Research Accomplishments

Identification of subunit configurations of the receptors for insulin and the insulin-like growth factors 

Prior to the 1980s, there was an appreciation that receptor proteins for these peptides were present on cell surface membranes, but there was no biochemical information on these receptors. To approach this problem, Paul Pilch, then a postdoc in our lab, synthesized disuccinimidyl suberate as an affinity-labeling reagent for peptide hormone receptors and identified the two insulin receptor subunits, denoting these as alpha and beta in several publications. This approach was then used to deduce the disulfide-linked subunit configurations of the receptors for insulin and the insulin-like growth factors (IGF). This work clarified the overlapping binding affinities of insulin and the IGF ligands for these receptors and revealed that the IGF-2 receptor does not mediate the major bio-effects of these peptides. We showed the IGF-2 receptor is instead a target of insulin signaling, cloned the IGF-2 receptor in collaboration with Axel Ullrich and reported in Science it's identical to the mannose-6-phosphate receptor, i.e., it's a bi-functional receptor. This affinity crosslinking technology was then successfully extended to other systems, most notably to the transforming growth factor receptors by one of our former fellows, Dr. Joan Massague.

Identification of a key downstream target of insulin-stimulated PIP3 generation

As several groups identified PI 3-kinase as a component in insulin signaling, it became important to identify downstream signaling elements. Our group established an expression cloning screen for identifying targets of the PI 3-kinase pathway in insulin and IGF-1 receptor signaling, and discovered Grp1, a novel downstream effector of the signaling lipid PtdIns(3,4,5)P3 (PIP3). We reported in Science the identification of Grp1, showing that it defines a novel PI 3-kinase-mediated signaling pathway distinct from Akt, linking PIP3 to the activation of ArfGTPases. Jes Klarlund in our lab showed the Grp1 PH domain has the highest specificity for PIP3 of all the PH domains studied. The structural basis of selective PIP3 binding to the crystallized PH domain of Grp1 was solved in collaboration with David Lambright, published in Molecular Cell.  The Grp1 PH domain fused to GFP is also now  widely used by researchers as a unique  reagent  to define the generation of PIP3 at the plasma membrane.

Identification of lipid droplet proteins and mechanisms of lipid storage

A focal point of our lab over many years has been the mechanisms that control triglyceride storage in adipocytes and their relationships to insulin resistance. We discovered a family of Cide-domain containing proteins is associated with lipid droplets in adipocytes and hepatocytes, and regulate lipid storage and turnover in these metabolic cell types. Gene deletion of Cidec/FSP27 and Cidea in mice has corroborated their key roles in lipid metabolism and whole body energy expenditure, and a human subject with a disrupting mutation in CIDEC displays lipodystrophy, insulin resistance and type 2 diabetes. We also found evidence that decreased lipid droplet protein expression in human adipose tissue may correlate with the appearance of insulin resistance, consistent with the hypothesis that they help sequester neutral lipids within adipocytes to protect other tissues from lipotoxicity. We continue to identify pathways of triglyceride storage and release, for example the HIG2 pathway and their relationships to systemic glucose tolerance and insulin sensitivity. This current work includes de novo fatty acid synthesis in adipocytes, and recent data shows this pathway may regulate white adipose tissue “browning” through regulation of UCP1 and other “Beige” adipocyte genes.

Development of siRNA delivery technology for research and therapeutic strategies

A major area of our research addresses the cellular and molecular mechanisms of inflammation and metabolic disease, especially obesity and diabetes. Recognizing the power and potential of RNAi as a therapeutic tool, we developed siRNA screens and identified Map4k4 and RIP140 as metabolic regulators. Our group collaborated with Gary Ostroff to develop a novel siRNA delivery system (GeRPs) based on yeast cell wall glucan shells to encapsulate siRNA cargo. This system delivers siRNA specifically to phagocytic cells in vivo, and we've demonstrated silencing of inflammatory genes in mouse macrophages following administration in vivo. We've now published extensively on the effectiveness of siRNA silencing in vivo using GeRPs, in papers by our laboratory and in collaboration with other research groups. Thus this technology can now be transferred very successfully to other laboratories, which have independently reproduced our gene silencing results. We're particularly excited about our recent work showing gene silencing in selective subpopulations of macrophages in visceral adipose tissue and in Kupffer cells in the liver. These data show that tissue - localized macrophages and macrophage - like Kupffer cells do indeed release deleterious cytokines and factors that cause insulin resistance, but also play beneficial roles in cell physiology. This work has been extended to the use of  “self delivery” sdRNA and development of vehicles for delivery of CRISPR-based gene editing.