TCR/CD28-stimulated actin dynamics are required for NFAT1-mediated transcription of c-rel leading to CD28 response element activation.
TCR/CD28 engagement triggers the initiation of a variety of signal transduction pathways that lead to changes in gene transcription. Although reorganization of the actin cytoskeleton is required for T cell activation, the molecular pathways controlled by the actin cytoskeleton are ill defined. To this end, we analyzed TCR/CD28-stimulated signaling pathways in cytochalasin D-treated T cells to determine the cytoskeletal requirements for T cell activation. Cytochalasin D treatment impaired T cell activation by causing a reduction in TCR/CD28-mediated calcium flux, and blocked activation of two regulatory elements within the IL-2 promoter, NFAT/AP-1 and CD28RE/AP. Treatment had no effect on signaling leading to the activation of either AP-1 or NF-kappaB. Significantly, we found that NFAT1 is required for optimal c-rel up-regulation in response to TCR/CD28 stimulation. In fact, NFAT1 could be detected bound at the c-rel promoter in response to TCR/CD28 stimulation, and targeting of NFAT1 using RNA interference in human CD4(+) T cells abrogated c-rel transcription. Overall, these findings establish that disrupting actin cytoskeletal dynamics impairs TCR/CD28-mediated calcium flux required for NFAT1-mediated c-rel transcription and, thus, activation of the CD28RE/AP.
Nolz
Jeffrey C
JC
Department of Immunology, Mayo Clinic College of Medicine, MN 55905, USA.
Fernandez-Zapico
Martin E
ME
Billadeau
Daniel D
DD
D000199:Actins / Q000187:drug effects / Q000378:metabolism*; D001483:Base Sequence; D015153:Blotting, Western; D018106:CD28 Antigens / Q000235:genetics*; D002118:Calcium / Q000378:metabolism; D003572:Cytochalasins / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D019169:Jurkat Cells; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology*; D008969:Molecular Sequence Data; D050778:NFATC Transcription Factors / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D020569:Proto-Oncogene Proteins c-rel / Q000235:genetics* / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism; D014158:Transcription, Genetic
D000199:Actins; D018106:CD28 Antigens; D003572:Cytochalasins; D007376:Interleukin-2; D050778:NFATC Transcription Factors; C494006:NFATC2 protein, human; D020569:Proto-Oncogene Proteins c-rel; D011948:Receptors, Antigen, T-Cell; C031582:cytochalasin C; D002118:Calcium