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Divergent transcriptional programming of class-specific B cell memory by T-bet and RORα.

Wang NS, McHeyzer-Williams LJ, Okitsu SL, Burris TP, Reiner SL, McHeyzer-Williams MG - Nat. Immunol. (2012)

Bottom Line: Here we demonstrate that memory B cell subsets unexpectedly diverged across antibody class through differences in the effects of major transcriptional regulators.In contrast, the transcription factor RORα and not T-bet was expressed in IgA(+) memory B cells, with evidence that knockdown of RORα mRNA expression and chemical inhibition of transcriptional activity also resulted in lower survival and BCR expression of IgA(+) memory B cells.Thus, divergent transcriptional regulators dynamically maintain subset integrity to promote specialized immune function in class-specific memory B cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Microbial Science, The Scripps Research Institute, La Jolla, California, USA.

ABSTRACT
Antibody class defines function in B cell immunity, but how class is propagated into B cell memory remains poorly understood. Here we demonstrate that memory B cell subsets unexpectedly diverged across antibody class through differences in the effects of major transcriptional regulators. Conditional genetic deletion of the gene encoding the transcription factor T-bet selectively blocked the formation and antigen-specific response of memory B cells expressing immunoglobulin G2a (IgG2a) in vivo. Cell-intrinsic expression of T-bet regulated expression of the transcription factor STAT1, steady-state cell survival and transcription of IgG2a-containing B cell antigen receptors (BCRs). In contrast, the transcription factor RORα and not T-bet was expressed in IgA(+) memory B cells, with evidence that knockdown of RORα mRNA expression and chemical inhibition of transcriptional activity also resulted in lower survival and BCR expression of IgA(+) memory B cells. Thus, divergent transcriptional regulators dynamically maintain subset integrity to promote specialized immune function in class-specific memory B cells.

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Evidence for T-bet activity in IgG2a+ memory B cells(a) mRNA for T-bet targets at recall day 5 IgG2a+ memory versus naive B cells from day 5 memory NP-specific B cells from draining lymph nodes of C57BL/6 mice immunized with adjuvant and NP-KLH are FACS-purified and gated on Gr1−CD4−CD8−, B cell positive [CD138+ or CD19+], switched (IgM−IgD−), and NP+IgG2a+ memory B220hiCD38+CD138−. Naïve B cells are FACS-purified and gated on Gr1−CD4−CD8−, B cell positive (B220+CD19+CD138−) and IgM+IgD+CD23+ (top panel). IgG2a+ day 14 mature-GC (CD38−) versus recall day 5 (bottom panel). Mean±sem, n≥5. (b) mRNA for T-bet targets in recall day 5 IgG2a+ memory versus IgG2a+ plasma cells. NP+IgG2a+ plasma cells FACS-purified by gating on Gr1−CD4−CD8−, B cell positive [CD138+ or CD19+], switched (IgM−IgD−) and CD138+ are compared to recall day 5 IgG2a+ memory FACS-purified as in (a). (c) T-bet MFI in IgG2a+(G2a) and IgG1+(G1) B220hiCD38+ B cells (Gr1−CD4−CD8− IgM−IgD− CD19+CD138−) with control histograms from Tbx21−/− mice in grey. (d) T-bet and IFN-γ mRNA expression from single FACS-sorted cells (left panel) that are Gr1−CD4−CD8−, B cell positive [CD19+or CD138+], switched IgM−IgD−, and B220hiCD138−CD38+IgG2a+. Frequency of total single cells shown in left panel positive for T-bet or IFN-γ signal (right panel).
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Figure 4: Evidence for T-bet activity in IgG2a+ memory B cells(a) mRNA for T-bet targets at recall day 5 IgG2a+ memory versus naive B cells from day 5 memory NP-specific B cells from draining lymph nodes of C57BL/6 mice immunized with adjuvant and NP-KLH are FACS-purified and gated on Gr1−CD4−CD8−, B cell positive [CD138+ or CD19+], switched (IgM−IgD−), and NP+IgG2a+ memory B220hiCD38+CD138−. Naïve B cells are FACS-purified and gated on Gr1−CD4−CD8−, B cell positive (B220+CD19+CD138−) and IgM+IgD+CD23+ (top panel). IgG2a+ day 14 mature-GC (CD38−) versus recall day 5 (bottom panel). Mean±sem, n≥5. (b) mRNA for T-bet targets in recall day 5 IgG2a+ memory versus IgG2a+ plasma cells. NP+IgG2a+ plasma cells FACS-purified by gating on Gr1−CD4−CD8−, B cell positive [CD138+ or CD19+], switched (IgM−IgD−) and CD138+ are compared to recall day 5 IgG2a+ memory FACS-purified as in (a). (c) T-bet MFI in IgG2a+(G2a) and IgG1+(G1) B220hiCD38+ B cells (Gr1−CD4−CD8− IgM−IgD− CD19+CD138−) with control histograms from Tbx21−/− mice in grey. (d) T-bet and IFN-γ mRNA expression from single FACS-sorted cells (left panel) that are Gr1−CD4−CD8−, B cell positive [CD19+or CD138+], switched IgM−IgD−, and B220hiCD138−CD38+IgG2a+. Frequency of total single cells shown in left panel positive for T-bet or IFN-γ signal (right panel).

