Expression of TASK‐2 and its upregulation by B cell receptor stimulation in WEHI‐231 mouse immature B cells

Stimulation of B cell receptors (BCR ligation) induces apoptosis of immature B cells, which is critical to the elimination of self-reactive clones. In the mouse immature B cell line WEHI-231, the authors previously reported two types of background K+ channels with large (∼300 pS, LKbg) and medium (∼100 pS, MKbg) conductance in divalent cation-free conditions. While the authors have recently identified LKbg as TREK-2, the molecular nature of MKbg is unknown yet. In the present study, the authors found that BCR ligation markedly increased the background K+ conductance of WEHI-231. A single-channel study revealed that MKbg activity is increased by BCR ligation and that the biophysical properties (unitary conductance and pH sensitivity) of MKbg are consistent with those of TWIK-related acid-sensitive K+ channel 2 (TASK-2). The expression of TASK-2 and its upregulation by BCR ligation were confirmed by RT-PCR and immunoblot assays in WEHI-231. The BCR ligation-induced increase of K+ current was prevented by calcineurin inhibitors (cyclosporine A or FK506), and also by TASK-2-specific small interfering RNA (siRNA) transfection (si-TASK-2). Furthermore, si-TASK-2 attenuated the apoptosis of WEHI-231 caused by BCR ligation. TASK-2 activity and its mRNA were also confirmed in the primary splenic B cells of mouse. Summarizing, the authors report for the first time the expression of TASK-2 in B cells and surmise that the upregulation of TASK-2 by BCR ligation is associated with the apoptosis of immature B cells.

Download Full-text

RasGRP1 Sensitizes an Immature B Cell Line to Antigen Receptor-induced Apoptosis

Journal of Biological Chemistry ◽

10.1074/jbc.m314273200 ◽

2004 ◽

Vol 279 (19) ◽

pp. 19523-19530 ◽

Cited By ~ 23

Author(s):

Benoit Guilbault ◽

Robert J. Kay

Keyword(s):

T Cells ◽

B Cells ◽

B Cell ◽

Cell Line ◽

Cell Receptor ◽

B Cell Receptor ◽

Dominant Negative ◽

Dominant Negative Mutant ◽

Wehi 231 ◽

Induced Apoptosis

RasGRP1 is a guanine nucleotide exchange factor that activates Ras GTPases and is activated downstream of antigen receptors on both T and B lymphocytes. Ras-GRP1 provides signals to immature T cells that confer survival and proliferation, but RasGRP1 also promotes T cell receptor-mediated deletion of mature T cells. We used the WEHI-231 cell line as an experimental system to determine whether RasGRP1 can serve as a quantitative modifier of B cell receptor-induced deletion of immature B cells. A 2-fold elevation in RasGRP1 expression markedly increased apoptosis of WEHI-231 cells following B cell receptor ligation, whereas a dominant negative mutant of RasGRP1 suppressed B cell receptor-induced apoptosis. Activation of ERK1 or ERK2 kinases was not required for RasGRP1-mediated apoptosis. Instead, elevated RasGRP1 expression caused down-regulation of NF-κB and Bcl-xL, which provide survival signals counter-acting apoptosis induction by B cell receptor. Inhibition of NF-κB was sufficient to enhance B cell receptor-induced apoptosis of WEHI-231 cells, and ligation of co-stimulatory receptors that activate NF-κB suppressed the ability of RasGRP1 to promote B cell receptor-induced apoptosis. These experiments define a novel apoptosis-promoting pathway leading from B cell receptor to the inhibition of NF-κB and demonstrate that differential expression of RasGRP1 has the potential to modulate the sensitivities of B cells to negative selection following antigen encounter.

Download Full-text