Additional file 3: of ERK1 and ERK2 present functional redundancy in tetrapods despite higher evolution rate of ERK1
Year: 2015
Type: article
Abstract: A. The epitope of the antibody targeting active ERK is conserved from sea anemone to human. Alignment of ERK1/2 residues which surround the sites phosphorylated by MEK kinase. The sequence HTGFLTRYVAT corresponds to the epitope recognized by the monoclonal anti-phosho ERK antibody from Sigma (#M8159). Phosphorylation sites, threonine, and tyrosine are indicated by arrows and are highlighted in yellow. ERK1 list in red, ERK2 list in blue, and single ERK list in black. Species names, sequences, accession numbers and common names are presented in that order. Note: yeast sequences of the two ERK orthologous genes, Fus3 and Kss1 are presented to illustrate that the phospho-ERK epitope is not fully conserved in yeast, unlike in sea anemone for example. B: Three distinct antibodies recognize phosphorylated ERK in extracts from mouse, lizard and hagfish. 10 % acrylamide gels were loaded with protein samples from mouse fibroblasts NIH3T3 or lizard embryo fibroblasts (A. sageii and A. carolinensis) or hagfish brain extracts as indicated in the figure (top). Mouse fibroblasts were pre-treated for one hour with 20 μM UO126 prior cell lysis to block activation of the ERK signaling cascade (lane1), or were stimulated with 10 % serum in presence of 1 mM sodium orthovanadate (NaVO4--) for one hour to block phosphatases and activate maximally the ERK signaling cascade (lanes 2). Lizard fibroblasts, from A. carolinensis (lane 3) or A. sagreii (lane 4) were stimulated as mouse fibroblasts of lane 2. One gel was stained with coomassie blue (i), other gels were transferred onto nitrocellulose membrane as described in materials and methods and immunoblots performed by incubating in the following antibodies ii) Anti phospho-ERKs sigma #M7802 iii) Anti phospho-ERKs Cell signaling #9101 iv) Anti phospho-ERKs Abcam #32538 v) Anti total-ERKs from Cell-Signaling #9102 and vi) Anti total-ERKs homemade E1B [10]. (PDF 2233 kb)
Source: Figshare
Language: English
Topic: Melanoma and MAPK Pathways
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W7136193786