased DNA damage, cell death, and tissue-dependent cytological deformities. The breadth of cytological, tissue, and organ phenotypes of <i>hht </i>mutants can be used as a model for developing computational tools for quantitative 3D characterization of pleiotropy, including cytological and tissue phenotypes across organ systems.</p> 2020-04-20 23:14:48 DNA Polymerase Alpha Cell Death DNA damage Nuclear Atypia Nuclear Fragmentation Maternal RNA Pleiotropy Genetics