UNIVERSITY OF MASSACHUSETTS
Our current work under this grant has focused on understanding the function of one of the FYVE-domain containing proteins, WDFY2 in cultured cells. We identified this previously uncharacterized protein, which contains a FYVE domain embedded within WD40 repeats, in an siRNA screen in C. elegans using a background that revealed endocytic defects (Hayakawa et al. (2006) Proc Natl Acad Sci U S A. 103(32):11928-33)The mammalian homologue of WDFY2 is expressed ubiquitously, and its depletion 3T3-L1 adipocytes results in an inhibition of insulin-stimulated glucose transport. We determined that this effect is at least in part due to an isoform-specific depletion of the downstream serine-threnonine kinase Akt2 in response to WDFY2 knockdown. We have now determined that in intact cells recruitment of endogenous Akt2 to WDY2 is isoform-specific, and that signaling events downstream of Akt2 are affected differentially by WDFY2 depletion. In the remaining two years of the current project, we seek to take advantage of emerging technologies to achieve more quickly and with increased value the research goals of the original project, which are to understand the function of the FYVE-domain-containing protein family. In addition, in keeping with the goals and spirit of ARRA, we will train personnel to be proficient in these new technologies, increasing their ability to keep or obtain work in academia or industry after the two years contemplated in the supplement. With resources from this supplement we seek to study, in parallel, the entire group of 14 highly conserved FYVE-domain-containing proteins at the organism level using the vertebrate model Dario rerio (Zebrafish). This will be possible due to the recent establishment at UMASS Medical School of a Zebra Fish Functional Genomics Core Facility, which will allow ?non-fish? researchers to initiate studies using zebrafish. We requested an administrative supplement to achieve these goals, and received an award of $ 92,845, which was used entirely for the purchase of a new microscope required for these studies.