MAP Kinase Signaling ProtocolsRony Seger Mitogen-activated protein kinase (MAPK) signaling cascades are a group of protein kinases that play a central role in the intracellular transmission of extracellular signals. These cascades operate as major lines of communication within a complicated signaling network that regulates many cellular processes, including proliferation, differentiation, development, stress response, and apoptosis. More than 15,000 papers on MAPKs have been published over the past few years, with the number of publications increasing each year. More and more laboratories embark on the study of MAPK cascades in many d- tinct cellular systems and in particular their role in disease. Future challenges in the study of MAPK cascades remain in understa- ing the role of the various components and isoforms of the cascades in the multiple critical functions that they regulate in the whole organism, as well as the diseases caused by their malfunction. Data from gene-disrupted mice s- gest that inhibition of the MAPK cascades may have serious consequences on the development and growth of the animals. For example, targeted deletion of MEK1 is lethal, owing to developmental problems of placental vasculature and abnormal fibroblast migration. This lethality occurs in spite of the normal expression of MEK2, indicating that although the two MEK isoforms are apparently similar, they do have distinct functions, at least during embryog- esis. The ERK cascade was also shown to play a central role in brain function and in learning and memory. |
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Contents
The ERK Cascade As a Prototype of MAPK Signaling Pathways | 1 |
AntiphosphoERK Antibodies | 29 |
Detection of ERK12 Activities Using Affinity Reagents | 49 |
Kyriakis Hong Liu and Deborah N Chadee | 61 |
Investigating the Cellular BMK1ERK5 Signaling Pathway | 89 |
PullDown Assays for Guanosine 5TriphosphateBound RasLike | 97 |
Regulation of MAPK Cascades by Protein Tyrosine Phosphatases | 103 |
Use of Inhibitors in the Study of MAPK Signaling | 113 |
Makoto Adachi and Eisuke Nishida 11 Study of MAPK Signaling Using Knockout Mice | 145 |
Aguda and Herbert M Sauro 13 Signaling by Growth Factor Receptors | 167 |
Rikiro Fukunaga and Tony Hunter 16 Study of Substrate Specificity of MAPKs | 211 |
Michael B Yaffe 17 Ras Signaling Pathway for Analysis | 237 |
Ami Aronheim | 251 |
and Natalie G Ahn 20 Use of Xenopus Oocytes and Early Embryos | 283 |
Lessons From Learning | 299 |
323 | |
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Common terms and phrases
agarose amino acids antibody aprotinin beads binding Biol BMK1 c-Jun cDNA cellular centrifuge Chem clones column containing detection diluted domain electrophoresis EMBO enzyme ERK cascade ERK1 ERK2 expression extracellular extracellular signal-regulated kinase extract filters gene growth factor GST-PTP-SL HEPES immunoblot immunoprecipitation Incubate inhibitor interaction IPTG isoforms kinase activity kinase assay kinase kinase ligand lysis buffer mammalian MAP Kinase Signaling MAP3K MAPK activation MAPK cascades medium MEK1 MEKK1 MEKs membrane Methods mg/mL mitogen-activated protein kinase molecular motif mutant N-terminal NaCl nitrocellulose nuclear oocytes p38MAPK peptide library phage phosphatases phosphorylation plasmid plates PMSF prepared PTP-SL purified reaction reagents receptor recombinant regulation residues room temperature SAPK/JNK screening SDS-PAGE Seger sequence serum Sigma signaling pathways solution specific stimulation Subheading substrate supernatant tein tion transcription factor transfection transfer Tris-HCl tube tyrosine ubiquitination vector vitro washing buffer Western blot