scholarly journals Cryo-EM reconstructions of inhibitor-bound SMG1 kinase reveal an autoinhibitory state dependent on SMG8

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Lukas M Langer ◽  
Fabien Bonneau ◽  
Yair Gat ◽  
Elena Conti
Keyword(s):  
Small Molecule ◽  
Molecular Basis ◽  
Biochemical Analysis ◽  
Kinase Activity ◽  
Regulatory Mechanisms ◽  
Cryo Electron Microscopy ◽  
State Dependent ◽  
Molecule Inhibitor ◽  
Nonsense Mediated Mrna Decay ◽  
Complex Regulatory Network

The PI3K-related kinase (PIKK) SMG1 monitors progression of metazoan nonsense-mediated mRNA decay (NMD) by phosphorylating the RNA helicase UPF1. Previous work has shown that the activity of SMG1 is impaired by small molecule inhibitors, is reduced by the SMG1 interactors SMG8 and SMG9, and is downregulated by the so-called SMG1 insertion domain. However, the molecular basis for this complex regulatory network has remained elusive. Here, we present cryo-electron microscopy reconstructions of human SMG1-9 and SMG1-8-9 complexes bound to either a SMG1 inhibitor or a non-hydrolyzable ATP analogue at overall resolutions ranging from 2.8 to 3.6 Å. These structures reveal the basis with which a small molecule inhibitor preferentially targets SMG1 over other PIKKs. By comparison with our previously reported substrate-bound structure (Langer et al. 2020), we show that the SMG1 insertion domain can exert an autoinhibitory function by directly blocking the substrate binding path as well as overall access to the SMG1 kinase active site. Together with biochemical analysis, our data indicate that SMG1 autoinhibition is stabilized by the presence of SMG8. Our results explain the specific inhibition of SMG1 by an ATP-competitive small molecule, provide insights into regulation of its kinase activity within the NMD pathway, and expand the understanding of PIKK regulatory mechanisms in general.

scholarly journals Cryo-EM reconstructions of inhibitor-bound SMG1 kinase reveal an autoinhibitory state dependent on SMG8

2021 ◽  
Author(s):  
Lukas M Langer ◽  
Fabien Bonneau ◽  
Yair Gat ◽  
Elena Conti
Keyword(s):  
Small Molecule ◽  
Molecular Basis ◽  
Biochemical Analysis ◽  
Kinase Activity ◽  
Regulatory Mechanisms ◽  
Cryo Electron Microscopy ◽  
State Dependent ◽  
Molecule Inhibitor ◽  
Nonsense Mediated Mrna Decay ◽  
Complex Regulatory Network

The PI3K-related kinase (PIKK) SMG1 monitors progression of metazoan nonsense-mediated mRNA decay (NMD) by phosphorylating the RNA helicase UPF1. Previous work has shown that the activity of SMG1 is impaired by small molecule inhibitors, is reduced by the SMG1 interactors SMG8 and SMG9, and is downregulated by the so-called SMG1 insertion domain. However, the molecular basis for this complex regulatory network has remained elusive. Here, we present cryo-electron microscopy reconstructions of human SMG1-9 and SMG1-8-9 complexes bound to either a SMG1 inhibitor or a non-hydrolyzable ATP analogue at overall resolutions ranging from 2.8 to 3.6 Å. These structures reveal the basis with which a small molecule inhibitor preferentially targets SMG1 over other PIKKs. By comparison with our previously reported substrate-bound structure (Langer et al. 2020), we show that the SMG1 insertion domain can exert an autoinhibitory function by directly blocking the substrate binding path as well as overall access to the SMG1 kinase active site. Together with biochemical analysis, our data indicate that SMG1 autoinhibition is stabilized by the presence of SMG8. Our results explain the specific inhibition of SMG1 by an ATP-competitive small molecule, provide insights into regulation of its kinase activity within the NMD pathway, and expand the understanding of PIKK regulatory mechanisms in general.

scholarly journals CDK10 in Gastrointestinal Cancers: Dual Roles as a Tumor Suppressor and Oncogene

2021 ◽  
Vol 11 ◽  
Author(s):  
Zainab A. Bazzi ◽  
Isabella T. Tai
Keyword(s):  
Cell Proliferation ◽  
Tumor Suppressor ◽  
Small Molecule ◽  
High Throughput Screening ◽  
Kinase Activity ◽  
Screening Assay ◽  
Cellular Processes ◽  
Molecule Inhibitor ◽  
High Throughput Screening Assay ◽  
Kinase Activity Assay