Mentions: As evidence for T-bet activity in antigen-specific IgG2a+ CD38hi memory B cells, transcription for a series of known T-bet target genes23 were elevated at day 5 of the memory response (Fig. 4a, top). Differential expression of these target genes implies that T-bet enables separate functions in IgG2a+ memory B cells compared to naive B cells. Recent studies have shown that Bcl-6 can directly bind to T-bet and repress T-bet target gene expression in T cells24. To test whether something similar occurs in B cells, we assayed the same T-bet target genes in germinal center B cells that contain elevated Bcl-6 and found that expression of these targets was diminished (Fig. 4a, bottom). Furthermore, Blimp-1 has been shown to directly antagonize T-bet expression25. Analysis of T-bet target genes in plasma cells that contain elevated Blimp-1 protein revealed reduced expression for most of the genes tested (Fig. 4b). Although T-bet activity is present in IgG2a+ memory B cells, Bcl-6 and Blimp-1 expression significantly decreases this activity within IgG2a+ GC B cells and IgG2a+ plasma cells respectively.


Divergent transcriptional programming of class-specific B cell memory by T-bet and RORα.

Wang NS, McHeyzer-Williams LJ, Okitsu SL, Burris TP, Reiner SL, McHeyzer-Williams MG - Nat. Immunol. (2012)

