Identification of Essential Functions of GRP94 in Metazoan Growth Control and Epithelial Homeostasis

Abstract

<p>GRP94, the endoplasmic reticulum Hsp90, is a metazoan-restricted chaperone essential for early development in mammals, yet dispensable for mammalian cell viability. These data suggest that GRP94 is required for important developmental processes relying on cell-cell communication and cell-cell interaction. Consistent with this hypothesis, loss of GRP94 expression in mouse is embryonic lethal yet tissue culture cells expressing no GRP94 are viable. To date, functional studies of GRP94 have relied on cell-autonomous model systems, the use of which has lead to discoveries of proteins that GRP94 chaperones also called client proteins. These systems give limited insight into the essential role(s) played by GRP94 in metazoan biology. The dichotomy that GRP94 is necessary for metazoan life, but dispensable for cellular viability suggests that the chaperone is required for the functional expression of secretory and/or membrane proteins that enable cells to function in the context of tissues.</p><p>To explore this hypothesis, the <italic>Drosophila</italic> ortholog of <italic>GRP94</italic>, <italic>Gp93</italic>, was identified and <italic>Gp93</italic> mutants were created using imprecise P-element excision. <italic>Gp93</italic> was found to be an essential gene in <italic>Drosophila</italic>. Loss of zygotic <italic>Gp93</italic> expression is late larval lethal and causes prominent defects in the larval midgut, the sole endoderm-derived larval tissue. <italic>Gp93</italic> mutant larvae display pronounced defects in the midgut epithelium, with aberrant copper cell structure, markedly reduced gut acidification, atypical septate junction structure, depressed gut motility, and deficits in intestinal nutrient uptake. The metabolic consequences of the loss of <italic>Gp93</italic>-expression are profound; <italic>Gp93</italic> mutant larvae exhibit a starvation-like metabolic phenotype, including suppression of insulin signaling and extensive mobilization of amino acids and triglycerides. The defects in copper cell structure/function accompanying loss of <italic>Gp93</italic> expression resemble those reported for mutations in <italic>labial</italic>, an endodermal homeotic gene required for copper cell specification, and &alpha;-spectrin, thus suggesting an essential role for Gp93 in the functional expression of secretory/integral membrane protein-encoding labial protein target genes and/or integral membrane protein(s) that interact with the spectrin cytoskeleton.</p><p>The creation of <italic>Gp93</italic> mutant <italic>Drosophila</italic> has allowed for the study of GRP94 function <italic>in vivo</italic> and will be of upmost importance to future studies examining the function of this chaperone in all aspects of metazoan biology. This dissertation focuses on the morphological and physiological defects that accompany loss of <italic>Gp93</italic> expression in <italic>Drosophila</italic> larvae. It will also outline future studies utilizing this model.</p>