Determining the Genotype‐Phenotype Relationship of atg18 Mutants

Abstract

8J16 and 9E6 are two independent allelic point mutations in Drosophila melanogaster autophagy‐specific gene 18a (atg18a) located in introns three and four, respectively, of its 5 exon locus at 66B11. The atg188J16 and atg189E6 intronic mutations are not in sequences expected to disrupt transcript processing, yet these mutations cause pupal lethality and neurodegenerative phenotypes by an unknown molecular mechanism. The purpose of this research is to examine the genotype‐phenotype relationship of atg188J16and atg189E6on neuronal maturation using the Drosophila eye as a model. RT‐PCR analysis revealed no detectable effects on atg18 mRNA splicing in either mutant background. In adults, atg188J16 and atg189E6 homozygous mutant eyes are small and display a rough eye phenotype. To understand the basis of this phenotype, homozygous mutant larval imaginal eye discs were examined. ELAV, a pan neuronal marker, showed that photoreceptors differentiate normally, but those that differentiate early are being lost by apoptosis as determined by the increased cleaved caspase 3. These mutations also caused significant death at the morphogenetic furrow, which may result in a smaller pool of photoreceptor progenitors. Hence, autophagy may have roles in both photoreceptor differentiation and maintenance and its absence leads to apoptotic death. Senseless and Prospero expression showed that photoreceptors R8 and R7, respectively, are more refractory to apoptosis as these photoreceptors predominate near the optic stalk. This suggests autophagy is more important for the maintenance of photoreceptors R1, R2, R3 R4, R5 and R6 than R7 and R8.Acknowledgements given to the Howard Hughes Medical Institute and University of Delaware Undergraduate Research Program for financial support.