The unique membrane organization of the rod outer segment (ROS), the specialized sensory cilium of rod photoreceptor cells, provides the foundation for phototransduction, the initial step in vision. ROS architecture is characterized by a stack of identically shaped and tightly packed membrane disks loaded with the visual receptor rhodopsin. A wide range of genetic aberrations have been reported to compromise ROS ultrastructure, impairing photoreceptor viability and function. Yet, the structural basis giving rise to the remarkably precise arrangement of ROS membrane stacks and the molecular mechanisms underlying genetically inherited diseases remain elusive. Here, cryo-electron tomography (cryo-ET) performed on native ROS at molecular resolution provides insights into key structural determinants of ROS membrane architecture. Our data confirm the existence of two previously observed molecular connectors/spacers which likely contribute to the nanometer-scale precise stacking of the ROS disks. We further provide evidence that the extreme radius of curvature at the disk rims is enforced by a continuous supramolecular assembly composed of peripherin-2 (PRPH2) and rod outer segment membrane protein 1 (ROM1) oligomers. We suggest that together these molecular assemblies constitute the structural basis of the highly specialized ROS functional architecture. Our Cryo-ET data provide novel quantitative and structural information on the molecular architecture in ROS and substantiate previous results on proposed mechanisms underlying pathologies of certain PRPH2 mutations leading to blindness.
The subvolume averages of ROS disk rims are deposited in EMDB under the accession codes: EMD-13321, EMD-13322, EMD-13323 and EMD-13324.Two representative 4x binned tomograms for each acquisition scheme or mouse strain were deposited in EMPIAR under the accession codes: EMPIAR-10771, EMPIAR-10772 and EMPIAR-10773
- Krzysztof Palczewski
- Krzysztof Palczewski
- Matthias Pöge
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: The animals used were bred for scientific purposes. At the University of California, Irvine, all mice were housed in the vivarium where they were maintained on a normal mouse chow diet and a 12 h / 12 h light / dark cycle. All animal procedures were approved by the Institutional Animal Care and Use Committee (IACUC, AUP-21-096) of the University of California, Irvine, and were conducted in accordance with the Association for Research in Vision and Ophthalmology Statement for the Use of Animals in Ophthalmic and Visual Research. The research at the Max Planck Institute of Biochemistry did not involve experiments on an animal. All animals were sacrificed prior to the removal of organs in accordance with the European Commission Recommendations for the euthanasia of experimental animals (Part 1 and Part 2). Breeding and housing as well as the euthanasia of the animals are fully compliant with all German (e.g. German Animal Welfare Act) and EC (e.g. Directive 86/609/EEC) applicable laws and regulations concerning care and use of laboratory animals. The Max Planck Institute of Biochemistry has a licence for breeding and housing of Iaboratory animals which includes the killing of animals solely for the use of their organs or tissues (No.4.3.2-5682/MPI für Biochemie - rural districts office).
- Giulia Zanetti, Institute of Structural and Molecular Biology, Birkbeck, University of London, United Kingdom
© 2021, Pöge 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.
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