Non-specific X-linked mental retardation (MRX) is characterized by mental impairment without any other distinctive clinical features. More than a dozen loci have been implicated in this condition, and importantly, of the eight genes that have been identified to date, three encode regulators or effectors of members of the Rho family of GTPases. These findings have led to the hypothesis that abnormal Rho GTPase signaling may be a prominent cause of MRX. However, how alterations in Rho signaling result in changes in neuronal connectivity and/or plasticity that give rise to MRX remain unknown. In this proposal, we will focus on the functional characterization of oliophrenin-1, a putative Rho GTPase activating protein (RHO GAP) that is lacking in unrelated MRX families.
Our goal is to elucidate how a lack of oligophrenin-1 results in a mental retardation phenotype and how its interaction(s) with Rho family GTPases (which include RhoA, Rac and Cdc42) contributes to this phenotype. To achieve this goal, we will employ complementary cellular and molecular experimental approaches. First, we will determine what effect(s) absence and ectopic expression of oligophrenin-1 has on the cellular morphology of developing neurons. This will involve performing live cell imaging of biolistically-transfected pyramidal neurons in hippocampal slices using two-photon miscroscopy. Second, we will investigate the signaling pathways in which oligophrenin-1 participates, by determining which target Rho GTPase(s) it acts upon in neuronal cells. To this end, we will perform RhoA, Rac1 and Cdc42 activity assays in vivo and compare the phenotypes of dominant negative and activated mutant forms of the Rho GTPases with those generated by gain-of and loss-of function conditions for oligophrenin-1.
Finally, we will further define the molecular and signaling functions of oligophrenin-1 by identifying new interacting proteins. The effects of selected oligophrenin-1 interacting proteins on dendritic arborization and spine formation will be examined in the context of oligophrenin-1 signaling using the above hippocampal slice system.
We believe that studies on oligophrenin-1 will correlate basic cellular and molecular biology with the disease phenotype of mental retardation. These studies will advance our understanding of neuronal development and determine how the disruption of proteins intimately associated with Rho GTPases contributes to MRX.