Limits...
Bad can act as a key regulator of T cell apoptosis and T cell development.

Mok CL, Gil-Gómez G, Williams O, Coles M, Taga S, Tolaini M, Norton T, Kioussis D, Brady HJ - J. Exp. Med. (1999)

Bottom Line: Here, we show that Bad expression levels are greatly increased in thymocytes during apoptosis.The numbers of T cells are greatly depleted and the processes of T cell development and selection are perturbed.These data suggest that Bad can act as a key regulator of T cell apoptosis and that this is a consequence of its upregulation after exposure to death stimuli.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Immunology, MRC National Institute for Medical Research, London NW7 1AA, United Kingdom.

ABSTRACT
Bad is a distant relative of Bcl-2 and acts to promote cell death. Here, we show that Bad expression levels are greatly increased in thymocytes during apoptosis. We generated bad transgenic mice to study the action of upregulated Bad expression on T cell apoptosis. The T cells from these mice are highly sensitive to apoptotic stimuli, including anti-CD95. The numbers of T cells are greatly depleted and the processes of T cell development and selection are perturbed. We show that the proapoptotic function of Bad in primary T cells is regulated by Akt kinase and that Bad overexpression enhances both cell cycle progression and interleukin 2 production after T cell activation. These data suggest that Bad can act as a key regulator of T cell apoptosis and that this is a consequence of its upregulation after exposure to death stimuli.

Show MeSH

Related in: MedlinePlus

Upregulation of Bad expression during thymocyte apoptosis.  Total cell lysate from 107 thymocytes per lane was resolved by SDS  PAGE, Western blotted, and Bad expression detected by probing with  anti-Bad antibody. Cell lysate was prepared from thymocytes immediately  after removal of the thymus from C57 Bl/10 mice (Control, 0 h). Lysates  were also prepared from thymocytes 5 h after γ-radiation, 5 h with 5 μM  dexamethasone, or 5 h in culture alone. The percent apoptosis in the  samples before cell lysate preparation is shown. The positive control for  Bad expression is a thymocyte lysate from a bad transgenic mouse; equal  numbers of cells were not added in this lane. Tubulin expression is a loading control to show that each track contains approximately equivalent  amounts of protein.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2192908&req=5

Figure 1: Upregulation of Bad expression during thymocyte apoptosis. Total cell lysate from 107 thymocytes per lane was resolved by SDS PAGE, Western blotted, and Bad expression detected by probing with anti-Bad antibody. Cell lysate was prepared from thymocytes immediately after removal of the thymus from C57 Bl/10 mice (Control, 0 h). Lysates were also prepared from thymocytes 5 h after γ-radiation, 5 h with 5 μM dexamethasone, or 5 h in culture alone. The percent apoptosis in the samples before cell lysate preparation is shown. The positive control for Bad expression is a thymocyte lysate from a bad transgenic mouse; equal numbers of cells were not added in this lane. Tubulin expression is a loading control to show that each track contains approximately equivalent amounts of protein.

Mentions: To study the action of Bad, we first determined the effect of apoptosis on Bad expression levels in thymocytes. Thymocytes were taken from C57 Bl/10 mice and cultured for 5 h, either unstimulated or after exposure to 10 Gy of γ-radiation or incubation with 5 μM dexamethasone. The level of apoptosis induced in the thymocytes was determined using flow cytometric analysis following cell lysis in a hypotonic buffer and DNA staining by propidium iodide (26). Equivalent numbers of thymocytes were loaded in each track for immunoblot analysis and the Western blot was probed with a polyclonal antibody specific for Bad. To determine the level of Bad in normal thymocytes, cells were frozen immediately after removal from the thymus (Control, 0 h in Fig. 1). As shown in Fig. 1, the level of Bad in normal thymocytes is very low, although there is a massive induction of Bad as thymocytes enter apoptosis particularly in response to strong apoptotic stimuli such as dexamethasone and γ-radiation. This suggests that Bad induction is concomitant with apoptosis in primary thymocytes and may well play an important regulatory role. To investigate this hypothesis further and to study the action of Bad in primary cells, we decided to establish lines of transgenic mice overexpressing Bad exclusively within their T cells.


