Russell T. Matthews, Ph.D. - US grants

DESCRIPTION (provided by applicant): Primary tumors of the central nervous system, gliomas, are notoriously difficult to control, due in large measure to their highly invasive behavior. The ability of cells to migrate through tissue depends, in part, on the composition of the tissues extracellular matrix (ECM). Understanding the molecular composition of the extracellular environment of brain tumors is one approach to developing methods to prevent tumor invasion. BEHAB/brevican, a recently identified brain-specific ECM protein, is markedly upregulated in human glioma and in experimental models of glioma. In human glioma BEHAB/brevican expression is 700 percent over that in the normal adult human brain and expression is not detected in any non-glial derived tumor, even when these tumors grow within the brain. Our work during the previous funding period has provided evidence that BEHAB/brevican plays a role in glioma invasion. The experiments proposed here will test this hypothesis and, in addition, will test the possibility that BEHAB/brevican may provide a novel therapeutic target for glioma. In experimental glioma cel models, BEHAB/brevican expression is induced by a brain-specific factor. The first specific aim of this application is to characterize the mechanism of BEHAB/brevican induction and to identify potential inducing factors. Several converging lines of evidence suggest that proteolytic processing is required for BEHAB/brevican function in glioma invasion. The second specific aim of this application is to characterize BEHAB/brevican Droteolytic cleava2e in glioma. Slowing or preventing glioma cell motility could increase the efficacy of regional therapies; the third specific aim of this application is to determine whether inhibitors of BEHAB/brevican cleavage can slow tumor invasion and, therefore, might provide a new therapeutic avenue foi glioma. Our work has led to the hypothesis that production, cleavage, and turnover of BEHAB/brevican together modulate the ECM and can regulate cell motility. Our final specific aim is to test a role for BEHAB/brevican turnover in glioma invasion. Our long-term goal is to determine if functional reduction or elimination of BEHAB could slow the progression of primary brain tumor.

The PILOT PROJECT PROGRAM is designed to bring new investigators into the field of alcohol research-by providing funds (a) to young investigators and (b) to established investigators who propose projects departing from their ongoing research. Funding from this program will be sufficient to obtain pilot data that can be used to support applications for independent research grants or can seed future reapplications for DEARC funding. The process for competing for pilot funding includes internal solicitation of proposals and a rigorous review process relying on internal and external reviews to select studies of the highest quality. This process is described in the ADMINISTRATIVE CORE. We have allotted eight slots for Pilot Projects, two projects will commence in each of the first four years of the DEARC funding. The current application lists four potential pilot projects that have passed through our internal and external review processes. The other projects have been left open to allow for the vibrancy and fluidity of the field of alcohol research (though two potential projects have been included in the description). The initial three studies are as follows. (1) Russell Matthews (Asst. Prof., Dept. of Neuroscience and Phyiology, Upstate) is an expert in the role of extracellular matrix in the developing nervous system. He proposes to examine the effects of prenatal exposure to ethanol on plastic events in the development of barrels in somatosensory cortex. (2) John Lombardo (Prof., Dept. of Psychology, SUNY- Cortland) studies the toxicity of polychlorinated biphenyls. Among the issues that he will explore is the effect of perinatal exposure to PCBs to increase ethanol intake on sexual dimorphisms. (3) Sandra Mooney (Asst. Prof., Dept. of Neuroscience and Physiology, Upstate) is investigating fetal programming. She proposes that exposure to ethanol at a key time, gastrulation or neuronogenesis, can lead to long-term changes in social behavior in adolescents. (4) Michael Nishnikov (Instructor, Binghamton University, Binghamton) will explore the role of kappa opioid receptors (a) as mediators of perinatal ethanol exposure and (b) in ethanol acting as an appetitive/aversive substance. Each of the proposed Pilot Projects integrates with at least one MAIN PROJECT and uses at least two of the scientific CORES. In summary, the PILOT PROJECT PROGRAM will serve as the seed for alcohol research in Central New York. It will encourage new researchers and new ideas to explore ethanol toxicity in developing nervous systems.

