Requirement of the Caspase-3 Homologue DrICE in Drosophila Tracheal Size Control

McSharry, Saoirse Susan

doi:https://doi.org/10.21985/n2-gxph-9p86

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Requirement of the Caspase-3 Homologue DrICE in Drosophila Tracheal Size Control

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Proper size control of organs and tissues is critical to their function, and it is necessary for the millions of precisely sized tubes that make up those organs— for example, excessive cell growth can lead to devastating diseases such as Polycystic Kidney Disease. The regulation of tube growth is therefore essential to organismal health. Many key molecular pathways are known to control aspects of growth through managing a desirable balance between cell division and cell death. In this work, I describe a novel feature of this management whereby a protein known for executing cell death, DrICE, is required for the proper lengthening of an epithelial tube in Drosophila, the trachea, but it does so without causing apoptosis. In Chapter 2, I describe a novel mechanism by which DrICE, a component of cell death machinery, is required for proper tracheal size control in the fruit fly Drosophila melanogaster. Although well known for its role in apoptosis, DrICE, an executioner caspase, has a non-apoptotic function that is required for elongation of the epithelial tubes of the Drosophila tracheal system. I show that DrICE acts downstream of the Hippo Network to regulate endocytic trafficking of at least seven cell polarity, cell junction and apical extracellular matrix proteins involved in tracheal tube size control. I further show that tracheal cells are competent to undergo apoptosis, even though developmentally-regulated DrICE function rarely kills tracheal cells. My results reveal a novel developmental role for caspases, a previously unidentified pool of DrICE that colocalizes with endocytic markers, and a mechanism by which the Hippo Network controls endocytic trafficking. Given published reports of in vitro regulation of endocytosis by mammalian caspases during apoptosis, I propose that caspase-mediated regulation of endocytic trafficking is an evolutionarily conserved function of caspases that can be deployed during morphogenesis. In Chapter 3, I explore the results of the candidate approach for caspase substrate identification, in which I probed the Sterol Regulatory Element Binding Protein (SREBP) as a potential tracheal-elongating substrate of DrICE. A series of epistasis experiments with new transgenic and CRISPR alleles of dSREBP shows that DrICE may cleave and activate dSREBP in the trachea. I show that overexpression of the canonical S2P-processed form of dSREBP is lethal to tracheal cells, and that our new CRISPR-mediated loss-of-function allele, dSREBPORF, causes dramatic synthetic lethality when combined with yorkieB5 and DrICE17 mutant alleles. I also demonstrate that dSREBPORF has the same epistatic relationships with septate junction loss-of-function alleles as DrICE17, which is consistent with a model in which there is interplay between dSREBP and DrICE. The exact molecular mechanism of the relationship between DrICE and lipid levels is complex and remains to be elucidated. Clarifying this relationship, however, could have important consequences for our understanding of the interactions between lipid metabolism, apoptosis, and disease.

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