Cyclin-dependent kinase 10 (CDK10) is a CDC2-related serine/threonine kinase involved in cellular processes including cell proliferation, transcription regulation and cell cycle regulation. CDK10 has been identified as both a candidate tumor suppressor in hepatocellular carcinoma, biliary tract cancers and gastric cancer, and a candidate oncogene in colorectal cancer (CRC). CDK10 has been shown to be specifically involved in modulating cancer cell proliferation, motility and chemosensitivity. Specifically, in CRC, it may represent a viable biomarker and target for chemoresistance. The development of therapeutics targeting CDK10 has been hindered by lack a specific small molecule inhibitor for CDK10 kinase activity, due to a lack of a high throughput screening assay. Recently, a novel CDK10 kinase activity assay has been developed, which will aid in the development of small molecule inhibitors targeting CDK10 activity. Discovery of a small molecular inhibitor for CDK10 would facilitate further exploration of its biological functions and affirm its candidacy as a therapeutic target, specifically for CRC.

scholarly journals The role of Lyn kinase in the development of imatinib resistance in chronic myelogenous leukemia

2011 ◽  
Vol 2 (2) ◽  
pp. 67
Author(s):  
Camillo Porta ◽  
Federica Tagliani
Keyword(s):  
Cost Effectiveness ◽  
Chronic Myelogenous Leukemia ◽  
Small Molecule ◽  
Kinase Activity ◽  
Myelogenous Leukemia ◽  
Tyrosine Kinase Activity ◽  
Imatinib Resistance ◽  
Lyn Kinase ◽  
Molecule Inhibitor

Imatinib mesylate, a small-molecule inhibitor of BCRABL tyrosine kinase activity, has emerged as the well-recognized standard of treatment for chronic myelogenous leukemia (CML). Indeed, both its efficacy, tolerability, as well as cost-effectiveness have been clearly proven...

scholarly journals The role of Lyn kinase in the development of imatinib resistance in chronic myelogenous leukemia

2011 ◽  
pp. 67-68
Author(s):  
Camillo Porta ◽  
Federica Tagliani
Keyword(s):  
Cost Effectiveness ◽  
Chronic Myelogenous Leukemia ◽  
Small Molecule ◽  
Kinase Activity ◽  
Myelogenous Leukemia ◽  
Tyrosine Kinase Activity ◽  
Imatinib Resistance ◽  
Lyn Kinase ◽  
Molecule Inhibitor

Imatinib mesylate, a small-molecule inhibitor of BCRABL tyrosine kinase activity, has emerged as the well-recognized standard of treatment for chronic myelogenous leukemia (CML). Indeed, both its efficacy, tolerability, as well as cost-effectiveness have been clearly proven...

scholarly journals Structure of substrate-bound SMG1-8-9 kinase complex reveals molecular basis for phosphorylation specificity

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Lukas M Langer ◽  
Yair Gat ◽  
Fabien Bonneau ◽  
Elena Conti
Keyword(s):  
Active Site ◽  
Molecular Basis ◽  
Phosphorylation Site ◽  
Hydrophobic Residue ◽  
Cellular Processes ◽  
Cryo Electron Microscopy ◽  
Biochemical Assays ◽  
Rna Quality Control ◽  
Nonsense Mediated Mrna Decay ◽  
Site Recognition

PI3K-related kinases (PIKKs) are large Serine/Threonine (Ser/Thr)-protein kinases central to the regulation of many fundamental cellular processes. PIKK family member SMG1 orchestrates progression of an RNA quality control pathway, termed nonsense-mediated mRNA decay (NMD), by phosphorylating the NMD factor UPF1. Phosphorylation of UPF1 occurs in its unstructured N- and C-terminal regions at Serine/Threonine-Glutamine (SQ) motifs. How SMG1 and other PIKKs specifically recognize SQ motifs has remained unclear. Here, we present a cryo-electron microscopy (cryo-EM) reconstruction of a human SMG1-8-9 kinase complex bound to a UPF1 phosphorylation site at an overall resolution of 2.9 Å. This structure provides the first snapshot of a human PIKK with a substrate-bound active site. Together with biochemical assays, it rationalizes how SMG1 and perhaps other PIKKs specifically phosphorylate Ser/Thr-containing motifs with a glutamine residue at position +1 and a hydrophobic residue at position -1, thus elucidating the molecular basis for phosphorylation site recognition.

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