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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.

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Regulation of MZB cell development by Mib1 in the splenic microenvironment. (A) The lethally irradiated CD45.1 mice were injected i.v. with BM cells from the 12–15-wk-old MMTV-Cre;Mib1+/f (left) and MMTV-Cre;Mib1f/f (right) mice. At 12 wk after transplantation, the splenocytes were stained for the expression of CD21 and CD23 gated on the CD45.1+B220+ cells (top), CD1d and CD9 gated on the CD45.1+B220+ cells (middle), and CD21 and IgM gated on CD45.1+CD23− cells (bottom). Percentages indicated are mean ± SD from five independent experiments. (B) The lethally irradiated MMTV-Cre;Mib1+/f (left) and MMTV-Cre;Mib1f/f (right) mice (7–9 wk old) were transplanted with CD45.1 BM cells. At 5–6 wk after reconstitution with CD45.1 BM cells, the splenocytes were analyzed as in A. Percentages indicated are mean ± SD from five independent experiments. (C) Genomic DNA was prepared from the CD45− cells sorted from spleens and was analyzed by quantitative real-time PCR with deleted allele-specific primers. Data are mean ± SD from triplicate experiments. *, P < 0.0001.
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fig6: Regulation of MZB cell development by Mib1 in the splenic microenvironment. (A) The lethally irradiated CD45.1 mice were injected i.v. with BM cells from the 12–15-wk-old MMTV-Cre;Mib1+/f (left) and MMTV-Cre;Mib1f/f (right) mice. At 12 wk after transplantation, the splenocytes were stained for the expression of CD21 and CD23 gated on the CD45.1+B220+ cells (top), CD1d and CD9 gated on the CD45.1+B220+ cells (middle), and CD21 and IgM gated on CD45.1+CD23− cells (bottom). Percentages indicated are mean ± SD from five independent experiments. (B) The lethally irradiated MMTV-Cre;Mib1+/f (left) and MMTV-Cre;Mib1f/f (right) mice (7–9 wk old) were transplanted with CD45.1 BM cells. At 5–6 wk after reconstitution with CD45.1 BM cells, the splenocytes were analyzed as in A. Percentages indicated are mean ± SD from five independent experiments. (C) Genomic DNA was prepared from the CD45− cells sorted from spleens and was analyzed by quantitative real-time PCR with deleted allele-specific primers. Data are mean ± SD from triplicate experiments. *, P < 0.0001.

Mentions: Because Mib1 is expressed in both CD45+ and CD45− cells in the spleen (Fig. 1 B), we investigated whether Mib1 functions in the hematopoietic cells or in the microenvironments for MZB cell development. When the lethally irradiated CD45.1 WT mice were reconstituted with BM cells from either the MMTV-Cre;Mib1+/f or MMTV-Cre;Mib1f/f mice, all of the CD45.1 recipient mice displayed not only T1 B and T2 B cells but also normal MZB cell development (Fig. 6 A and not depicted), indicating that the impaired MZB cell development in the MMTV-Cre;Mib1f/f mice is not caused by an autonomous defect in hematopoietic cells. However, in the reciprocal BMT experiment, in which the lethally irradiated MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were reconstituted with BM cells from CD45.1 congenic mice, B220+CD21+CD23lo/− and B220+CD1d+CD9+ MZB cell development was completely blocked in the MMTV-Cre;Mib1f/f recipient mice, whereas it was unaffected in the control mice (Fig. 6 B). In addition to MZB cells, cycling T2 B cell development, but not T1 B cell development, was severely affected in the MMTV-Cre;Mib1f/f recipient mice (Fig. 6 B and not depicted). These results show that the impaired MZB cell development in the MMTV-Cre;Mib1f/f mice originated from a nonhematopoietic microenvironment. Consistent with these findings, Mib1 was deleted ∼90% in isolated CD45− splenic stromal cells from MMTV-Cre;Mib1f/f mice as compared with that of the control mice (Fig. 6 C), suggesting that Mib1 regulates MZB and cycling T2 B cell development in the splenic microenvironments.


