Analysis of mitogen-activated protein kinase phosphorylation in response to stimulation of histidine kinase signaling pathways in Neurospora.
In eukaryotes, two-component regulatory systems have been demonstrated to regulate phosphorylation of mitogen-activated protein kinases (MAPKs). Here, we describe a method implementing preparation of a protein extract under denaturing conditions, followed by Western analysis using MAPK antibodies that can be used to observe the effects of components of two-component signaling pathways or other proteins on the phosphorylation status of MAPKs. The protein extraction method presented may also be used to concentrate cellular proteins for additional applications, such as metabolic labeling or analysis of other posttranslational modifications.
Jones
Carol A
CA
Department of Plant Pathology and Microbiology, University of California, Riverside, California, USA.
Borkovich
Katherine A
KA
D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D000071677:Histidine Kinase; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D009491:Neurospora / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology
D005656:Fungal Proteins; D011494:Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D000071677:Histidine Kinase