Ronald R. Waclaw, Ph.D. - US grants

? DESCRIPTION (provided by applicant): Oligodendrocytes (OLs) are the cells that myelinate axons in the central nervous system. OL development begins with ventricular zone (VZ) progenitors that give rise to oligodendrocyte progenitor cells (OPCs), which subsequently proliferate, migrate, and differentiate into mature myelinating OLs. This multistep process requires precise regulation of intracellular signaling. The overall goal of this proposal is to identify cooperating signaling pathways that affect distinct stages of OL development. Using mouse genetics, we found that the intracellular protein tyrosine phosphatase Shp2, a RASopathy gene that has a positive role in the RAS/MAPK pathway, is a key regulator of multiple stages of OL development. Specifically, we found that altering Shp2 phosphatase activity impacts the generation of OPCs and also OL myelination. Interestingly, downstream of Shp2 function, we detect changes in MAPK and WNT signaling. Here we propose to test the hypothesis that altering the balance of Shp2 phosphatase activity results in changes in MAPK and WNT pathway activation that results in abnormal OPC generation and OL myelination. In the first aim, we will test if Shp2 phosphatase activity and/or adaptor function(s), independent of phosphatase activity, are critical in OPC/OLs and determine if the OPC and myelination abnormalities caused by altered Shp2 phosphatase activity impacts animal behaviors. In the second aim, we will identify MAPK pathway dependent and independent roles downstream of Shp2 signaling during OL development. In the third aim, we will identify the role of the WNT pathway in abnormal OL development caused by Shp2 RASopathy mutations. These studies will identify the specific roles of MAPK versus WNT signaling downstream of altered Shp2 function OL development, and provide a new understanding of the complex relationships between multiple signaling pathways during OL development.

The basal ganglia are known to regulate motor function and have recently been implicated in both social and cognitive functions as well. As a result, these brain nuclei have been implicated in childhood disorders, ADHD, OCD, Tourette's syndrome and autism, which display a spectrum of behavioral abnormalities. These childhood disorders have been proposed to result from abnormal development/function of basal ganglia circuitry. The striatum represents the major nucleus of the basal ganglia and the striatal projection neurons (SPNs) comprise the major neuronal subtype on which the basal ganglia circuit is dependent. The direct pathway (d)SPNs project to the output nuclei of the basal ganglia, while the indirect pathway (i)SPN axons innervate an intermediate nucleus and indirectly influencing the output nuclei though a polysynaptic circuit. Balanced activity between these two pathways is fundamental for normal brain function. Despite the importance of these striatal output pathways, little is known about the molecular genetic mechanisms controlling their formation. In the previous funding cycle, we showed that the transcription factor Isl1 is required for the normal formation of dSPNs. In its absence, these neurons are generated but do not survive and as a result, innervation of the output nuclei is severely compromised. Isl1 conditional mutants (cKOs) exhibit behavioral abnormalities reminiscent of ADHD as they are hyperactive and blunted to psychostimulant treatment. Moreover, we identified the transcription factor Sox8 in dSPNs. Our data indicate that the direct pathway axons do not project properly in Sox8 homozygous mutants. However, unlike the Isl1 cKOs, no SPN cell death was observed. Interestingly, Sox8 heterozygotes showed a partial phenotype with reduced direct pathway axonal innervation. Both the heterozygous and homozygous Sox8 animals exhibited hyperactivity, reminiscent of Isl1 cKOs, as well as, cognitive impairments. The main goal of this proposal is to understand the molecular genetic pathways controlling the development of dSPNs and specifically the roles of the transcription factors Sox8, Bach2 and Arx with respect to neuronal survival/differentiation and axon outgrowth. We will achieve this by testing the following hypotheses: 1) Sox8 regulates dSPN axon outgrowth downstream of Ebf1 by controlling the timing of maturation, 2) Isl1 regulates a Foxo/Bach2-mediated survival/differentiation pathway in developing dSPNs and 3) Arx is required for development of dSPNs and their altered development in Arx mutants accounts for certain behavioral defects observed in these mutants. Our approach will combine molecular and cellular analysis of genetic mouse mutants exhibiting defined alterations in dSPN connectivity and correlate this with specific behavioral abnormalities in motor and cognitive function. The genetic models in this proposal may inform human studies of ADHD, OCD, Tourette's as well as autism and intellectual disabilities.