Abstract
The immunosuppressive cytokines transforming growth factor β1 (TGFβ1) and interleukin-10 (IL-10) regulate a variety of biological processes including differentiation, proliferation, tissue repair, tumorigenesis, inflammation, and host defense. Aberrant expression of TGFβ1 and IL-10 has been associated with many types of autoimmune and inflammatory disorders, as well as with many types of cancer and leukemia. Patients with cutaneous T cell lymphoma (CTCL) have high levels of malignant CD4+ T cells expressing IL-10 and TGFβ1 that suppress the immune system and diminish the antitumor responses. The transcriptional regulation of TGFβ1 and IL-10 expression is orchestrated by several transcription factors, including NFκB. However, while the transcriptional regulation of pro-inflammatory and anti-apoptotic genes by NFκB has been studied extensively, much less is known about the NFκB regulation of immunosuppressive genes. In this chapter, we describe a protocol that uses chromatin immunoprecipitation (ChIP) to analyze the transcriptional regulation of TGFβ1 and IL-10 by measuring recruitment of NFκB p65, p50, c-Rel, Rel-B, and p52 subunits to TGFβ1 and IL-10 promoters in human CTCL Hut-78 cells.
Publication types
-
Research Support, N.I.H., Extramural
MeSH terms
-
CD4-Positive T-Lymphocytes / metabolism*
-
CD4-Positive T-Lymphocytes / pathology
-
Chromatin Immunoprecipitation
-
Humans
-
Interleukin-10 / genetics*
-
Interleukin-10 / metabolism
-
Lymphoma, T-Cell, Cutaneous / genetics*
-
Lymphoma, T-Cell, Cutaneous / metabolism
-
Lymphoma, T-Cell, Cutaneous / pathology
-
NF-kappa B p50 Subunit / genetics
-
NF-kappa B p50 Subunit / metabolism
-
NF-kappa B p52 Subunit / genetics
-
NF-kappa B p52 Subunit / metabolism
-
Promoter Regions, Genetic
-
Proto-Oncogene Proteins c-rel / genetics
-
Proto-Oncogene Proteins c-rel / metabolism
-
Real-Time Polymerase Chain Reaction
-
Signal Transduction
-
Transcription Factor RelA / genetics
-
Transcription Factor RelA / metabolism
-
Transcription Factor RelB / genetics
-
Transcription Factor RelB / metabolism
-
Transcription, Genetic
-
Transforming Growth Factor beta1 / genetics*
-
Transforming Growth Factor beta1 / metabolism
-
Tumor Cells, Cultured
Substances
-
IL10 protein, human
-
NF-kappa B p50 Subunit
-
NF-kappa B p52 Subunit
-
Proto-Oncogene Proteins c-rel
-
RELA protein, human
-
RELB protein, human
-
Transcription Factor RelA
-
Transforming Growth Factor beta1
-
Interleukin-10
-
Transcription Factor RelB