Cerebral Cavernous Malformation (CCM) is a disease caused by the progressive development of large mulberry-like lesions in blood capillaries, afflicting approximately 1 in 500 individuals. CCM can arise sporadically by unknown mechanisms or by inheriting a mutation in one of three genes (CCM1, CCM2 or CCM3), which encode structurally unrelated scaffold proteins. CCM1 and CCM2 form a complex that functions to stabilize the vasculature by inhibiting the RhoA GTPase. Although CCM3 has been found to associate with CCM1/2, there is some debate about whether this is its primary mechanism of action. Loss of the CCM3 gene leads to early onset of the disease, often presenting in children, and has the most severe prognosis for patients. Previous work from co-author Anne-Claude Gingras’ lab showed that human CCM3 protein preferentially associates with the striatin interacting phosphatase and kinase complex, rather than CCM1/2.
To gain insight into the mechanism by which CCM3 functions in vivo we developed a disease model using the C. elegans excretory cell, a unicellular tube that extend canals along both sides of the body. The excretory cell resembles blood capillaries of vertebrates and serves as a primitive renal system for this organism. In this paper we report a novel mechanism by which the worm CCM3 gene (ccm-3) regulates extension and membrane integrity of the excretory canal. Specifically, we found that CCM-3 and its associated kinase GCK-1 (a member of the Ste20/GCKIII family) promote CDC-42 activity and endocytic trafficking. Consistent with biochemical data from the Gingras lab, we found that ablation of components of the worm STRIPAK complex caused similar defects in excretory canals as ccm-3 mutants, and we observed no role for the CCM1 homologue kri-1.
Taking advantage of the powerful genetics of C. elegans we performed a screen and identified myotonic dystrophy-related CDC-42-binding kinase (MRCK-1) and the exocyst complex as a components of the CCM-3 pathway. Ultrastructural analysis carried out in collaboration with Dr. Mei Zhen revealed a breakdown in Golgi and perturbations to the apical and basolateral membranes of excretory cells in ccm-3 mutants, consistent with CCM-3/STRIPAK regulating endocytic trafficking. The C. elegans model offers new insights into the mechanism by which CCM3 functions in biological tube development, which should help in the development of therapeutic strategies for treating this disease. Towards this goal, Dr. Derry and Dr. Peter Roy were recently awarded an E-RARE grant to screen for small compounds that reverse the phenotypes of ccm-3 and kri-1/CCM1 mutants in C. elegans. Together with international collaborators in Germany and France they are using worms, zebrafish and mouse models of CCM disease to identify potential drugs for treating this disease.
This work was supported by a CIHR grant to Dr. Gingras and Dr. Derry. Dr. Lant was supported by a CIHR-funded Excellence in Radiation Research for the 21st Century (EIRR21) training grant.
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