Genetic and environmental factors, such as metals, interact to determine neurological traits. We reasoned that interactomes of molecules handling metals in neurons should include novel metal homeostasis pathways. We focused on copper and its transporter ATP7A because ATP7A null mutations cause neurodegeneration. We performed ATP7A immunoaffinity chromatography and identified 541 proteins co-isolating with ATP7A. The ATP7A interactome concentrated gene products implicated in neurodegeneration and neurodevelopmental disorders, including subunits of the Golgi-localized conserved oligomeric Golgi (COG) complex. COG null cells possess altered content and subcellular localization of ATP7A and CTR1 (SLC31A1), the transporter required for copper uptake, as well as decreased total cellular copper, and impaired copper-dependent metabolic responses. Changes in the expression of ATP7A and COG subunits in Drosophila neurons altered synapse development in larvae and copper-induced mortality of adult flies. We conclude that the ATP7A interactome encompasses a novel COG-dependent mechanism to specify neuronal development and survival.
ATP7A IP DSP treated cells after incubation with BCS or CuCl2Available at the PeptideAtlas database.
ATP7A IP Menkes and Rescued Menkes CellsAvailable at the PeptideAtlas database.
- Victor Faundez
- Michael Petris
- Vladimir Lupashin
- Dana Boyd Barr
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Andrew B West, University of Alabama at Birmingham, United States
© 2017, Comstra et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Proliferating cells undergo metabolic changes in synchrony with cell cycle progression and cell division. Mitochondria provide fuel, metabolites, and ATP during different phases of the cell cycle, however it is not completely understood how mitochondrial function and the cell cycle are coordinated. CLUH is a post-transcriptional regulator of mRNAs encoding mitochondrial proteins involved in oxidative phosphorylation and several metabolic pathways. Here, we show a role of CLUH in regulating the expression of astrin, which is involved in metaphase to anaphase progression, centrosome integrity, and mTORC1 inhibition. We find that CLUH binds both the SPAG5 mRNA and its product astrin, and controls the synthesis and the stability of the full-length astrin-1 isoform. We show that CLUH interacts with astrin-1 specifically during interphase. Astrin-depleted cells show mTORC1 hyperactivation and enhanced anabolism. On the other hand, cells lacking CLUH show decreased astrin levels and increased mTORC1 signaling, but cannot sustain anaplerotic and anabolic pathways. In absence of CLUH, cells fail to grow during G1, and progress faster through the cell cycle, indicating dysregulated matching of growth, metabolism and cell cycling. Our data reveal a role of CLUH in coupling growth signaling pathways and mitochondrial metabolism with cell cycle progression.
A developing understanding suggests that spatial compartmentalisation in pancreatic β cells is critical in controlling insulin secretion. To investigate the mechanisms, we have developed live-cell sub-cellular imaging methods using the mouse organotypic pancreatic slice. We demonstrate that the organotypic pancreatic slice, when compared with isolated islets, preserves intact β cell structure, and enhances glucose dependent Ca2+ responses and insulin secretion. Using the slice technique, we have discovered the essential role of local activation of integrins and the downstream component, focal adhesion kinase, in regulating β cells. Integrins and focal adhesion kinase are exclusively activated at the β cell capillary interface and using in situ and in vitro models we show their activation both positions presynaptic scaffold proteins, like ELKS and liprin, and regulates glucose dependent Ca2+ responses and insulin secretion. We conclude that focal adhesion kinase orchestrates the final steps of glucose dependent insulin secretion within the restricted domain where β cells contact the islet capillaries.