Dynamic regulation of notch 1 and notch 2 surface expression during T cell development and activation revealed by novel monoclonal antibodies.
It is well established that Notch signaling plays a critical role at multiple stages of T cell development and activation. However, detailed analysis of the cellular and molecular events associated with Notch signaling in T cells is hampered by the lack of reagents that can unambiguously measure cell surface Notch receptor expression. Using novel rat mAbs directed against the extracellular domains of Notch1 and Notch2, we find that Notch1 is already highly expressed on common lymphoid precursors in the bone marrow and remains at high levels during intrathymic maturation of CD4(-)CD8(-) thymocytes. Notch1 is progressively down-regulated at the CD4(+)CD8(+) and mature CD4(+) or CD8(+) thymic stages and is expressed at low levels on peripheral T cells. Immunofluorescence staining of thymus cryosections further revealed a localization of Notch1(+)CD25(-) cells adjacent to the thymus capsule. Notch1 was up-regulated on peripheral T cells following activation in vitro with anti-CD3 mAbs or infection in vivo with lymphocytic chorio-meningitis virus or Leishmania major. In contrast to Notch1, Notch2 was expressed at intermediate levels on common lymphoid precursors and CD117(+) early intrathymic subsets, but disappeared completely at subsequent stages of T cell development. However, transient up-regulation of Notch2 was also observed on peripheral T cells following anti-CD3 stimulation. Collectively our novel mAbs reveal a dynamic regulation of Notch1 and Notch2 surface expression during T cell development and activation. Furthermore they provide an important resource for future analysis of Notch receptors in various tissues including the hematopoietic system.
Fiorini
Emma
E
Ludwig Institute for Cancer Research Ltd., Lausanne Branch, 1066 Epalinges, Switzerland.
Merck
Estelle
E
Wilson
Anne
A
Ferrero
Isabel
I
Jiang
Wei
W
Koch
Ute
U
Auderset
Floriane
F
Laurenti
Elisa
E
Tacchini-Cottier
Fabienne
F
Pierres
Michel
M
Radtke
Freddy
F
Luther
Sanjiv A
SA
Macdonald
H Robson
HR
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D002454:Cell Differentiation; D002462:Cell Membrane / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D008213:Lymphocyte Activation / Q000276:immunology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D051381:Rats; D051881:Receptor, Notch1 / Q000096:biosynthesis* / Q000276:immunology; D051883:Receptor, Notch2 / Q000096:biosynthesis* / Q000276:immunology; D015398:Signal Transduction / Q000276:immunology; D013234:Stem Cells / Q000378:metabolism; D013601:T-Lymphocytes / Q000166:cytology / Q000378:metabolism
D000911:Antibodies, Monoclonal; C497763:Notch1 protein, rat; C497766:Notch2 protein, rat; D051881:Receptor, Notch1; D051883:Receptor, Notch2