The LINC complex mechanically couples cytoskeletal and nuclear components across the nuclear envelope to fulfil a myriad of cellular functions, including nuclear shape and positioning, hearing and meiotic chromosome movements. The canonical model is that 3:3 interactions between SUN and KASH proteins underlie the nucleocytoskeletal linkages provided by the LINC complex. Here, we provide crystallographic and biophysical evidence that SUN-KASH is a constitutive 6:6 complex in which two constituent 3:3 complexes interact head-to-head. A common SUN-KASH topology is achieved through structurally diverse 6:6 interaction mechanisms by distinct KASH proteins, including zinc-coordination by Nesprin-4. The SUN-KASH 6:6 interface provides a molecular mechanism for the establishment of integrative and distributive connections between 3:3 structures within a branched LINC complex network. In this model, SUN-KASH 6:6 complexes act as nodes for force distribution and integration between adjacent SUN and KASH molecules, enabling the coordinated transduction of large forces across the nuclear envelope.
Crystallographic structure factors and atomic coordinates have been deposited in the Protein Data Bank (PDB) under accession numbers 6R15, 6R16 and 6R2I, and raw diffraction data have been uploaded to https://proteindiffraction.org/. SEC-SAXS data have been deposited in the Small Angle Scattering Biological Data Bank (https://www.sasbdb.org/) under accession numbers SASDJC5, SASDJD5, SASDJE5 and SASDJF5. Uncropped gel images relating to Figures 4b and 5a are available in source data files.
- Owen Richard Davies
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Megan C King, Yale School of Medicine, United States
© 2021, Gurusaran & Davies
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.
In eukaryotic cells, stressors reprogram the cellular proteome by activating the integrated stress response (ISR). In its canonical form, stress-sensing kinases phosphorylate the eukaryotic translation initiation factor eIF2 (eIF2-P), which ultimately leads to reduced levels of ternary complex required for initiation of mRNA translation. Previously we showed that translational control is primarily exerted through a conformational switch in eIF2’s nucleotide exchange factor, eIF2B, which shifts from its active A-State conformation to its inhibited I-State conformation upon eIF2-P binding, resulting in reduced nucleotide exchange on eIF2 (Schoof et al. 2021). Here, we show functionally and structurally how a single histidine to aspartate point mutation in eIF2B’s β subunit (H160D) mimics the effects of eIF2-P binding by promoting an I-State like conformation, resulting in eIF2-P independent activation of the ISR. These findings corroborate our previously proposed A/I-State model of allosteric ISR regulation.
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