Ramin Homayouni - US grants

[unreadable] DESCRIPTION (provided by applicant): The mammalian brain is formed through a series of intricately orchestrated events whereby neurons born in germinal zones migrate great distances to reach their final positions and form specific connections. Abnormalities in neuronal migration and positioning are believed to be responsible in part for disorders such as lissencephaly, pediatric epilepsy, schizophrenia and autism. Recent genetic studies in mice have identified a key signaling pathway that controls cell positioning and formation of laminated structures throughout the mammalian brain. Mice with disruptions in reelin, disabled-1 (Dab1), or both very low-density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2) genes exhibit nearly identical histopathological abnormalities. Reelin is an extracellular protein that directly binds to the lipoprotein receptors and induces tyrosine phosphorylation of Dab1. Dab1 is an intracellular adapter protein that is required for Reelin signaling. The long-range goal of this project is to identify molecular components downstream of Dab1 in the Reelin signaling pathway and to understand the mechanism by which Reelin controls neuronal positioning, dendritic maturation, and synaptic function. Using a yeast two-hybrid strategy, it was found that Dab1 interacts with amyloid precursor family proteins, protocadherin-18 and the novel GTPase activating protein Dab2 interacting protein (Dab2IP). The deduced amino acid sequence of Dab2IP encodes a Ras GAP related domain and several protein-protein interaction domains, including an NPxY PTB-interacting motif. It is hypothesized that Dab2IP functions as a regulator of GTPases and, by virtue of its interaction with Dab1 and other intracellular proteins, is the downstream effector in the Reelin signaling pathway. The specific aims of this proposal are to: 1) Characterize the activity, regulation and cellular function of Dab2IP; 2) Determine if Dab2IP plays a critical role in Reelin signaling; 3) Characterize the Dab2IP-P1-/- mice which have recently been generated in this laboratory. Understanding the biological function of Dab2IP and its role in Reelin signaling will provide valuable insight into the molecular mechanisms of neuronal migration, cell positioning and dendrite maturation during brain development. [unreadable] [unreadable]

The Chemistry Department at the University of Memphis will acquire a spectropolarimeter with a stopped flow module with this award from the Major Research Instrumentation (MRI) program. The requested instrument will facilitate ongoing research projects including the detection and quantitation of water disinfection by-products; the structure and function of phospholipid receptors; development of nanobioreactors for hydrogen production and biosensor application; characterization of a novel mitochondrial protein, NIPSNAP1.

The spectropolarimeter will produce circular dichroism spectral data which provide information on molecular structure, especially for proteins. The stopped flow capability will provide reaction kinetics data which give information on the speed of chemical and biological reactions. These are important tools in the training of young scientists. The instrument will be used in research by graduate and undergraduate students and in laboratory classes.

Abstract Mitochondrial dysfunction is associated with ageing as well as a number of age-related neurodegenerative diseases including Alzheimer?s Disease (AD). Early onset AD has been linked to mutations in amyloid precursor protein (APP) and presenilins 1 and 2 (PS1 and PS2), which result in abnormal cleavage of APP and release of toxic amyloid beta (A???peptides. Accumulating evidence suggests that APP intracellular domain (AICD) may also contribute to pathogenesis of AD. To gain insights into the normal and pathological roles of AICD, we previously used a biochemical affinity proteomic strategy and found that AICD directly interacts with a novel mitochondrial protein, Nipsnap1 (4-nitrophenyl phosphatase domain and non-neuronal SNAP-25 like protein homolog1). Although Nipsnap1 is evolutionarily conserved, very little is known about its function. Our long-term goal is to investigate the molecular and cellular function of Nipsnap1 and to determine its role in neurodegeneration. Toward this end, we generated a mouse with a targeted disruption of the Nipsnap1 gene. Disruption of Nipsnap1 expression profoundly affects intermediate metabolism and significantly increased apoptosis and neurodegeneration in the brain. Protein structure modeling and virtual ligand screening suggested that Nipsnap1 may bind to NADH and NADPH. Using in vitro biochemical assays, we found for the first time that Nipsnap1 directly binds to both NADH and NADPH. Moreover, we found significantly lower NAD+/NADH ratios in Nipsnap1 deficient brain. The balance between NAD+ and NADH is critical for production of ATP, maintenance of mitochondrial potential and regeneration of reducing agents within cells to counteract reactive oxygen radicals. Based on these preliminary results, we hypothesize that Nipsnap1 plays an important role in neuronal survival by modulating dehydrogenase activities and NAD(P)H levels. In this project, we will use biochemical approaches and primary neuronal cultures derived from WT and Nipsnap1 deficient mice to determine if: 1) Nipsnap1 interacts with and regulates multiple dehydrogenases in the mitochondria; 2) AICD interaction with Nipsnap1 affects dehydrogenase activity and neuronal NAD+/NADH levels. Our work will provide insights into the molecular function of Nipsnap1 and possibly a new mechanism by which AICD produces neurotoxicity.