Regulation of Nutrient Homeostasis by COMMD proteins

PROJECT SUMMARY Copper (Cu) is a vital nutrient required by many essential enzymes. Cu-transporting ATPases, ATP7A and ATP7B, play pivotal roles in delivering Cu to proteins in the secretory pathway or to vesicles that mediate extracellular excretion of Cu. In response to varying levels in cellular Cu, ATP7A and B undergo dynamic movement between the trans-Golgi network and the endolysosomal system. Whole body Cu homeostasis is maintained by the liver, which is responsible for Cu excretion through ATP7B-mediated Cu delivery to the biliary canaliculus. Impaired Cu excretion, which can occur as a result of loss-of-function mutations in ATP7B, leads in Cu accumulation and tissue injury in the liver and brain, a condition known as Wilson’s disease. In addition, alterations in cellular systems that control ATP7A/B trafficking across the endolysosomal system can lead to altered Cu balance. An example of the latter is Cu toxicosis resulting from loss-of-function mutations in the COMMD1 gene, encoding an essential component of a larger complex known as the COMMD / CCDC22 / CCDC93 or CCC complex, which our group identified. The CCC complex sits at the apex of a hierarchy of regulatory systems that play essential roles in endosomal regulation, including endosomal recycling of transporters, such as ATP7A/B. Studies supported by the prior competitive period of this grant defined the principles by which the CCC complex regulates early endosomal function and recycling of endocytosed surface proteins back to the plasma membrane (PM), including elucidating the structural organization of the CCC complex. Interestingly, ATP7A/B trafficking to the plasma membrane has been shown to also involve late endolysosomal vesicles, implicating the CCC complex in their regulation through hitherto unknown mechanisms. In this proposal, we seek to uncover how the CCC complex regulates Cu excretion, focused on CCC-interacting factors in the late endolysosomal compartment. The project includes two specific aims, and each aim integrates biochemical and cellular studies, with animal models. In Aim 1 we will examine the role of Rab27a in supporting CCC-dependent trafficking of ATP7B. This aim is founded on preliminary studies showing that Rab27a binds to CCC directly and regulates ATP7B trafficking. Rab27a is known to be associated with late endolysosomal vesicles and to promote vesicle fusion events with the PM. In Aim 2, we will examine the contribution of DENND10 to Cu excretion. DENND10 is a CCC-binding protein of unknown function. We find that a proportion of DENND10 is localized on late endolysosomes and that DENND10 deficiency leads to an accumulation of ATP7B in these vesicles, indicating that DENND10 participates in regulating ATP7B trafficking. Altogether, the studies proposed in this project will close key knowledge gaps in Cu metabolism. More broadly, this work explores an unappreciated intersection between the CCC complex and the regulation of late endolysosomal vesicles, with potential implications to multiple other fields.