Abstract
Calnexin was initially identified as an endoplasmic reticulum (ER) type I integral membrane protein, phosphorylated on its cytosolic domain by ER-associated protein kinases. Although the role of the ER luminal domain of calnexin has been established as a constituent of the molecular chaperone machinery of the ER, less is known about the role of the cytosolic phosphorylation of calnexin. Analysis by two-dimensional phosphopeptide maps revealed that calnexin was in vitro phosphorylated in isolated microsomes by casein kinase 2 (CK2) and extracellular-signal regulated kinase-1 (ERK-1) at sites corresponding to those for in vivo phosphorylation. In canine pancreatic microsomes, synergistic phosphorylation by CK2 and ERK-1 led to increased association of calnexin with membrane-bound ribosomes. In vivo, calnexin-associated ERK-1 activity was identified by co-immunoprecipitation. This activity was abolished in cells expressing a dominant-negative MEK-1. Activation of ERK-1 in cells by addition of serum led to a 4-fold increase in ribosome-associated calnexin over unstimulated cells. Taken together with studies revealing calnexin association with CK2 and ERK-1, a model is proposed whereby phosphorylation of calnexin leads to a potential increase in glycoprotein folding close to the translocon.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Blood Proteins / pharmacology
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Calcium-Binding Proteins / metabolism*
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Calcium-Calmodulin-Dependent Protein Kinases / metabolism*
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Calnexin
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Casein Kinase II
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Cell Line
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Cytosol / metabolism
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Dogs
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Endoplasmic Reticulum, Rough / drug effects
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Endoplasmic Reticulum, Rough / enzymology
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Endoplasmic Reticulum, Rough / metabolism
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Enzyme Activation / drug effects
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Intracellular Membranes / drug effects
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Intracellular Membranes / enzymology
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Intracellular Membranes / metabolism
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MAP Kinase Kinase 1
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Microsomes / drug effects
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Microsomes / enzymology
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Microsomes / metabolism
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Mitogen-Activated Protein Kinase 3
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Mitogen-Activated Protein Kinase Kinases*
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Mitogen-Activated Protein Kinases*
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Pancreas / cytology
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Phosphorylation
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Precipitin Tests
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Protein Binding / drug effects
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism*
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Protein-Tyrosine Kinases / genetics
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Protein-Tyrosine Kinases / metabolism
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Rats
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Recombinant Fusion Proteins / metabolism
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Ribosomes / drug effects
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Ribosomes / metabolism*
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Serine / metabolism
Substances
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Blood Proteins
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Calcium-Binding Proteins
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Recombinant Fusion Proteins
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Calnexin
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Serine
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Protein-Tyrosine Kinases
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Casein Kinase II
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Protein Serine-Threonine Kinases
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Calcium-Calmodulin-Dependent Protein Kinases
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Mitogen-Activated Protein Kinase 3
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Mitogen-Activated Protein Kinases
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MAP Kinase Kinase 1
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Mitogen-Activated Protein Kinase Kinases