The Med1 subunit of transcriptional mediator plays a central role in regulating CCAAT/enhancer-binding protein-beta-driven transcription in response to interferon-gamma.
Transcription factor CCAAT/enhancer-binding protein (C/EBP)-beta is crucial for regulating transcription of genes involved in a number of diverse cellular processes, including those involved in some cytokine-induced responses. However, the mechanisms that contribute to its diverse transcriptional activity are not yet fully understood. To gain an understanding into its mechanisms of action, we took a proteomic approach and identified cellular proteins that associate with C/EBP-beta in an interferon (IFN)-gamma-dependent manner. Transcriptional mediator (Mediator) is a multisubunit protein complex that regulates signal-induced cellular gene transcription from enhancer-bound transcription factor(s). Here, we report that the Med1 subunit of the Mediator as a C/EBP-beta-interacting protein. Using gene knock-out cells and mutational and RNA interference approaches, we show that Med1 is critical for IFN-induced expression of certain genes. Med1 associates with C/EBP-beta through a domain located between amino acids 125 and 155 of its N terminus. We also show that the MAPK, ERK1/2, and an ERK phosphorylation site within regulatory domain 2, more specifically the Thr(189) residue, of C/EBP-beta are essential for it to bind to Med1. Last, an ERK-regulated site in Med1 protein is also essential for up-regulating IFN-induced transcription although not critical for binding to C/EBP-beta.
Li
Hui
H
Department of Microbiology and Immunology, Greenebaum Cancer Center, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
Gade
Padmaja
P
Nallar
Shreeram C
SC
Raha
Abhijit
A
Roy
Sanjit K
SK
Karra
Sreenivasu
S
Reddy
Janardan K
JK
Reddy
Sekhar P
SP
Kalvakolanu
Dhananjaya V
DV
D000595:Amino Acid Sequence; D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D064550:Death-Associated Protein Kinases; D004706:Endodeoxyribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit / Q000378:metabolism; D007371:Interferon-gamma / Q000494:pharmacology*; D051379:Mice; D018345:Mice, Knockout; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014158:Transcription, Genetic / Q000187:drug effects* / Q000235:genetics*
D051017:Apoptosis Regulatory Proteins; D022782:CCAAT-Enhancer-Binding Protein-beta; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; C494228:Isgf3g protein, mouse; D021122:Protein Subunits; D011994:Recombinant Proteins; D007371:Interferon-gamma; D064550:Death-Associated Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D004706:Endodeoxyribonucleases; C118712:MBD4 protein, human; C492249:Mbd4 protein, mouse