InsP6 binding to PIKK kinases revealed by the cryo-EM structure of an SMG1–SMG8–SMG9 complex

Gat, Yair; Schuller, Jan M.; Lingaraju, Mahesh; Weyher, Elisabeth; Bonneau, Fabien; Strauß, Michael; Murray, Peter J.; Conti, Elena

doi:10.1038/s41594-019-0342-7

Cited by 33 publications

(62 citation statements)

References 33 publications

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“…S14A). Notably, the mutations in I2 are equivalent to those reported previously to abolish completely the kinase activity of an N-terminally truncated 'naked' mTOR fragment toward a C-terminal peptide of Akt1 (47). A possible explanation for this 15 apparent discrepancy is provided by a reduced stability of mTORC2 assembled using the I2 variant (but not the I3 variant) (Fig.…”

Section: Cryo-em Reconstructions Of Variantssupporting

confidence: 63%

“…S17 and S18A-F). InsP6 binds in a region, 10 which is incomplete in related PI3Ks (46), but generally conserved in members of the PIKK family of kinases (47). Indeed, InsP6 was previously reported to associate with DNA-PKcs (48).…”

Section: Cryo-em Reconstructions Of Variantsmentioning

confidence: 99%

“…Indeed, InsP6 was previously reported to associate with DNA-PKcs (48). Recently, structure determination of the PIKK family pseudo-kinase SMG1 revealed InsP6 binding in a region corresponding to the I-site and led the authors to postulate a corresponding binding site in mTOR but involving both the kinase domain and FAT domain (47). InsP6 has previously been 5 observed as a structural component of multi-subunit assemblies, including the splicesome (49) and proteasome activator complex (50), and helical repeat regions have been identified as InsP6 interaction sites (51).…”

Section: Cryo-em Reconstructions Of Variantsmentioning

confidence: 99%

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The 3.2Å resolution structure of human mTORC2

Scaiola

Mangia

Imseng

et al. 2020

Preprint

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The protein kinase mammalian target of rapamycin (mTOR) is the central regulator of cell growth. Aberrant mTOR signaling is linked to cancer, diabetes and neurological disorders. mTOR exerts its functions in two distinct multiprotein complexes, mTORC1 and mTORC2. Here we report a 3.2 Å resolution cryo-EM reconstruction of mTORC2. It reveals entangled folds of the defining Rictor and the substrate-binding SIN1 subunits, identifies the C-terminal domain of Rictor as the source of the rapamycin insensitivity of mTORC2, and resolves mechanisms for mTORC2 regulation by complex destabilization. Two novel small molecule binding sites are visualized, an inositol hexakisphosphate (InsP6) pocket in mTOR and an mTORC2-specific nucleotide binding site in Rictor which also forms a zinc finger. Structural and biochemical analyses suggest that InsP6 and nucleotide binding do not control mTORC2 activity directly but rather have roles in folding or ternary interactions. These insights provide a firm basis for studying mTORC2 signaling and for developing mTORC2-specific inhibitors.

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Section: Cryo-em Reconstructions Of Variantssupporting

confidence: 63%

Section: Cryo-em Reconstructions Of Variantsmentioning

confidence: 99%

Section: Cryo-em Reconstructions Of Variantsmentioning

confidence: 99%

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The 3.2Å resolution structure of human mTORC2

Scaiola

Mangia

Imseng

et al. 2020

Preprint

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“…The inhibitory effects of this compound depends on the induction of MYC expression (it is shown to inhibit NMD) [104]. Mago-Y14-eIF4AIII-Barentsz-UPF3b 2xb2 3.4 Å X-Ray diffraction [108] Translation is necessary for NMD [107]; thus, it is suggested that inhibitors of translation may also be effective inhibitors of NMD [104]. Moreover, Martin et al have recently demonstrated that NMD inhibition can be achieved via other mechanisms [16,93] and also determined that 80% depletion of UPF1 can suppress NMD activity not influencing the proliferation or survival of cells [55,93].…”

