Coordinated Drosophila Class II PI3-kinase Variants Interact with Myotubularin Phosphatase to Regulate Autophagy and Longevity

Autophagy(definition) is the degradation of cytoplasm in lysosomes to enable cellular homeostasis. It is also the adaptation strategy to deal with cellular damage and nutrient shortages. Autophagy is critical to cell health and relevant to diseases, such as Huntington’s disease and cancers. Class I and III PI3-kinases are well-studied and have been shown to regulate autophagy, but roles for class II PI3-Kinase (PI3KC2) have not been fully investigated. We utilized a well-built system in fat body of Drosophila larvae to study functions of PI3KC2 in autophagy. In flies, we found two alternatively spliced PI3KC2 variants: PI3KC2 kinase and a truncated, non-catalytic form(PI3KC2-short). Using targeted deletions, we showed that wild-type PI3KC2 is required to restrict basal autophagy levels, while PI3KC2-short is needed to derepress this inhibition to induce autophagy. Our past data uncovered indirect protein interactions between PI3KC2 and Mtm (myotubularin) PI3-phosphatase, raising questions on the significance of PI3KC2-short in protein complex with Mtm and enzyme regulation of autophagy. This thesis presents strong evidence of protein-protein interactions between PI3KC2-short with both PI3KC2 and Mtm, and further genetically supports that PI3KC2-short acts upstream of both enzymes to promote autolysosomal maturation. Although deletion of PI3KC2-short causes a block in autolysosomal maturation with a lack of cathepsin B activity, another cysteine cathepsin protein L was processed to normal mature forms. The two PI3KC2 variants were both found to be broadly co-expressed and co-required for normal adult fly longevity, motivating initial studies to explore PI3KC2 requirements for protein aggregate clearance with fly aging. In contrast, mutants lacking only PI3KC2 resulted in extended survival in response to adult fly starvation, suggesting that the enhanced autophagy levels in this condition may provide a stress response advantage. In conclusion, PI3KC2 is a key regulator of autophagy levels. Future study of PI3KC2 in flies allows better understanding of its functions which can be applied to solve pathological issues in humans.