Proteomic analysis of the cortisol-mediated stress response in THP-1 monocytes using DIGE technology.
The glucocorticoid (GC) cortisol, the main mediator of the hypothalamic-pituitary-adrenal axis has many implications in metabolism, stress response and the immune system. Its function is mediated via binding to the glucocorticoid receptor (GR), a member of the superfamily of ligand-activated nuclear hormone receptors. The activity of the ligated GR results from its binding as a transcription factor to glucocorticoid response elements (GREs). Two-dimensional gel electrophoresis with DIGE (fluorescence difference gel electrophoresis) technology was applied to study the effects of cortisol on the human THP-1 monocytic cell line. A total of 28 cortisol-modulated proteins were identified belonging to five functional groups: cytoskeleton (8), chaperones (9), immune response (4), metabolism (3) and transcription/translation (4). Their corresponding genes were screened for putative GREs in their + 10 kb/- 0.2 kb promoter regions including all alternative promoters available within the Database for Transcription Start Sites (DBTSS). FKBP51, known to be induced by cortisol, was identified as the strongest differentially expressed protein, and contains the highest number of strict GREs. Genomic analysis of five alternative FKBP5 promoter regions suggests GC inducibility of all transcripts. Additionally, proteomics (2D DIGE and 2D immunoblotting) revealed the existence of several FKBP51 isoforms, which were not previously described. To our knowledge this is the first proteomic study that addresses the effects of cortisol on immune cells. FKBP51 isoforms found on the gel map were linked to alternative promoter usage on the genetic level, successfully correlating both the specific proteomic and genomic findings.
Billing
Anja M
AM
Institute of Immunology, National Public Health Laboratory, 20A, rue Auguste Lumière, L-1950 Luxembourg, Grand Duchy of Luxembourg.
Fack
Fred
F
Renaut
Jenny
J
Olinger
Christophe M
CM
Schote
Andrea B
AB
Turner
Jonathan D
JD
Muller
Claude P
CP
D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D015536:Down-Regulation / Q000187:drug effects; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D015870:Gene Expression / Q000187:drug effects; D005938:Glucocorticoids / Q000494:pharmacology; D006801:Humans; D006854:Hydrocortisone / Q000494:pharmacology*; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism; D009000:Monocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D010449:Peptide Mapping; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000235:genetics; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D020543:Proteome / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D022021:Tacrolimus Binding Proteins / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects; D000099159:Tacrolimus Binding Protein 5
D005938:Glucocorticoids; D006854:Hydrocortisone; D008840:Microfilament Proteins; D020033:Protein Isoforms; D020543:Proteome; D022021:Tacrolimus Binding Proteins; D000099159:Tacrolimus Binding Protein 5; C047262:LCP1 protein, human