Section: Development Of Nmd Inhibitorsmentioning

confidence: 99%

“…Three-dimensional protein structures can reveal the precise interactions defining the protein-protein interface for in silico drug design, which can be targeted for drug discovery [109][110][111]. These protein-protein structures available in the Protein Data Bank (http://www.rcsb.org/pdb) [111] for NMD pathway are ( Figure 2 and Table 1): UPF1-UPF2 [34], UPF2-UPF3b [36], SMG5-SMG7 [103,105,112], SMG8-SMG9 [104,106], SMG1-SMG8-SMG9 [107] and the EJC (Mago-Y14-eIF4AIII-Barentsz-UPF3b) [108]. Figure 2, illustrates different protein-protein or protein-RNA interactions from the NMD pathways that could represent a base or the platform to design inhibitors or peptide-like molecules.…”

Section: Development Of Nmd Inhibitorsmentioning

confidence: 99%

Nonsense-Mediated mRNA Decay: Pathologies and the Potential for Novel Therapeutics

Pawlicka

Kalathiya

Alfaro

2020

Cancers

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Nonsense-mediated messenger RNA (mRNA) decay (NMD) is a surveillance pathway used by cells to control the quality mRNAs and to fine-tune transcript abundance. NMD plays an important role in cell cycle regulation, cell viability, DNA damage response, while also serving as a barrier to virus infection. Disturbance of this control mechanism caused by genetic mutations or dys-regulation of the NMD pathway can lead to pathologies, including neurological disorders, immune diseases and cancers. The role of NMD in cancer development is complex, acting as both a promoter and a barrier to tumour progression. Cancer cells can exploit NMD for the downregulation of key tumour suppressor genes, or tumours adjust NMD activity to adapt to an aggressive immune microenvironment. The latter case might provide an avenue for therapeutic intervention as NMD inhibition has been shown to lead to the production of neoantigens that stimulate an immune system attack on tumours. For this reason, understanding the biology and co-option pathways of NMD is important for the development of novel therapeutic agents. Inhibitors, whose design can make use of the many structures available for NMD study, will play a crucial role in characterizing and providing diverse therapeutic options for this pathway in cancer and other diseases.

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Recessive, Deleterious Variants in SMG8 Expand the Role of Nonsense-Mediated Decay in Developmental Disorders in Humans

Alzahrani

Kuwahara

Long

et al. 2020

The American Journal of Human Genetics

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We have previously described a heart-, eye-, and brain-malformation syndrome caused by homozygous loss-of-function variants in SMG9, which encodes a critical component of the nonsense-mediated decay (NMD) machinery. Here, we describe four consanguineous families with four different likely deleterious homozygous variants in SMG8, encoding a binding partner of SMG9. The observed phenotype greatly resembles that linked to SMG9 and comprises severe global developmental delay, microcephaly, facial dysmorphism, and variable congenital heart and eye malformations. RNA-seq analysis revealed a general increase in mRNA expression levels with significant overrepresentation of core NMD substrates. We also identified increased phosphorylation of UPF1, a key SMG1-dependent step in NMD, which most likely represents the loss of SMG8-mediated inhibition of SMG1 kinase activity. Our data show that SMG8 and SMG9 deficiency results in overlapping developmental disorders that most likely converge mechanistically on impaired NMD.

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InsP6 binding to PIKK kinases revealed by the cryo-EM structure of an SMG1–SMG8–SMG9 complex

Cited by 33 publications

References 33 publications

The 3.2Å resolution structure of human mTORC2

The 3.2Å resolution structure of human mTORC2

Nonsense-Mediated mRNA Decay: Pathologies and the Potential for Novel Therapeutics

Recessive, Deleterious Variants in SMG8 Expand the Role of Nonsense-Mediated Decay in Developmental Disorders in Humans

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