DESCRIPTION (provided by applicant): Mutations in enzymes involved in protein O-mannosyl glycosylation produce a group of devastating diseases characterized by congenital muscular dystrophy (CMD), type II lissencephaly, and eye abnormalities. Previous studies using animal models have demonstrated that hypoglycosylation of a-dystroglycan critically contributes to the pathogenesis of these disorders. A growing body of evidence, however, suggests that other substrates for O-mannosyl glycosylation likely contribute to particular brain abnormalities found both in animal models and in patients with CMDs. Unfortunately, previous studies have failed to identify the additional neural substrates of O-mannosylation that contribute to these brain pathologies. Using an animal model of CMD in which the glycosyltransferase POMGnT1 is knocked out we identified RPTP? as another important substrate for O-mannosyl glycosylation in the brain. Receptor tyrosine phosphatase zeta/beta is a receptor phosphatase that is found predominately in the central nervous system and is highly expressed during key stages of neural development. Both membrane-bound receptor and secreted variants of the protein are expressed in the brain and are high affinity ligands for a number of developmentally important adhesion molecules, growth factors and extracellular matrix proteins. We hypothesize that the hypoglycosylation of RPTP6 due to disrupted O-mannosylation alters the interactions of RPTP? with these key ligands and contributes to abnormal brain development. Consistent with this hypothesis we found that cortical neuron development in cultured cells from POMGnT1 knockouts is abnormal when they are plated on a subset of known RPTP? ligands. In this proposal we investigate the hypothesis that the hypoglycosylaton of RPTP? in O-mannosyl glycosylation mutant animals alters its ligand-binding characteristics thereby leading to aberrant cellular interactions and abnormal brain development. The proposal is focused on the following 3 specific aims: 1) To determine whether altered glycosylation of RPTP? modulates its ligand binding characteristics. 2) To determine whether disrupted glycosylation of RPTP? alter cell development. And 3) To identify the 1-dystroglycan-independent neural abnormalities in animal models of CMDs and to determine if these abnormalities are due to altered RPTP?/phosphacan glycosylation. To explore these questions, we will utilize a variety of genetic mouse model systems in which we will disrupt O-mannosyl glycosylation, a-dystrglycan expression or RPTP?expression. Studies will be conducted both in vitro and in vivo to determine how O-mannosyl glycans modulate RPTP? function and if this contributes to brain abnormalities found in animal models of CMDs. These studies will determine if RPTP? is an important contributor to CMD pathogenesis and will provide important insights into the pathomechanisms of the neural phenotypes in these disorders. PUBLIC HEALTH RELEVANCE: Altered protein glycosylation is a well-known cause of congenital muscular dystrophies. However, all the protein targets of altered glycosylation in these disorders are not known. In this proposal we investigate RPTP? as an important new target of the disrupted glycosylation in congenital muscular dystrophies in investigate its role in animals models of these disorders.

Perineuronal nets (PNNs) are conspicuous neural extracellular matrix (ECM) structures that have garnered significant interest over the last decade for the critical roles they play in neural developmental plasticity. These complex macromolecular structures are implicated in an array of cognitive functions, and are altered in a variety of neurological disorders. Despite the growing interest in PNN functions, the mechanisms by which they modulate neural functions are poorly understood, because there are currently no tools or techniques to manipulate PNNs specifically. We surmise that our inability to target and disrupt PNNs is primarily driven by a lack of understanding of their molecular composition or structure. Our goal in this proposal is to conduct a structure-function analysis of known PNN components as well as to identify proteins that anchor nets to neuronal surfaces. Using a powerful combination of in vitro and in vivo approaches, we have obtained strong preliminary data detailing how the newly identified PNN component receptor protein tyrosine phosphatase zeta (RPTP?) associates with tenascin-R (TNR) within PNNs at a molecular level. Furthermore, our data indicate that the RPTP??TNR complex anchors PNNs to the neuronal cell surface via the GPI-linked protein contactin-1 (CNTN1), which makes CNTN1 the first surface binding protein for PNNs ever identified. Our central hypothesis is that there are a set of unique components and receptors of PNNs that nucleate PNNs and anchor them to specific neuronal cell surfaces, thereby defining their unique structure and functions. The overall objective of this proposal is to identify PNN-specific components and dissect the formation of PNNs through a unique combination of proximity-labeling assays, protein-binding assays, and protein X-ray crystallography in order to create the tools to target and manipulate these structures specifically and precisely. Our long-term goal is then to use these tools to dissect PNN function in order to better understand disease pathogenesis and ultimately to target PNNs therapeutically. Guided by our strong preliminary data, this proposal seeks to discover the unique components that guide the assembly of PNNs by pursuing three non-overlapping specific aims: 1) defining the role of the RPTP??TNR complex in anchoring PNNs to neuronal surfaces; 2) pursuing the biochemical and structural characterization of interactions between ACAN, HAPLN1, and TNR; and 3) identifying cell surface receptors and novel components of PNNs. The proposed work is significant because it will attempt to identify the key unique components that contribute to the formation and thereby function of PNNs. Successful completion of the aims will provide key insights and reagents to manipulate PNNs specifically and precisely and ultimately understand their functional mechanisms. This approach is innovative because it brings together a novel combination of physiological, biochemical and structural approaches to investigate these important macromolecular assemblies in the central nervous system. Ultimately, the proposed work could be transformative for the field and lead to key mechanistic insights into of PNN function in health and disease.