Regulation of Drosophila larval fat body development and function by tissue-specific SPARC variants

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Basement membranes (BMs) play an indispensable role in development, growth, regenerative capacity, and tissue homeostasis of all multicellular organisms. A core component of BMs includes network-forming Collagen IV (ColIV). However, the molecular processes controlling the assembly of BMs are poorly understood. Non-core components are required for BM assembly and remodelling, often in a tissue-specific manner. Work in my laboratory using Drosophila has established SPARC as an essential molecular chaperone for ColIV, enabling diffusion of non-polymerized ColIV from its central site of production in the larval fat body to distal sites, such as the wing imaginal discs. We have shown that loss of SPARC results in fibrotic-like accumulation of ColIV within the fat body and 2nd instar larval lethality. SPARC consists of an acidic calcium-binding N-terminal domain, a central follistatin-like domain, and a C-terminal domain containing two highly conserved collagen-binding epitopes and two high-affinity calcium-binding EF-hands. The highly conserved collagen-binding epitopes and EF-hands of SPARC enable association of SPARC with ColIV. We determined that a loss in the ColIV-binding capacity of SPARC is responsible for the larval lethality. Moreover, loss of SPARC expression in the fat body is sufficient for lethality. Importantly, SPARC generated by the wing imaginal disc is able to migrate to the fat body, but does not associate with BMs in either tissue, implying distinct roles for SPARC derived from these tissues. We determined that SPARC first colocalizes with ColIV within the trans-Golgi of hemocytes and fat body adipocytes indicating a role in preventing intracellular ColIV polymerization. Collectively, our data provide the first in vivo evidence that different variants of SPARC are expressed by tissues with distinct impact on BM dynamics, fat body function and larval development. We hypothesize that wing imaginal disc-derived SPARC, which has a similar molecular weight to that of SPARC secreted by the fat body, represents a glycoform incapable of binding ColIV and with distinct functions from that produced by the fat body. In this proposal, a series of strategies are proposed that continue to take advantage of the genetic tractability and molecular resources of Drosophila to gain further mechanistic insight into the diverse morphoregulatory contributions of SPARC during larval development. Our work to date provides strong evidence that SPARC functions as a chaperone for Col(IV) enabling appropriate tissue distribution. However, key mechanistic questions remain that need to be addressed to fully accept this unique function of a secreted extracellular protein. The work outlined in this proposal will provide this information and will test our hypothesis that wing imaginal disc-derived SPARC represents a glycoform incapable of binding Col(IV) and exerts distinct functions from that produced by the fat body-derived SPARC.