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)

Regulation of MZB cell development by Mib1 in the splenic microenvironment. (A) The lethally irradiated CD45.1 mice were injected i.v. with BM cells from the 12–15-wk-old MMTV-Cre;Mib1+/f (left) and MMTV-Cre;Mib1f/f (right) mice. At 12 wk after transplantation, the splenocytes were stained for the expression of CD21 and CD23 gated on the CD45.1+B220+ cells (top), CD1d and CD9 gated on the CD45.1+B220+ cells (middle), and CD21 and IgM gated on CD45.1+CD23− cells (bottom). Percentages indicated are mean ± SD from five independent experiments. (B) The lethally irradiated MMTV-Cre;Mib1+/f (left) and MMTV-Cre;Mib1f/f (right) mice (7–9 wk old) were transplanted with CD45.1 BM cells. At 5–6 wk after reconstitution with CD45.1 BM cells, the splenocytes were analyzed as in A. Percentages indicated are mean ± SD from five independent experiments. (C) Genomic DNA was prepared from the CD45− cells sorted from spleens and was analyzed by quantitative real-time PCR with deleted allele-specific primers. Data are mean ± SD from triplicate experiments. *, P < 0.0001.
© Copyright Policy
Related In: Results  -  Collection

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fig6: Regulation of MZB cell development by Mib1 in the splenic microenvironment. (A) The lethally irradiated CD45.1 mice were injected i.v. with BM cells from the 12–15-wk-old MMTV-Cre;Mib1+/f (left) and MMTV-Cre;Mib1f/f (right) mice. At 12 wk after transplantation, the splenocytes were stained for the expression of CD21 and CD23 gated on the CD45.1+B220+ cells (top), CD1d and CD9 gated on the CD45.1+B220+ cells (middle), and CD21 and IgM gated on CD45.1+CD23− cells (bottom). Percentages indicated are mean ± SD from five independent experiments. (B) The lethally irradiated MMTV-Cre;Mib1+/f (left) and MMTV-Cre;Mib1f/f (right) mice (7–9 wk old) were transplanted with CD45.1 BM cells. At 5–6 wk after reconstitution with CD45.1 BM cells, the splenocytes were analyzed as in A. Percentages indicated are mean ± SD from five independent experiments. (C) Genomic DNA was prepared from the CD45− cells sorted from spleens and was analyzed by quantitative real-time PCR with deleted allele-specific primers. Data are mean ± SD from triplicate experiments. *, P < 0.0001.
Mentions: Because Mib1 is expressed in both CD45+ and CD45− cells in the spleen (Fig. 1 B), we investigated whether Mib1 functions in the hematopoietic cells or in the microenvironments for MZB cell development. When the lethally irradiated CD45.1 WT mice were reconstituted with BM cells from either the MMTV-Cre;Mib1+/f or MMTV-Cre;Mib1f/f mice, all of the CD45.1 recipient mice displayed not only T1 B and T2 B cells but also normal MZB cell development (Fig. 6 A and not depicted), indicating that the impaired MZB cell development in the MMTV-Cre;Mib1f/f mice is not caused by an autonomous defect in hematopoietic cells. However, in the reciprocal BMT experiment, in which the lethally irradiated MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were reconstituted with BM cells from CD45.1 congenic mice, B220+CD21+CD23lo/− and B220+CD1d+CD9+ MZB cell development was completely blocked in the MMTV-Cre;Mib1f/f recipient mice, whereas it was unaffected in the control mice (Fig. 6 B). In addition to MZB cells, cycling T2 B cell development, but not T1 B cell development, was severely affected in the MMTV-Cre;Mib1f/f recipient mice (Fig. 6 B and not depicted). These results show that the impaired MZB cell development in the MMTV-Cre;Mib1f/f mice originated from a nonhematopoietic microenvironment. Consistent with these findings, Mib1 was deleted ∼90% in isolated CD45− splenic stromal cells from MMTV-Cre;Mib1f/f mice as compared with that of the control mice (Fig. 6 C), suggesting that Mib1 regulates MZB and cycling T2 B cell development in the splenic microenvironments.

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