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The GTPase rho controls a p53-dependent survival checkpoint during thymopoiesis.

Costello PS, Cleverley SC, Galandrini R, Henning SW, Cantrell DA - J. Exp. Med. (2000)

Bottom Line: Previous work has shown that the guanine nucleotide binding protein Rho controls cell survival in T cell progenitors.Here we define the survival pathways controlled by Rho in pre-T cells and show that this GTPase is a pivotal regulator of the p53-mediated checkpoint operating at the time of beta selection: loss of Rho function results in apoptosis in pre-T cells, but this cell death is prevented by loss of p53.The prevention of cell death by loss of p53 restored numbers of early T cell progenitors but did not fully restore thymic cellularity.

View Article: PubMed Central - PubMed

Affiliation: Lymphocyte Activation Laboratory, Imperial Cancer Research Fund, London WC2A 3PX, United Kingdom.

ABSTRACT
During the early stages of thymopoiesis, cell survival is controlled by cytokines that regulate the expression of antiapoptotic proteins such as Bcl-2. At the pre-T cell stage, a critical checkpoint for beta chain selection is monitored by the tumor suppressor p53: pre-T cells can survive and differentiate when p53 is removed genetically or when its proapoptotic function is inactivated physiologically as a consequence of signaling through the pre-T cell receptor complex. Previous work has shown that the guanine nucleotide binding protein Rho controls cell survival in T cell progenitors. Here we define the survival pathways controlled by Rho in pre-T cells and show that this GTPase is a pivotal regulator of the p53-mediated checkpoint operating at the time of beta selection: loss of Rho function results in apoptosis in pre-T cells, but this cell death is prevented by loss of p53. The prevention of cell death by loss of p53 restored numbers of early T cell progenitors but did not fully restore thymic cellularity. Further analysis revealed that loss of Rho function caused survival defects in CD4/8 double-positive thymocytes that is independent of p53 but can be prevented by ectopic expression of Bcl-2. These studies highlight that the GTPase Rho is a crucial component of survival signaling pathways in at least two different thymocyte subpopulations: Rho controls the p53 survival checkpoint in pre-T cells and is also crucial for a p53 independent survival signaling pathway in CD4/8 double positives.

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Intracellular β chain staining of CD25+ thymocytes from lck-C3/p53−/−mice. Thymocytes from p53+/−, p53−/−, and lck-C3/p53−/− transgenic mice were stained with anti-CD25–PE and a panel of biotinylated antibodies (CD4bio, CD8bio, CD3bio, B220bio, Mac-1bio, NKbio, γδ-bio, and Gr-1bio) revealed with streptavidin–TRI-COLOR before saponin permeabilization and staining with an antibody to the common β chain epitope.
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Figure 2: Intracellular β chain staining of CD25+ thymocytes from lck-C3/p53−/−mice. Thymocytes from p53+/−, p53−/−, and lck-C3/p53−/− transgenic mice were stained with anti-CD25–PE and a panel of biotinylated antibodies (CD4bio, CD8bio, CD3bio, B220bio, Mac-1bio, NKbio, γδ-bio, and Gr-1bio) revealed with streptavidin–TRI-COLOR before saponin permeabilization and staining with an antibody to the common β chain epitope.

Mentions: The proliferation of DN3 and DN4 cells is dependent on the successful rearrangement and expression of TCR β chains. We therefore wished to exclude the possibility that loss of p53 in the Rho-deficient thymi was allowing survival of non–β-selected pre-T cells. Intracellular staining of DN3 cells with an antibody reactive to a common β chain epitope 31 quantitates levels of intracellular β subunits and thus monitors successful β chain rearrangements. The results show the presence of normal levels of intracellular β chains in CD44−25+ cells from lck-C3/p53−/− mice, indicating that they have apparently undergone normal β selection (Fig. 2).


The GTPase rho controls a p53-dependent survival checkpoint during thymopoiesis.

Costello PS, Cleverley SC, Galandrini R, Henning SW, Cantrell DA - J. Exp. Med. (2000)

Intracellular β chain staining of CD25+ thymocytes from lck-C3/p53−/−mice. Thymocytes from p53+/−, p53−/−, and lck-C3/p53−/− transgenic mice were stained with anti-CD25–PE and a panel of biotinylated antibodies (CD4bio, CD8bio, CD3bio, B220bio, Mac-1bio, NKbio, γδ-bio, and Gr-1bio) revealed with streptavidin–TRI-COLOR before saponin permeabilization and staining with an antibody to the common β chain epitope.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1887705&req=5

Figure 2: Intracellular β chain staining of CD25+ thymocytes from lck-C3/p53−/−mice. Thymocytes from p53+/−, p53−/−, and lck-C3/p53−/− transgenic mice were stained with anti-CD25–PE and a panel of biotinylated antibodies (CD4bio, CD8bio, CD3bio, B220bio, Mac-1bio, NKbio, γδ-bio, and Gr-1bio) revealed with streptavidin–TRI-COLOR before saponin permeabilization and staining with an antibody to the common β chain epitope.
Mentions: The proliferation of DN3 and DN4 cells is dependent on the successful rearrangement and expression of TCR β chains. We therefore wished to exclude the possibility that loss of p53 in the Rho-deficient thymi was allowing survival of non–β-selected pre-T cells. Intracellular staining of DN3 cells with an antibody reactive to a common β chain epitope 31 quantitates levels of intracellular β subunits and thus monitors successful β chain rearrangements. The results show the presence of normal levels of intracellular β chains in CD44−25+ cells from lck-C3/p53−/− mice, indicating that they have apparently undergone normal β selection (Fig. 2).

Bottom Line: Previous work has shown that the guanine nucleotide binding protein Rho controls cell survival in T cell progenitors.Here we define the survival pathways controlled by Rho in pre-T cells and show that this GTPase is a pivotal regulator of the p53-mediated checkpoint operating at the time of beta selection: loss of Rho function results in apoptosis in pre-T cells, but this cell death is prevented by loss of p53.The prevention of cell death by loss of p53 restored numbers of early T cell progenitors but did not fully restore thymic cellularity.

View Article: PubMed Central - PubMed

Affiliation: Lymphocyte Activation Laboratory, Imperial Cancer Research Fund, London WC2A 3PX, United Kingdom.

ABSTRACT
During the early stages of thymopoiesis, cell survival is controlled by cytokines that regulate the expression of antiapoptotic proteins such as Bcl-2. At the pre-T cell stage, a critical checkpoint for beta chain selection is monitored by the tumor suppressor p53: pre-T cells can survive and differentiate when p53 is removed genetically or when its proapoptotic function is inactivated physiologically as a consequence of signaling through the pre-T cell receptor complex. Previous work has shown that the guanine nucleotide binding protein Rho controls cell survival in T cell progenitors. Here we define the survival pathways controlled by Rho in pre-T cells and show that this GTPase is a pivotal regulator of the p53-mediated checkpoint operating at the time of beta selection: loss of Rho function results in apoptosis in pre-T cells, but this cell death is prevented by loss of p53. The prevention of cell death by loss of p53 restored numbers of early T cell progenitors but did not fully restore thymic cellularity. Further analysis revealed that loss of Rho function caused survival defects in CD4/8 double-positive thymocytes that is independent of p53 but can be prevented by ectopic expression of Bcl-2. These studies highlight that the GTPase Rho is a crucial component of survival signaling pathways in at least two different thymocyte subpopulations: Rho controls the p53 survival checkpoint in pre-T cells and is also crucial for a p53 independent survival signaling pathway in CD4/8 double positives.

Show MeSH
Related in: MedlinePlus