Germinal Center Centroblasts Transition to a Centrocyte Phenotype According to a Timed Program and Depend on the Dark Zone for Effective Selection

Abstract

SummaryGerminal center (GC) B cells cycle between the dark zone (DZ) and light zone (LZ) during antibody affinity maturation. Whether this movement is necessary for GC function has not been tested. Here we show that CXCR4-deficient GC B cells, which are restricted to the LZ, are gradually outcompeted by WT cells indicating an essential role for DZ access. Remarkably, the transition between DZ centroblast and LZ centrocyte phenotypes occurred independently of positioning. However, CXCR4-deficient cells carried fewer mutations and were overrepresented in the CD73+ memory compartment. These findings are consistent with a model where GC B cells change from DZ to LZ phenotype according to a timed cellular program but suggest that spatial separation of DZ cells facilitates more effective rounds of mutation and selection. Finally, we identify a network of DZ CXCL12-expressing reticular cells that likely support DZ functions.

Clathrin light chains’ role in selective endocytosis influences antibody isotype switching

“…Because we observed increased surface TGFβR2 on KO B cells, we assessed surface levels of other receptors critical for B-cell function in GCs (29). GC B cells from Pp and mesenteric LNs (mLNs) of KO mice had higher surface levels of CXCR4, compared with equivalent B cells from WT controls, whereas B220 levels were not affected in the KO (Fig.…”

“…Spleen, mesenteric lymph node, and Peyer's patch cells were isolated and labeled for FACS analysis as described previously (29). Antibodies used for GC B-cell identification, Ig isotype detection, mixed bone chimera analysis, and labeling of TGFβR2, CXCR4, and CXCR5 are specified in SI Materials and Methods.…”

“…We speculate that the elevated TGFβR signaling caused by CLCa loss dominates over signaling from factors that promote IgG1 switching (e.g., IL4), which may have been most evident in the Peyer's patch where sufficient amounts of TGFβ are available to stimulate elevated surface TGFβRs. The reduced frequency of germinal center (GC) B cells in CLCa-null mice could be explained by increased surface expression and reduced internalization of CXCR4, which plays a role in cell distribution within the GC (29). Additionally, CLCa deficiency might affect internalization of sphingosine-1-phosphate receptor 2 (S1PR2), a GPCR that negatively regulates GC B-cell numbers (33), but lack of reagents to track S1PR2 surface expression prevented us from testing this hypothesis.…”

Differential expression of APE1 and APE2 in germinal centers promotes error-prone repair and A:T mutations during somatic hypermutation

“…1C). Recent reports have also shown low levels of APE1 transcripts in the GC (34,35), especially in the rapidly proliferating centroblasts of the dark zone, relative to centrocytes in the light zone where selection occurs. Cultured spleen B cells express roughly equivalent amounts of APE1 and APE2 mRNA (Fig.…”

A Stochastic Model of the Germinal Center Integrating Local Antigen Competition, Individualistic T–B Interactions, and B Cell Receptor Signaling

“…These results suggest that accessory molecules regulating individual cross-talks between Tfh and GC B cells may significantly impinge on qualitative and quantitative nature of affinity maturation. Furthermore, experimental studies of chemotaxis-based GC dynamics and zoning have revealed that genetic ablation of CXCR4 from the majority of GC B cells abrogates the GC LZ-DZ segregation but leaves GC maintenance and cyclic reaction relatively normal (22). FOXO1-mediated transcription regulation intrinsically dictates the DZ state (23,24).…”