Bad can act as a key regulator of T cell apoptosis and T cell development.

Mok CL, Gil-Gómez G, Williams O, Coles M, Taga S, Tolaini M, Norton T, Kioussis D, Brady HJ - J. Exp. Med. (1999)

Upregulation of Bad expression during thymocyte apoptosis.  Total cell lysate from 107 thymocytes per lane was resolved by SDS  PAGE, Western blotted, and Bad expression detected by probing with  anti-Bad antibody. Cell lysate was prepared from thymocytes immediately  after removal of the thymus from C57 Bl/10 mice (Control, 0 h). Lysates  were also prepared from thymocytes 5 h after γ-radiation, 5 h with 5 μM  dexamethasone, or 5 h in culture alone. The percent apoptosis in the  samples before cell lysate preparation is shown. The positive control for  Bad expression is a thymocyte lysate from a bad transgenic mouse; equal  numbers of cells were not added in this lane. Tubulin expression is a loading control to show that each track contains approximately equivalent  amounts of protein.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2192908&req=5

Figure 1: Upregulation of Bad expression during thymocyte apoptosis. Total cell lysate from 107 thymocytes per lane was resolved by SDS PAGE, Western blotted, and Bad expression detected by probing with anti-Bad antibody. Cell lysate was prepared from thymocytes immediately after removal of the thymus from C57 Bl/10 mice (Control, 0 h). Lysates were also prepared from thymocytes 5 h after γ-radiation, 5 h with 5 μM dexamethasone, or 5 h in culture alone. The percent apoptosis in the samples before cell lysate preparation is shown. The positive control for Bad expression is a thymocyte lysate from a bad transgenic mouse; equal numbers of cells were not added in this lane. Tubulin expression is a loading control to show that each track contains approximately equivalent amounts of protein.
Mentions: To study the action of Bad, we first determined the effect of apoptosis on Bad expression levels in thymocytes. Thymocytes were taken from C57 Bl/10 mice and cultured for 5 h, either unstimulated or after exposure to 10 Gy of γ-radiation or incubation with 5 μM dexamethasone. The level of apoptosis induced in the thymocytes was determined using flow cytometric analysis following cell lysis in a hypotonic buffer and DNA staining by propidium iodide (26). Equivalent numbers of thymocytes were loaded in each track for immunoblot analysis and the Western blot was probed with a polyclonal antibody specific for Bad. To determine the level of Bad in normal thymocytes, cells were frozen immediately after removal from the thymus (Control, 0 h in Fig. 1). As shown in Fig. 1, the level of Bad in normal thymocytes is very low, although there is a massive induction of Bad as thymocytes enter apoptosis particularly in response to strong apoptotic stimuli such as dexamethasone and γ-radiation. This suggests that Bad induction is concomitant with apoptosis in primary thymocytes and may well play an important regulatory role. To investigate this hypothesis further and to study the action of Bad in primary cells, we decided to establish lines of transgenic mice overexpressing Bad exclusively within their T cells.

Bottom Line: Here, we show that Bad expression levels are greatly increased in thymocytes during apoptosis.The numbers of T cells are greatly depleted and the processes of T cell development and selection are perturbed.These data suggest that Bad can act as a key regulator of T cell apoptosis and that this is a consequence of its upregulation after exposure to death stimuli.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Immunology, MRC National Institute for Medical Research, London NW7 1AA, United Kingdom.

ABSTRACT
Bad is a distant relative of Bcl-2 and acts to promote cell death. Here, we show that Bad expression levels are greatly increased in thymocytes during apoptosis. We generated bad transgenic mice to study the action of upregulated Bad expression on T cell apoptosis. The T cells from these mice are highly sensitive to apoptotic stimuli, including anti-CD95. The numbers of T cells are greatly depleted and the processes of T cell development and selection are perturbed. We show that the proapoptotic function of Bad in primary T cells is regulated by Akt kinase and that Bad overexpression enhances both cell cycle progression and interleukin 2 production after T cell activation. These data suggest that Bad can act as a key regulator of T cell apoptosis and that this is a consequence of its upregulation after exposure to death stimuli.

Show MeSH
Related in: MedlinePlus