Limits...
Hierarchy of Notch-Delta interactions promoting T cell lineage commitment and maturation.

Besseyrias V, Fiorini E, Strobl LJ, Zimber-Strobl U, Dumortier A, Koch U, Arcangeli ML, Ezine S, Macdonald HR, Radtke F - J. Exp. Med. (2007)

Bottom Line: However, N2-DL1-mediated signaling does not allow further T cell maturation beyond the CD25(+) stage due to a lack of T cell receptor beta expression.Moreover, comparative binding studies show preferential interaction of DL4 with N1, whereas binding of DL1 to N1 is weak.Collectively, our results establish a hierarchy of Notch-Delta interactions in which N1-DL4 exhibits the greatest capacity to induce and support T cell development.

View Article: PubMed Central - PubMed

Affiliation: Ludwig Institute for Cancer Research, Lausanne Branch, University of Lausanne, 1066 Epalinges, Switzerland.

ABSTRACT
Notch1 (N1) receptor signaling is essential and sufficient for T cell development, and recently developed in vitro culture systems point to members of the Delta family as being the physiological N1 ligands. We explored the ability of Delta1 (DL1) and DL4 to induce T cell lineage commitment and/or maturation in vitro and in vivo from bone marrow (BM) precursors conditionally gene targeted for N1 and/or N2. In vitro DL1 can trigger T cell lineage commitment via either N1 or N2. N1- or N2-mediated T cell lineage commitment can also occur in the spleen after short-term BM transplantation. However, N2-DL1-mediated signaling does not allow further T cell maturation beyond the CD25(+) stage due to a lack of T cell receptor beta expression. In contrast to DL1, DL4 induces and supports T cell commitment and maturation in vitro and in vivo exclusively via specific interaction with N1. Moreover, comparative binding studies show preferential interaction of DL4 with N1, whereas binding of DL1 to N1 is weak. Interestingly, preferential N1-DL4 binding reflects reduced dependence of this interaction on Lunatic fringe, a glycosyl transferase that generally enhances the avidity of Notch receptors for Delta ligands. Collectively, our results establish a hierarchy of Notch-Delta interactions in which N1-DL4 exhibits the greatest capacity to induce and support T cell development.

Show MeSH

Related in: MedlinePlus

Comparative in vivo analysis of DL1 and DL4 for their ability to promote ectopic T cell development. (A) Bar diagrams show percentages of GFP+ Ctrl and N1−/− BM cells after transduction with the control virus (MIG) and virus expressing DL1 and DL4. (B) Histograms show the percentage of GFP+ cells within total PBLs in host mice 9 wk after BM transplantation. Flow cytometric analysis for the presence of CD4- and CD8-expressing T cells was performed on PBLs (B) and BM and spleen (C) of host animals that were transplanted with WT and N1−/− BM cells expressing the indicated ligands.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2118717&req=5

fig8: Comparative in vivo analysis of DL1 and DL4 for their ability to promote ectopic T cell development. (A) Bar diagrams show percentages of GFP+ Ctrl and N1−/− BM cells after transduction with the control virus (MIG) and virus expressing DL1 and DL4. (B) Histograms show the percentage of GFP+ cells within total PBLs in host mice 9 wk after BM transplantation. Flow cytometric analysis for the presence of CD4- and CD8-expressing T cells was performed on PBLs (B) and BM and spleen (C) of host animals that were transplanted with WT and N1−/− BM cells expressing the indicated ligands.

Mentions: In vitro, DL4 is unable to induce T lineage commitment in the absence of a functional N1 receptor, suggesting that DL4 must specifically interact with N1 to specify the T lineage. To exclude that this observation is caused by a peculiarity of our in vitro culture system, we investigated the ability of DL1 and DL4 to induce T cell development in vivo in the presence and absence of N1. Previous studies demonstrated that retroviral overexpression of DL4 in hematopoietic cells is sufficient to promote thymus-independent T cell development to the DP stage in vivo (13, 23, 24). We therefore transduced CD45.2+ Ctrl and N1−/− BM cells with a retrovirus expressing either GFP alone (MIG), or DL1 or DL4 together with GFP, and subsequently transplanted these cells into lethally irradiated CD45.1+ C57BL/6 mice. The BM transduction efficiency of MIG and MIG expressing either DL1 or DL4 virus (based on GFP expression) was between 55 and 60% for both Ctrl and N1−/− BM cells (Fig. 8 A). Reconstituted hosts were analyzed 9 wk after transplantation for the presence of GFP+ donor-derived cells in PBLs. 72% of Ctrl PBLs and 64% of N1−/− PBLs in host mice that were transplanted with MIG-transduced BM cells were GFP+. Comparable percentages of PBLs were GFP+ in hosts receiving either DL1- or DL4-expressing Ctrl and N1−/− BM cells, indicating that the relative number of virus-expressing Ctrl and N1−/− donor cells was comparable even 9 wk after transplantation (Fig. 8 B, right). Only forced expression of DL4 but not DL1 resulted in the efficient development of DP T cells (Fig. 8, B–D). DL4-induced DP T cells were exclusively found in the PBLs (13%), BM (86%), and spleen (48%) of Ctrl but not of N1−/− chimeras, suggesting that enforced DL4 expression can only induce T cell development of N1-expressing progenitors in vivo. These results confirm our in vitro results using the DL4-expressing OP9 cells.


Hierarchy of Notch-Delta interactions promoting T cell lineage commitment and maturation.

Besseyrias V, Fiorini E, Strobl LJ, Zimber-Strobl U, Dumortier A, Koch U, Arcangeli ML, Ezine S, Macdonald HR, Radtke F - J. Exp. Med. (2007)

Comparative in vivo analysis of DL1 and DL4 for their ability to promote ectopic T cell development. (A) Bar diagrams show percentages of GFP+ Ctrl and N1−/− BM cells after transduction with the control virus (MIG) and virus expressing DL1 and DL4. (B) Histograms show the percentage of GFP+ cells within total PBLs in host mice 9 wk after BM transplantation. Flow cytometric analysis for the presence of CD4- and CD8-expressing T cells was performed on PBLs (B) and BM and spleen (C) of host animals that were transplanted with WT and N1−/− BM cells expressing the indicated ligands.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2118717&req=5

fig8: Comparative in vivo analysis of DL1 and DL4 for their ability to promote ectopic T cell development. (A) Bar diagrams show percentages of GFP+ Ctrl and N1−/− BM cells after transduction with the control virus (MIG) and virus expressing DL1 and DL4. (B) Histograms show the percentage of GFP+ cells within total PBLs in host mice 9 wk after BM transplantation. Flow cytometric analysis for the presence of CD4- and CD8-expressing T cells was performed on PBLs (B) and BM and spleen (C) of host animals that were transplanted with WT and N1−/− BM cells expressing the indicated ligands.
Mentions: In vitro, DL4 is unable to induce T lineage commitment in the absence of a functional N1 receptor, suggesting that DL4 must specifically interact with N1 to specify the T lineage. To exclude that this observation is caused by a peculiarity of our in vitro culture system, we investigated the ability of DL1 and DL4 to induce T cell development in vivo in the presence and absence of N1. Previous studies demonstrated that retroviral overexpression of DL4 in hematopoietic cells is sufficient to promote thymus-independent T cell development to the DP stage in vivo (13, 23, 24). We therefore transduced CD45.2+ Ctrl and N1−/− BM cells with a retrovirus expressing either GFP alone (MIG), or DL1 or DL4 together with GFP, and subsequently transplanted these cells into lethally irradiated CD45.1+ C57BL/6 mice. The BM transduction efficiency of MIG and MIG expressing either DL1 or DL4 virus (based on GFP expression) was between 55 and 60% for both Ctrl and N1−/− BM cells (Fig. 8 A). Reconstituted hosts were analyzed 9 wk after transplantation for the presence of GFP+ donor-derived cells in PBLs. 72% of Ctrl PBLs and 64% of N1−/− PBLs in host mice that were transplanted with MIG-transduced BM cells were GFP+. Comparable percentages of PBLs were GFP+ in hosts receiving either DL1- or DL4-expressing Ctrl and N1−/− BM cells, indicating that the relative number of virus-expressing Ctrl and N1−/− donor cells was comparable even 9 wk after transplantation (Fig. 8 B, right). Only forced expression of DL4 but not DL1 resulted in the efficient development of DP T cells (Fig. 8, B–D). DL4-induced DP T cells were exclusively found in the PBLs (13%), BM (86%), and spleen (48%) of Ctrl but not of N1−/− chimeras, suggesting that enforced DL4 expression can only induce T cell development of N1-expressing progenitors in vivo. These results confirm our in vitro results using the DL4-expressing OP9 cells.

Bottom Line: However, N2-DL1-mediated signaling does not allow further T cell maturation beyond the CD25(+) stage due to a lack of T cell receptor beta expression.Moreover, comparative binding studies show preferential interaction of DL4 with N1, whereas binding of DL1 to N1 is weak.Collectively, our results establish a hierarchy of Notch-Delta interactions in which N1-DL4 exhibits the greatest capacity to induce and support T cell development.

View Article: PubMed Central - PubMed

Affiliation: Ludwig Institute for Cancer Research, Lausanne Branch, University of Lausanne, 1066 Epalinges, Switzerland.

ABSTRACT
Notch1 (N1) receptor signaling is essential and sufficient for T cell development, and recently developed in vitro culture systems point to members of the Delta family as being the physiological N1 ligands. We explored the ability of Delta1 (DL1) and DL4 to induce T cell lineage commitment and/or maturation in vitro and in vivo from bone marrow (BM) precursors conditionally gene targeted for N1 and/or N2. In vitro DL1 can trigger T cell lineage commitment via either N1 or N2. N1- or N2-mediated T cell lineage commitment can also occur in the spleen after short-term BM transplantation. However, N2-DL1-mediated signaling does not allow further T cell maturation beyond the CD25(+) stage due to a lack of T cell receptor beta expression. In contrast to DL1, DL4 induces and supports T cell commitment and maturation in vitro and in vivo exclusively via specific interaction with N1. Moreover, comparative binding studies show preferential interaction of DL4 with N1, whereas binding of DL1 to N1 is weak. Interestingly, preferential N1-DL4 binding reflects reduced dependence of this interaction on Lunatic fringe, a glycosyl transferase that generally enhances the avidity of Notch receptors for Delta ligands. Collectively, our results establish a hierarchy of Notch-Delta interactions in which N1-DL4 exhibits the greatest capacity to induce and support T cell development.

Show MeSH
Related in: MedlinePlus