Evidence for T-bet activity in IgG2a+ memory B cells(a) mRNA for T-bet targets at recall day 5 IgG2a+ memory versus naive B cells from day 5 memory NP-specific B cells from draining lymph nodes of C57BL/6 mice immunized with adjuvant and NP-KLH are FACS-purified and gated on Gr1−CD4−CD8−, B cell positive [CD138+ or CD19+], switched (IgM−IgD−), and NP+IgG2a+ memory B220hiCD38+CD138−. Naïve B cells are FACS-purified and gated on Gr1−CD4−CD8−, B cell positive (B220+CD19+CD138−) and IgM+IgD+CD23+ (top panel). IgG2a+ day 14 mature-GC (CD38−) versus recall day 5 (bottom panel). Mean±sem, n≥5. (b) mRNA for T-bet targets in recall day 5 IgG2a+ memory versus IgG2a+ plasma cells. NP+IgG2a+ plasma cells FACS-purified by gating on Gr1−CD4−CD8−, B cell positive [CD138+ or CD19+], switched (IgM−IgD−) and CD138+ are compared to recall day 5 IgG2a+ memory FACS-purified as in (a). (c) T-bet MFI in IgG2a+(G2a) and IgG1+(G1) B220hiCD38+ B cells (Gr1−CD4−CD8− IgM−IgD− CD19+CD138−) with control histograms from Tbx21−/− mice in grey. (d) T-bet and IFN-γ mRNA expression from single FACS-sorted cells (left panel) that are Gr1−CD4−CD8−, B cell positive [CD19+or CD138+], switched IgM−IgD−, and B220hiCD138−CD38+IgG2a+. Frequency of total single cells shown in left panel positive for T-bet or IFN-γ signal (right panel).
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Figure 4: Evidence for T-bet activity in IgG2a+ memory B cells(a) mRNA for T-bet targets at recall day 5 IgG2a+ memory versus naive B cells from day 5 memory NP-specific B cells from draining lymph nodes of C57BL/6 mice immunized with adjuvant and NP-KLH are FACS-purified and gated on Gr1−CD4−CD8−, B cell positive [CD138+ or CD19+], switched (IgM−IgD−), and NP+IgG2a+ memory B220hiCD38+CD138−. Naïve B cells are FACS-purified and gated on Gr1−CD4−CD8−, B cell positive (B220+CD19+CD138−) and IgM+IgD+CD23+ (top panel). IgG2a+ day 14 mature-GC (CD38−) versus recall day 5 (bottom panel). Mean±sem, n≥5. (b) mRNA for T-bet targets in recall day 5 IgG2a+ memory versus IgG2a+ plasma cells. NP+IgG2a+ plasma cells FACS-purified by gating on Gr1−CD4−CD8−, B cell positive [CD138+ or CD19+], switched (IgM−IgD−) and CD138+ are compared to recall day 5 IgG2a+ memory FACS-purified as in (a). (c) T-bet MFI in IgG2a+(G2a) and IgG1+(G1) B220hiCD38+ B cells (Gr1−CD4−CD8− IgM−IgD− CD19+CD138−) with control histograms from Tbx21−/− mice in grey. (d) T-bet and IFN-γ mRNA expression from single FACS-sorted cells (left panel) that are Gr1−CD4−CD8−, B cell positive [CD19+or CD138+], switched IgM−IgD−, and B220hiCD138−CD38+IgG2a+. Frequency of total single cells shown in left panel positive for T-bet or IFN-γ signal (right panel).
Mentions: As evidence for T-bet activity in antigen-specific IgG2a+ CD38hi memory B cells, transcription for a series of known T-bet target genes23 were elevated at day 5 of the memory response (Fig. 4a, top). Differential expression of these target genes implies that T-bet enables separate functions in IgG2a+ memory B cells compared to naive B cells. Recent studies have shown that Bcl-6 can directly bind to T-bet and repress T-bet target gene expression in T cells24. To test whether something similar occurs in B cells, we assayed the same T-bet target genes in germinal center B cells that contain elevated Bcl-6 and found that expression of these targets was diminished (Fig. 4a, bottom). Furthermore, Blimp-1 has been shown to directly antagonize T-bet expression25. Analysis of T-bet target genes in plasma cells that contain elevated Blimp-1 protein revealed reduced expression for most of the genes tested (Fig. 4b). Although T-bet activity is present in IgG2a+ memory B cells, Bcl-6 and Blimp-1 expression significantly decreases this activity within IgG2a+ GC B cells and IgG2a+ plasma cells respectively.

Bottom Line: Here we demonstrate that memory B cell subsets unexpectedly diverged across antibody class through differences in the effects of major transcriptional regulators.In contrast, the transcription factor RORα and not T-bet was expressed in IgA(+) memory B cells, with evidence that knockdown of RORα mRNA expression and chemical inhibition of transcriptional activity also resulted in lower survival and BCR expression of IgA(+) memory B cells.Thus, divergent transcriptional regulators dynamically maintain subset integrity to promote specialized immune function in class-specific memory B cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Microbial Science, The Scripps Research Institute, La Jolla, California, USA.

ABSTRACT
Antibody class defines function in B cell immunity, but how class is propagated into B cell memory remains poorly understood. Here we demonstrate that memory B cell subsets unexpectedly diverged across antibody class through differences in the effects of major transcriptional regulators. Conditional genetic deletion of the gene encoding the transcription factor T-bet selectively blocked the formation and antigen-specific response of memory B cells expressing immunoglobulin G2a (IgG2a) in vivo. Cell-intrinsic expression of T-bet regulated expression of the transcription factor STAT1, steady-state cell survival and transcription of IgG2a-containing B cell antigen receptors (BCRs). In contrast, the transcription factor RORα and not T-bet was expressed in IgA(+) memory B cells, with evidence that knockdown of RORα mRNA expression and chemical inhibition of transcriptional activity also resulted in lower survival and BCR expression of IgA(+) memory B cells. Thus, divergent transcriptional regulators dynamically maintain subset integrity to promote specialized immune function in class-specific memory B cells.

Show MeSH
Related in: MedlinePlus