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In vivo identification of lymphocyte subsets exhibiting transcriptionally active NF-kappaB/Rel complexes
J Feuillard1, S Mémet, B Goudeau
1Unité de Biologie Moléculaire de l'Expression Génique, URA 1773 CNRS, Institut Pasteur, 28 rue du Dr Roux, 75724 Paris Cedex 15, France.
Insights
Nuclear factor kappa B (NF-kappaB) activity is crucial in immune cell development. This study reveals NF-kappaB/Rel complexes are key players in the in vivo differentiation of IgD(+) B lymphocytes and CD25(+) thymocytes.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- NF-kappaB/Rel signaling is vital for immune responses.
- Previous studies indicated limited NF-kappaB activity in lymphocytes.
- Transgenic mice with a lacZ reporter were generated to study NF-kappaB activity in vivo.
Purpose of the Study:
- To analyze the NF-kappaB/Rel activity pattern in primary and secondary lymphoid organs of living organisms.
- To re-examine NF-kappaB/Rel transcriptional activity in thymocytes and splenic lymphocytes using flow cytometry.
Main Methods:
- Generation of transgenic mice carrying a kappaB-dependent lacZ gene.
- In situ analysis of lymphoid organs.
- Flow cytometry using fluorescein-di-beta-D-galactopyranoside (FDG) as a vital substrate for beta-galactosidase.
Main Results:
- Constitutive NF-kappaB/Rel activity was detected in specific thymocyte subsets (CD44+CD25(-) and CD44(-)CD25(+)).
- NF-kappaB/Rel activity was found in most splenic B cells, particularly virgin IgD(+) B cells, but largely absent in T cells.
- Activity was also observed in antigen-presenting cells, some endothelial cells, and lymph node capsula lining cells.
Conclusions:
- NF-kappaB/Rel complexes play a significant role in the in vivo differentiation of IgD(+) B lymphocytes.
- NF-kappaB/Rel signaling may also be involved in the differentiation of CD25(+) thymocytes.
Abstract:
To analyze the NF-kappaB/Rel activity pattern in a living organism, we previously generated transgenic mice carrying a kappaB-dependent lacZ gene. In situ analysis of both primary and secondary lymphoid organs revealed a strong NF-kappaB transcriptional activity in antigen-presenting cells, some endothelial cells and sinus lining cells of the lymph node capsula with very little activity in lymphocytes and thymocytes. Using fluorescein-di-beta-D-galactopyranoside (FDG) as a vital substrate for the beta-galactosidase, we re-examined by flow cytometry the NF-kappaB/Rel transcriptional activity in our mouse model. We report here that such constitutive NF-kappaB/Rel activity was significantly detected in thymocytes at the CD44+CD25(-) stage. This constitutive activity extended with CD25 expression to the majority of the CD44(-)CD25(+) thymocytes and was then restricted to a few mature T cells. In the spleen, constitutive NF-kappaB/Rel activity was found in most B cells, unlike T cells which were largely negative. Virgin IgD(+) B cells expressed higher levels of NF-kappaB transcriptional activity than other B cell types. Altogether, these results suggest that NF-kappaB/Rel complexes are key players in the in vivo differentiation of IgD(+) B lymphocytes and possibly CD25(+) thymocytes.