Purification and cDNA cloning of SAPKK3, the major activator of RK/p38 in stress- and cytokine-stimulated monocytes and epithelial cells.
Two chromatographically distinct stress-activated protein kinase kinases (SAPKKs) have been identified in several mammalian cells, termed SAPKK2 and SAPKK3, which activate the MAP kinase family member RK/p38 but not JNK/SAPK in vitro. Here we demonstrate that SAPKK2 is identical or very closely related to the MAP kinase kinase family member MKK3. However, under our assay conditions, SAPKK3 was the major activator of RK/p38 detected in extracts prepared from stress- or interleukin-1-stimulated epithelial (KB) cells, from bacterial lipopolysaccharide and tumour necrosis factor alpha-stimulated THP1 monocytes or from rabbit skeletal muscle. The activated form of SAPKK3 was purified from muscle to near homogeneity, and tryptic peptide sequences were used to clone human and murine cDNAs encoding this enzyme. Human SAPKK3 comprised 334 amino acids and was 78% identical to MKK3. The murine and human SAPKK3 were 97% identical in their amino acid sequences. We also cloned a different murine cDNA that appears to encode a SAPKK3 protein truncated at the N-terminus. SAPKK3 is identical to the recently cloned MKK6.
Cuenda
A
A
MRC Protein Phosphorylation Unit, Department of Biochemistry, University of Dundee, UK.
Alonso
G
G
Morrice
N
N
Jones
M
M
Meier
R
R
Cohen
P
P
Nebreda
A R
AR
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism / Q000502:physiology; D002478:Cells, Cultured; D004789:Enzyme Activation; D004848:Epithelium / Q000187:drug effects* / Q000378:metabolism; D005796:Genes; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D007624:KB Cells; D007948:Leukemia, Monocytic, Acute / Q000473:pathology; D008070:Lipopolysaccharides / Q000494:pharmacology; D048371:MAP Kinase Kinase 3; D048669:MAP Kinase Kinase 6; D008322:Mammals; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases; D008969:Molecular Sequence Data; D009000:Monocytes / Q000187:drug effects* / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism; D009363:Neoplasm Proteins / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000235:genetics; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D011817:Rabbits; D016415:Sequence Alignment; D013312:Stress, Physiological / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D048051:p38 Mitogen-Activated Protein Kinases
D007375:Interleukin-1; D008070:Lipopolysaccharides; D009363:Neoplasm Proteins; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048371:MAP Kinase Kinase 3; D048669:MAP Kinase Kinase 6; C482530:MAP2K3 protein, human; C482566:MAP2K6 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases