Limits...
Mind bomb 1 in the lymphopoietic niches is essential for T and marginal zone B cell development.

Song R, Kim YW, Koo BK, Jeong HW, Yoon MJ, Yoon KJ, Jun DJ, Im SK, Shin J, Kong MP, Kim KT, Yoon K, Kong YY - J. Exp. Med. (2008)

Bottom Line: In addition, the endocytosis of Dll1 was impaired in the Mib1- microenvironment.Moreover, the block in T cell development and the failure of Dll1 endocytosis were also observed in coculture system by Mib1 knockdown.Our study reveals that Mib1 is the essential E3 ligase in T and MZB cell development, through the regulation of Notch ligands in the thymic and splenic microenvironments.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Science, Pohang University of Science and Technology, Pohang, Kyungbuk 790-784, South Korea.

ABSTRACT
Notch signaling regulates lineage decisions at multiple stages of lymphocyte development, and Notch activation requires the endocytosis of Notch ligands in the signal-sending cells. Four E3 ubiquitin ligases, Mind bomb (Mib) 1, Mib2, Neuralized (Neur) 1, and Neur2, regulate the Notch ligands to activate Notch signaling, but their roles in lymphocyte development have not been defined. We show that Mib1 regulates T and marginal zone B (MZB) cell development in the lymphopoietic niches. Inactivation of the Mib1 gene, but not the other E3 ligases, Mib2, Neur1, and Neur2, abrogated T and MZB cell development. Reciprocal bone marrow (BM) transplantation experiments revealed that Mib1 in the thymic and splenic niches is essential for T and MZB cell development. Interestingly, when BM cells from transgenic Notch reporter mice were transplanted into Mib1- mice, the Notch signaling was abolished in the double-negative thymocytes. In addition, the endocytosis of Dll1 was impaired in the Mib1- microenvironment. Moreover, the block in T cell development and the failure of Dll1 endocytosis were also observed in coculture system by Mib1 knockdown. Our study reveals that Mib1 is the essential E3 ligase in T and MZB cell development, through the regulation of Notch ligands in the thymic and splenic microenvironments.

Show MeSH

Related in: MedlinePlus

Impaired T cell development in the thymi of Mib1 conditional KO mice. (A and B) Thymocytes from ∼8–10-wk-old littermate controls (left) and the mutant (right) mice indicated were stained for CD4 and CD8 (A) and CD19 and B220 gated on CD4−CD8− DN thymocytes (B) and were analyzed by flow cytometry. Percentages of each population are indicated in the quadrants. Numbers are mean ± SD from 10 independent experiments. (C) Thymocytes from the indicated mice were analyzed by flow cytometry for the expression of CD44 and CD25 gated on CD4−CD8− DN thymocytes. Percentages indicated are mean ± SD from five independent experiments. (D) Absolute cell numbers for total thymocytes and thymocyte subsets of the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were calculated and are shown as histograms. The error bars represent mean ± SD from five independent experiments. *, P < 0.01. (E and F) The Mx1-Cre;Mib1+/f and Mx1-Cre;Mib1f/f mice were injected i.p. four times at 2-d intervals with 300 μg pIpC. At ∼8–14 wk after the final injection, the thymocytes were analyzed by flow cytometry for the expression of CD4 and CD8 (D) and CD19 and B220 (E) gated on CD4−CD8− DN thymocytes. Percentages indicated are mean ± SD from five independent experiments.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC2571928&req=5

fig2: Impaired T cell development in the thymi of Mib1 conditional KO mice. (A and B) Thymocytes from ∼8–10-wk-old littermate controls (left) and the mutant (right) mice indicated were stained for CD4 and CD8 (A) and CD19 and B220 gated on CD4−CD8− DN thymocytes (B) and were analyzed by flow cytometry. Percentages of each population are indicated in the quadrants. Numbers are mean ± SD from 10 independent experiments. (C) Thymocytes from the indicated mice were analyzed by flow cytometry for the expression of CD44 and CD25 gated on CD4−CD8− DN thymocytes. Percentages indicated are mean ± SD from five independent experiments. (D) Absolute cell numbers for total thymocytes and thymocyte subsets of the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were calculated and are shown as histograms. The error bars represent mean ± SD from five independent experiments. *, P < 0.01. (E and F) The Mx1-Cre;Mib1+/f and Mx1-Cre;Mib1f/f mice were injected i.p. four times at 2-d intervals with 300 μg pIpC. At ∼8–14 wk after the final injection, the thymocytes were analyzed by flow cytometry for the expression of CD4 and CD8 (D) and CD19 and B220 (E) gated on CD4−CD8− DN thymocytes. Percentages indicated are mean ± SD from five independent experiments.

Mentions: To determine which E3 ligase is required for T cell development, flow cytometric analyses were performed on the thymocytes from the MMTV-Cre;Mib1f/f, Mib2−/−, Neur1−/−, and Neur2−/− mice. Unexpectedly, the thymocyte development, as well as the percentages and numbers of CD4 single-positive (SP), CD8 SP, CD4+CD8+ DP, and TCRγδ T and NK cells in Mib2−/−, Neur1−/−, Neur2−/−, and even Neur1−/−;Neur2−/− mice was comparable to that of WT controls (Fig. 2 A and not depicted), demonstrating that Neur1, Neur2, and Mib2 are dispensable for T cell development. In contrast, T cell development was severely affected in the MMTV-Cre;Mib1f/f mice. The percentage of DP thymocytes was dramatically reduced, whereas the percentage of CD4−CD8− double-negative (DN) cells was relatively increased (Fig. 2 A). In addition, the percentage of B220+ cells was increased ∼7.8-fold in the CD4−CD8− DN thymocytes from the MMTV-Cre;Mib1f/f mice, as compared with the MMTV-Cre;Mib1+/f mice (Fig. 2 B). The absolute numbers of total thymocytes were reduced 3.6-fold in the MMTV-Cre;Mib1f/f mice as compared with those of the control mice. The CD4 SP, CD8 SP, and CD4CD8 DP thymocytes were also reduced 2.7-fold, 2.1-fold, and 4.3-fold, respectively, whereas the DN cells remained relatively unaffected (Fig. 2 C). Because most of the DN cells of the MMTV-Cre;Mib1f/f mice are B cells, DN numbers appear to be unaffected because of the increase in the number of B cells rather than a DN to DP transition defect.


Mind bomb 1 in the lymphopoietic niches is essential for T and marginal zone B cell development.

Song R, Kim YW, Koo BK, Jeong HW, Yoon MJ, Yoon KJ, Jun DJ, Im SK, Shin J, Kong MP, Kim KT, Yoon K, Kong YY - J. Exp. Med. (2008)

Impaired T cell development in the thymi of Mib1 conditional KO mice. (A and B) Thymocytes from ∼8–10-wk-old littermate controls (left) and the mutant (right) mice indicated were stained for CD4 and CD8 (A) and CD19 and B220 gated on CD4−CD8− DN thymocytes (B) and were analyzed by flow cytometry. Percentages of each population are indicated in the quadrants. Numbers are mean ± SD from 10 independent experiments. (C) Thymocytes from the indicated mice were analyzed by flow cytometry for the expression of CD44 and CD25 gated on CD4−CD8− DN thymocytes. Percentages indicated are mean ± SD from five independent experiments. (D) Absolute cell numbers for total thymocytes and thymocyte subsets of the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were calculated and are shown as histograms. The error bars represent mean ± SD from five independent experiments. *, P < 0.01. (E and F) The Mx1-Cre;Mib1+/f and Mx1-Cre;Mib1f/f mice were injected i.p. four times at 2-d intervals with 300 μg pIpC. At ∼8–14 wk after the final injection, the thymocytes were analyzed by flow cytometry for the expression of CD4 and CD8 (D) and CD19 and B220 (E) gated on CD4−CD8− DN thymocytes. Percentages indicated are mean ± SD from five independent experiments.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2571928&req=5

fig2: Impaired T cell development in the thymi of Mib1 conditional KO mice. (A and B) Thymocytes from ∼8–10-wk-old littermate controls (left) and the mutant (right) mice indicated were stained for CD4 and CD8 (A) and CD19 and B220 gated on CD4−CD8− DN thymocytes (B) and were analyzed by flow cytometry. Percentages of each population are indicated in the quadrants. Numbers are mean ± SD from 10 independent experiments. (C) Thymocytes from the indicated mice were analyzed by flow cytometry for the expression of CD44 and CD25 gated on CD4−CD8− DN thymocytes. Percentages indicated are mean ± SD from five independent experiments. (D) Absolute cell numbers for total thymocytes and thymocyte subsets of the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were calculated and are shown as histograms. The error bars represent mean ± SD from five independent experiments. *, P < 0.01. (E and F) The Mx1-Cre;Mib1+/f and Mx1-Cre;Mib1f/f mice were injected i.p. four times at 2-d intervals with 300 μg pIpC. At ∼8–14 wk after the final injection, the thymocytes were analyzed by flow cytometry for the expression of CD4 and CD8 (D) and CD19 and B220 (E) gated on CD4−CD8− DN thymocytes. Percentages indicated are mean ± SD from five independent experiments.
Mentions: To determine which E3 ligase is required for T cell development, flow cytometric analyses were performed on the thymocytes from the MMTV-Cre;Mib1f/f, Mib2−/−, Neur1−/−, and Neur2−/− mice. Unexpectedly, the thymocyte development, as well as the percentages and numbers of CD4 single-positive (SP), CD8 SP, CD4+CD8+ DP, and TCRγδ T and NK cells in Mib2−/−, Neur1−/−, Neur2−/−, and even Neur1−/−;Neur2−/− mice was comparable to that of WT controls (Fig. 2 A and not depicted), demonstrating that Neur1, Neur2, and Mib2 are dispensable for T cell development. In contrast, T cell development was severely affected in the MMTV-Cre;Mib1f/f mice. The percentage of DP thymocytes was dramatically reduced, whereas the percentage of CD4−CD8− double-negative (DN) cells was relatively increased (Fig. 2 A). In addition, the percentage of B220+ cells was increased ∼7.8-fold in the CD4−CD8− DN thymocytes from the MMTV-Cre;Mib1f/f mice, as compared with the MMTV-Cre;Mib1+/f mice (Fig. 2 B). The absolute numbers of total thymocytes were reduced 3.6-fold in the MMTV-Cre;Mib1f/f mice as compared with those of the control mice. The CD4 SP, CD8 SP, and CD4CD8 DP thymocytes were also reduced 2.7-fold, 2.1-fold, and 4.3-fold, respectively, whereas the DN cells remained relatively unaffected (Fig. 2 C). Because most of the DN cells of the MMTV-Cre;Mib1f/f mice are B cells, DN numbers appear to be unaffected because of the increase in the number of B cells rather than a DN to DP transition defect.

Bottom Line: In addition, the endocytosis of Dll1 was impaired in the Mib1- microenvironment.Moreover, the block in T cell development and the failure of Dll1 endocytosis were also observed in coculture system by Mib1 knockdown.Our study reveals that Mib1 is the essential E3 ligase in T and MZB cell development, through the regulation of Notch ligands in the thymic and splenic microenvironments.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Science, Pohang University of Science and Technology, Pohang, Kyungbuk 790-784, South Korea.

ABSTRACT
Notch signaling regulates lineage decisions at multiple stages of lymphocyte development, and Notch activation requires the endocytosis of Notch ligands in the signal-sending cells. Four E3 ubiquitin ligases, Mind bomb (Mib) 1, Mib2, Neuralized (Neur) 1, and Neur2, regulate the Notch ligands to activate Notch signaling, but their roles in lymphocyte development have not been defined. We show that Mib1 regulates T and marginal zone B (MZB) cell development in the lymphopoietic niches. Inactivation of the Mib1 gene, but not the other E3 ligases, Mib2, Neur1, and Neur2, abrogated T and MZB cell development. Reciprocal bone marrow (BM) transplantation experiments revealed that Mib1 in the thymic and splenic niches is essential for T and MZB cell development. Interestingly, when BM cells from transgenic Notch reporter mice were transplanted into Mib1- mice, the Notch signaling was abolished in the double-negative thymocytes. In addition, the endocytosis of Dll1 was impaired in the Mib1- microenvironment. Moreover, the block in T cell development and the failure of Dll1 endocytosis were also observed in coculture system by Mib1 knockdown. Our study reveals that Mib1 is the essential E3 ligase in T and MZB cell development, through the regulation of Notch ligands in the thymic and splenic microenvironments.

Show MeSH
Related in: MedlinePlus