Identification of anisomycin-activated kinases p45 and p55 in murine cells as MAPKAP kinase-2.
We recently showed that EGF and anisomycin activate two kinases, p45 and p55, whose distinguishing feature is that their detection in in-gel kinase assays is enhanced by copolymerised poly-Glu/Tyr or poly-Glu/Phe (Cano E, Hazzalin CA and Mahadevan LC, Mol. Cell. Biol., 20:117-121). Their activation characteristics and sizes are strikingly similar to those of JNK/SAPKs, which are also strongly activated by anisomycin. However, we show here that p45 and p55 are not JNK/SAPKs but murine forms of MAPKAP kinase-2 because: (i) Detection of immunoprecipitated JNK/SAPKs is completely dependent on the presence of c-Jun as substrate in the in-gel kinase assays, whereas detection of p45 and p55 is not. (ii) Detection of p45 and p55 activity is enhanced by the presence of poly-Glu/Tyr or poly-Glu/Phe, whereas JNK/SAPKs are not detectable under these conditions. (iii) Although the sizes of the murine JNK/SAPKs and MAPKAP K-2 are similar, human JNK/SAPKs migrate at 45 and 55 kDa whereas human MAPKAP K-2 migrates at 50 kDa; the poly-Glu/Tyr-enhanced activity in human cells migrates at 50 KDa. (iv) Purified rabbit muscle MAPKAP K-2 is detectable as two bands of activity on in-gel kinase assays and their detection is enhanced by poly-Glu/Tyr. (v) Finally, the anisomycin-activated poly-Glu/Tyr-enhanced p45 and p55 kinases can be immunoprecipitated from murine cells using an anti-MAPKAP K-2 antibody. Thus, EGF- and anisomycin-activated p45 and p55 are not JNK/SAPKs but MAPKAP K-2, implying that both these agents activate the p38/RK MAP kinase cascade.
Cano
E
E
Nuclear Signalling Laboratory, Randall Institute, King's College, London, U.K.
Doza
Y N
YN
Ben-Levy
R
R
Cohen
P
P
Mahadevan
L C
LC
D000595:Amino Acid Sequence; D000818:Animals; D000841:Anisomycin / Q000494:pharmacology*; D000906:Antibodies; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000302:isolation & purification / Q000378:metabolism; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006801:Humans; D015151:Immunoblotting; D036341:Intercellular Signaling Peptides and Proteins; D047908:Intracellular Signaling Peptides and Proteins; D048031:JNK Mitogen-Activated Protein Kinases; D051379:Mice; D020928:Mitogen-Activated Protein Kinases; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D013379:Substrate Specificity; D000099190:MAP-Kinase-Activated Kinase 2
D000841:Anisomycin; D000906:Antibodies; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D036341:Intercellular Signaling Peptides and Proteins; D047908:Intracellular Signaling Peptides and Proteins; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D010455:Peptides; D010750:Phosphoproteins; D017346:Protein Serine-Threonine Kinases; D000099190:MAP-Kinase-Activated Kinase 2; C027606:L-glutamic acid-L-tyrosine copolymer