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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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The expression of E3 ubiquitin ligases and the deletion of Mib1 in the lymphoid tissues. (A and B) The expression of E3 ligases in the thymus (A) and spleens (B). Total RNAs were prepared from the thymus and spleen (total) and nonhematopoietic (CD45−) and hematopoietic (CD45+) fractions sorted from collagenase-treated thymi and spleens. They were subjected to quantitative real-time RT-PCR using specific primers for Cd45, Mib1, Mib2, Neur1, and Neur2. Data are mean ± SD from triplicate experiments. The mean expression level of Mib1 mRNA in the total thymus and spleen was defined as 1 arbitrary unit, as compared with the expression levels of the E3 ubiquitin ligases. (C) Immunohistochemistry was performed on the cryosections from the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f thymi with the anti-Mib1 antibody. Note that Mib1 expression (arrowheads) was reduced in the MMTV-Cre;Mib1f/f mice thymus. Bars, 5 μm. (D) Total protein extracts of the spleens from the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were immunoblotted with the anti-Mib1 antibody.
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fig1: The expression of E3 ubiquitin ligases and the deletion of Mib1 in the lymphoid tissues. (A and B) The expression of E3 ligases in the thymus (A) and spleens (B). Total RNAs were prepared from the thymus and spleen (total) and nonhematopoietic (CD45−) and hematopoietic (CD45+) fractions sorted from collagenase-treated thymi and spleens. They were subjected to quantitative real-time RT-PCR using specific primers for Cd45, Mib1, Mib2, Neur1, and Neur2. Data are mean ± SD from triplicate experiments. The mean expression level of Mib1 mRNA in the total thymus and spleen was defined as 1 arbitrary unit, as compared with the expression levels of the E3 ubiquitin ligases. (C) Immunohistochemistry was performed on the cryosections from the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f thymi with the anti-Mib1 antibody. Note that Mib1 expression (arrowheads) was reduced in the MMTV-Cre;Mib1f/f mice thymus. Bars, 5 μm. (D) Total protein extracts of the spleens from the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were immunoblotted with the anti-Mib1 antibody.

Mentions: The expression of the E3 ubiquitin ligases of Notch signaling in lymphoid tissues suggests the involvement of these molecules in lymphopoiesis. To test the relevance of the E3 ligases in lymphocyte development, we examined the expression of E3 ligase transcripts in lymphoid tissues, thymus, and spleen. Because we expected that the E3 ligases might work in the nonhematopoietic stromal cells, we measured the expression levels of E3 ligases in isolated cell populations by quantitative real-time RT-PCR. In the thymus, transcripts of all four E3 ligases were detected in both CD45− nonhematopoietic and CD45+ hematopoietic cells (Fig. 1 A). In particular, both the Mib1 and Mib2 transcripts were highly expressed, whereas the Neur1 and Neur2 transcripts were slightly expressed in the CD45− and CD45+ compartments of thymocytes as well as the entire thymus.


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)

The expression of E3 ubiquitin ligases and the deletion of Mib1 in the lymphoid tissues. (A and B) The expression of E3 ligases in the thymus (A) and spleens (B). Total RNAs were prepared from the thymus and spleen (total) and nonhematopoietic (CD45−) and hematopoietic (CD45+) fractions sorted from collagenase-treated thymi and spleens. They were subjected to quantitative real-time RT-PCR using specific primers for Cd45, Mib1, Mib2, Neur1, and Neur2. Data are mean ± SD from triplicate experiments. The mean expression level of Mib1 mRNA in the total thymus and spleen was defined as 1 arbitrary unit, as compared with the expression levels of the E3 ubiquitin ligases. (C) Immunohistochemistry was performed on the cryosections from the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f thymi with the anti-Mib1 antibody. Note that Mib1 expression (arrowheads) was reduced in the MMTV-Cre;Mib1f/f mice thymus. Bars, 5 μm. (D) Total protein extracts of the spleens from the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were immunoblotted with the anti-Mib1 antibody.
© Copyright Policy
Related In: Results  -  Collection

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

fig1: The expression of E3 ubiquitin ligases and the deletion of Mib1 in the lymphoid tissues. (A and B) The expression of E3 ligases in the thymus (A) and spleens (B). Total RNAs were prepared from the thymus and spleen (total) and nonhematopoietic (CD45−) and hematopoietic (CD45+) fractions sorted from collagenase-treated thymi and spleens. They were subjected to quantitative real-time RT-PCR using specific primers for Cd45, Mib1, Mib2, Neur1, and Neur2. Data are mean ± SD from triplicate experiments. The mean expression level of Mib1 mRNA in the total thymus and spleen was defined as 1 arbitrary unit, as compared with the expression levels of the E3 ubiquitin ligases. (C) Immunohistochemistry was performed on the cryosections from the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f thymi with the anti-Mib1 antibody. Note that Mib1 expression (arrowheads) was reduced in the MMTV-Cre;Mib1f/f mice thymus. Bars, 5 μm. (D) Total protein extracts of the spleens from the MMTV-Cre;Mib1+/f and MMTV-Cre;Mib1f/f mice were immunoblotted with the anti-Mib1 antibody.
Mentions: The expression of the E3 ubiquitin ligases of Notch signaling in lymphoid tissues suggests the involvement of these molecules in lymphopoiesis. To test the relevance of the E3 ligases in lymphocyte development, we examined the expression of E3 ligase transcripts in lymphoid tissues, thymus, and spleen. Because we expected that the E3 ligases might work in the nonhematopoietic stromal cells, we measured the expression levels of E3 ligases in isolated cell populations by quantitative real-time RT-PCR. In the thymus, transcripts of all four E3 ligases were detected in both CD45− nonhematopoietic and CD45+ hematopoietic cells (Fig. 1 A). In particular, both the Mib1 and Mib2 transcripts were highly expressed, whereas the Neur1 and Neur2 transcripts were slightly expressed in the CD45− and CD45+ compartments of thymocytes as well as the entire thymus.

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