Nature Structural &Molecular Biology最新文献

The Seh1-associated complex (SEAC; GATOR in mammals) transduces amino acid signals to the Target of Rapamycin Complex 1 (TORC1), a master regulator of cell growth. The SEAC is composed of two subcomplexes, SEACIT (GATOR1), an inhibitor of TORC1 that has GAP activity against Gtr1, and SEACAT (GATOR2), which appears to regulate SEACIT. However, the molecular details of this regulation are unclear. Here we determined the cryo-electron microscopy structure of the SEAC bound to its substrate, the EGOC (Ragulator-Rag), and studied its function in TORC1 amino acid signaling. A single SEAC can interact with two EGOC molecules via SEACIT, binding exclusively to the 'active' version of the EGOC, without involvement of SEACAT. The GAP activity of the SEACIT is essential for the regulation of TORC1 by amino acids and its loss phenocopies the lack of Gtr1-Gtr2, establishing the SEAC-EGOC complex as an amino acid-sensing hub. Compared to other SEACAT subunits, the loss of Sea2, or its N-terminal β-propeller domain, yielded strong defects in amino acid signaling to TORC1. Our results suggest that the Sea2 β-propeller recruits a GAP inhibitor to mediate fast amino acid signaling to TORC1, with additional pathways acting with slower kinetics.

Short linear motifs (SLiMs) are the most ubiquitous protein interaction motifs within unstructured regions of the human proteome, yet their contribution to cellular homeostasis remains poorly understood. Here, to systematically assess SLiM function, we applied base editing to mutate all reported and a set of computationally predicted SLiMs defined by SLiM-like evolutionary patterns. By screening 7,293 SLiM-containing regions with 80,473 mutations in HAP1 cells, we define a SLiM dependency map identifying 450 reported and 264 predicted SLiMs required for normal cell proliferation. Mutational consequences were highly reproducible in RPE1 cells, with differences attributed to cell-line-specific gene essentiality. We show that many predicted SLiMs affecting proliferation do not belong to existing classes and identify binding partners for several of these, providing mechanistic insight into a disease-associated ANKRD17 mutation. Our study provides a proteome-wide resource on SLiM essentiality uncovering numerous uncharacterized essential SLiMs.

Mitochondria dynamically adapt to cellular stress to ensure cell survival. The stress-regulated mitochondrial peptidase OMA1 orchestrates these adaptive responses, which limit mitochondrial fusion and promote mitochondrial stress signaling and metabolic rewiring. Here, we show that cellular stress adaptation involves OMA1-mediated regulation of mitochondrial protein import and OXPHOS biogenesis. OMA1 cleaves the mitochondrial chaperone DNAJC15 and promotes its degradation by the m-AAA protease AFG3L2. Loss of DNAJC15 impairs mitochondrial protein import and restricts OXPHOS biogenesis under conditions of mitochondrial dysfunction. Non-imported mitochondrial preproteins accumulate at the endoplasmic reticulum, inducing an unfolded protein response. Our results demonstrate stress-dependent changes in mitochondrial protein import as part of the OMA1-mediated mitochondrial stress response and highlight the interdependence of proteostasis regulation between different organelles.

Cells use transcription-coupled nucleotide excision repair (TC-NER) to efficiently resolve transcription-blocking DNA lesions caused by genotoxic stress such as ultraviolet (UV) irradiation. However, UV also induces RNA damage, triggering a cytoplasmic ribotoxic stress response (RSR). Whether and how RSR affects nuclear TC-NER has remained unclear. Here we identify INTS12, a flexible, poorly characterized subunit of the Integrator complex, as a key mediator linking RSR to TC-NER. Specifically, RSR-activated ZAK signaling induces phosphorylation of INTS12, enhancing its interaction with CSB and promoting recruitment of the Integrator complex to lesion-stalled RNA polymerase II (Pol II). This facilitates Pol II clearance and enables efficient DNA repair through TC-NER. Disruption of this pathway compromises TC-NER and transcription recovery, thereby increasing cellular sensitivity to UV-induced damage. Notably, the requirement for INTS12-mediated Pol II removal is context dependent, as it is not advantageous during the transcription-coupled response to formaldehyde-induced DNA-protein crosslinks, which rely on a distinct proteasome-dependent degradation pathway. Together, these findings uncover a regulatory axis connecting RNA damage signaling to DNA repair and highlight a context-dependent role of INTS12 in maintaining genome integrity.