TY - JOUR
T1 - Cloning of a novel phosphatidylinositol kinase-related kinase. Characterization of the human SMG-1 RNA surveillance protein
AU - Denning, Gabriela
AU - Jamieson, Lee
AU - Maquat, Lynne E.
AU - Thompson, E. Aubrey
AU - Fields, Alan P.
PY - 2001/6/22
Y1 - 2001/6/22
N2 - We have cloned and characterized a new member of the phosphatidylinositol kinase (PIK)-related kinase family. This gene, which we term human SMG-1 (hSMG-1), is orthologous to Caenorhabditis elegans SMG-1, a protein that functions in nonsense-mediated mRNA decay (NMD). cDNA sequencing revealed that hSMG-1 encodes a protein of 3031 amino acids containing a conserved kinase domain, a C-terminal domain unique to the PIK-related kinases and an FKBP12-rapamycin binding-like domain similar to that found in the PIK-related kinase mTOR. Immunopurified FLAG-tagged hSMG-1 exhibits protein kinase activity as measured by autophosphorylation and phosphorylation of the generic PIK-related kinase substrate PHAS-1. hSMG-1 kinase activity is inhibited by high nanomolar concentrations of wortmannin (IC50 = 105 nM) but is not inhibited by a FKBP12-rapamycin complex. Mutation of conserved residues within the kinase domain of hSMG-1 abolishes both autophosphorylation and substrate phosphorylation, demonstrating that hSMG-1 exhibits intrinsic protein kinase activity. hSMG-1 phosphorylates purified hUpf1 protein, a phosphoprotein that plays a critical role in NMD, at sites that are also phosphorylated in whole cells. Based on these data, we conclude that hSMG-1 is the human orthologue to C. elegans SMG-1. Our data indicate that hSMG-1 may function in NMD by directly phosphorylating hUpf1 protein at physiologically relevant sites.
AB - We have cloned and characterized a new member of the phosphatidylinositol kinase (PIK)-related kinase family. This gene, which we term human SMG-1 (hSMG-1), is orthologous to Caenorhabditis elegans SMG-1, a protein that functions in nonsense-mediated mRNA decay (NMD). cDNA sequencing revealed that hSMG-1 encodes a protein of 3031 amino acids containing a conserved kinase domain, a C-terminal domain unique to the PIK-related kinases and an FKBP12-rapamycin binding-like domain similar to that found in the PIK-related kinase mTOR. Immunopurified FLAG-tagged hSMG-1 exhibits protein kinase activity as measured by autophosphorylation and phosphorylation of the generic PIK-related kinase substrate PHAS-1. hSMG-1 kinase activity is inhibited by high nanomolar concentrations of wortmannin (IC50 = 105 nM) but is not inhibited by a FKBP12-rapamycin complex. Mutation of conserved residues within the kinase domain of hSMG-1 abolishes both autophosphorylation and substrate phosphorylation, demonstrating that hSMG-1 exhibits intrinsic protein kinase activity. hSMG-1 phosphorylates purified hUpf1 protein, a phosphoprotein that plays a critical role in NMD, at sites that are also phosphorylated in whole cells. Based on these data, we conclude that hSMG-1 is the human orthologue to C. elegans SMG-1. Our data indicate that hSMG-1 may function in NMD by directly phosphorylating hUpf1 protein at physiologically relevant sites.
UR - http://www.scopus.com/inward/record.url?scp=0035933889&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0035933889&partnerID=8YFLogxK
U2 - 10.1074/jbc.C100144200
DO - 10.1074/jbc.C100144200
M3 - Article
C2 - 11331269
AN - SCOPUS:0035933889
SN - 0021-9258
VL - 276
SP - 22709
EP - 22714
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 25
ER -