Cerebral Cavernous Malformations (CCMs) are brain vascular lesions estimated to affect up to 0.5% of the population. CCM patients can suffer chronic headaches, epilepsy, seizures, stroke and focal neurological deficits. Three disease-associated genes, which encode CCM1/KRIT1, CCM2/malcavernin/OSM, and CCM3/PDCD10, respectively, have been identified. These CCM proteins are known to form a multi-protein complex (the CCM complex signaling platform) and a loss of function of any one of these proteins leads to CCM pathology. Despite major progress in our understanding of the genetics and molecular functions of CCM proteins in mouse models, precisely how CCM is developed in patients with mutations in CCM genes remains largely unclear. One of the known binding partners of the CCM complex is a mitogen-activated protein kinase kinase kinase, MEKK3. Importantly, the role of MEKK3 in the vascular system appears to be overlapping with that of CCM proteins, especially that of CCM2. Therefore the goal of this project is to understand the structural requirement and functional consequences of MEKK3:CCM2 interaction and the associated changes in downstream signaling that may impact the critical vasculature phenotypes associated with CCM disease. Towards this end, our studies are designed to comprehensively address our central hypothesis that recruitment of MEKK3 by CCM2 is critical for vascular integrity. We will address this hypothesis in three Aims.
In Aim 1 we will define the structural and functional mechanisms for MEKK3 recruitment to CCM2.
This aim will use structural, biochemical and biophysical tools to provide the basic molecular level framework for how MEKK3 is recruited to the CCM complex by its interaction with CCM2.
In Aim 2 we will investigate the role of CCM2 in regulation of MEKK3 signaling by probing MEKK3 regulation by CCM2 using biochemical and cell based assays. We will also investigate the specific phosphorylation targets of the MEKK3:CCM2 complex and investigate the target genes of the complex.
In Aim 3 we will discover the in vivo functional role of MEKK3 recruitment to CCM2. We utilize structure-directed in vivo studies, in which we specifically disrupt the MEKK3:CCM2 interaction without compromising the overall activity of MEKK3, to test whether critical vasculature phenotypes associated with CCM disease, and associated changes in downstream signaling, result from loss of the MEKK3:CCM2 interaction. Overall, we expect that the studies we propose will define the functional importance of MEKK3 interaction with CCM2 and resolve whether the vascular pathology associated with CCM disease result from loss of the MEKK3:CCM2 interaction and associated changes in downstream signaling.
The devastating familial form of Cerebral Cavernous Malformations (CCM) disease is linked to loss of function of CCM2 (cerebral cavernous malformations 2) protein, therefore the loss of function of CCM2 is therefore directly associated with stroke, focal neurological defects, seizures and vascular abnormalities. A binding partner of CCM2, termed MEKK3, is also associated with vasculature defects on its loss in mice models. The goal of this application is therefore to understand the molecular underpinnings for normal interaction of these proteins, and whether MEKK3 plays a role in CCM disease.
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