Mucosa-specific targets for regulation of IFN-gamma expression: lamina propria T cells use different cis-elements than peripheral blood T cells to regulate transactivation of IFN-gamma expression.
Activation of lamina propria (LP) T cells via the CD2 pathway enhances IFN-gamma (IFN-gamma) secretion with further enhancement after CD28 coligation. The molecular mechanisms regulating IFN-gamma expression in LP T cells remain unknown. Previous studies in PBL and T cell lines identified cis- and trans-regulatory elements in TCR-mediated expression of IFN-gamma. This study examines CD2 and PMA/ionophore-responsive IFN-gamma promoter elements. Activation of LPMC via CD2-induced IFN-gamma secretion and a parallel up-regulation of mRNA expression. CD28 coligation enhanced mRNA stability without up-regulating transcription as measured by nuclear run-on. Transfection of a -2.7-kb IFN-gamma promoter-reporter construct into PBL and LP mononuclear cells (LPMC) revealed significant promoter activity after CD2 activation, with additional transactivation after CD2/CD28 costimulation in PBL, but not in LPMC. Functional analysis using truncated promoter fragments identified distinct cis-regulatory regions selectively transactivating IFN-gamma expression in PBL compared with LPMC. In PBL, CD2 activation elements reside within the -108- to +64-bp region. However, in LPMC the upstream region between -204 and -108 bp was essential. Transfection of the proximal and distal AP-1-binding elements, as well as TRE/AP-1 constructs, revealed functional activation of AP-1 subsequent to CD2 signaling, with activation critical in PBL but diminished in LPMC. Electromobility shift analysis using oligonucleotides encompassing the proximal, distal, and BED/AP-1-binding regions failed to demonstrate selective transactivation after CD2 signaling of LPMC. This report provides evidence that activation of LPMC results in transactivation of multiple promoter elements regulating IFN-gamma expression distinct from those in PBL.
Gonsky
R
R
Inflammatory Bowel Disease Research Center, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA.
Deem
R L
RL
Bream
J H
JH
Lee
D H
DH
Young
H A
HA
Targan
S R
SR
D020121:5' Untranslated Regions / Q000276:immunology; D000276:Adjuvants, Immunologic / Q000502:physiology; D018801:CD2 Antigens / Q000502:physiology; D018106:CD28 Antigens / Q000502:physiology; D017124:Conserved Sequence; D006801:Humans; D007371:Interferon-gamma / Q000096:biosynthesis* / Q000235:genetics; D007413:Intestinal Mucosa / Q000166:cytology / Q000276:immunology* / Q000378:metabolism*; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000378:metabolism; D008213:Lymphocyte Activation; D011401:Promoter Regions, Genetic / Q000276:immunology; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D012333:RNA, Messenger / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000276:immunology*; D015398:Signal Transduction / Q000276:immunology; D016176:T-Lymphocyte Subsets / Q000276:immunology* / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000378:metabolism; D015533:Transcriptional Activation / Q000276:immunology*; D015854:Up-Regulation / Q000235:genetics / Q000276:immunology
D020121:5' Untranslated Regions; D000276:Adjuvants, Immunologic; D018801:CD2 Antigens; D018106:CD28 Antigens; D012333:RNA, Messenger; D018808:Transcription Factor AP-1; D007371:Interferon-gamma