The long-term goal of this application is to elucidate the role of a recently described family of proteins termed Clics (chloride intracellular channels) in epithelial morphogenesis. Although the acronym """"""""Clic"""""""" implies that these proteins may function as ion channels, emerging evidence indicates that they also have alternative functions distinct from ion transport. The central hypothesis to be tested is that Clics are regulatory proteins that control cytoskeletal activity and polarity during epithelial cell morphogenesis. Clics have been identified in association with the actin cytoskeleton and signal transduction through genetic, biochemical, and subcellular localization studies; however, their precise roles remain to be defined. Recent studies have shown that disruption of Clic genes causes tubular cyst formation in C. elegans and deafness due to structural degeneration of sensory epithelial cells in mice. In addition, human Clics have been implicated as diagnostic indicators and causative factors in tumor formation and metastasis. The function of Clics has not been rigorously tested. In this proposal, Drosophila will be used as a model system to unravel the functional significance and cellular roles of Clics. Unlike vertebrates, which have multiple Clic genes and splice variants, Drosophila has only one Clic gene. Our preliminary data indicate that Drosophila Clic functionally interacts with cytoskeletal proteins and signaling molecules essential to fly development. The major objectives are to examine the role of Clic in cytoskeletal regulation during embryonic epithelial morphogenesis and to broaden our perspective on the role of Clic in signaling during embryonic development as follows: 1. Investigate the role of Clic during embryonic epithelial morphogenesis. We will test whether Drosophila Clic is required for establishing the position of segmentation determinants in early embryogenesis. In addition, we will investigate the role of Clic in epithelial invagination and epithelial sheet movement late in embryogenesis. Potential interactions between Clic and known regulatory proteins key to epithelial polarity will be tested genetically and biochemically. 2. Identify genes which interact with Clic during embryonic development. We will identify genetic modifiers of Clic, verify the interactions, and assess the cellular functions of Clic modifiers during embryonic development. In addition, we will test candidate regulatory genes for functionally important interactions with Clic during morphogenetic movements of epithelial sheets. Since many proteins are functionally conserved among vertebrate and invertebrate species, it is likely that analysis of Clic function in Drosophila will lead to a better understanding of Clic function in humans. ? ? PUBLIC HEALTH REVELANCE: The main goal of this proposal is to determine how Clic proteins, which are highly conserved between humans and insects, control cell shape and movement. Clic proteins are relevant to human health because they are essential for hearing and balance and have been implicated in tumor formation and metastasis. The fruit fly will be used as a model system to understand how Clic works with other genes and proteins known to be important for cell shape and movement during embryonic development. ? ? ?
