αv Integrins regulate germinal center B cell responses through noncanonical autophagy
Fiona Raso, … , Adam Lacy-Hulbert, Mridu Acharya
Fiona Raso, … , Adam Lacy-Hulbert, Mridu Acharya
Published July 12, 2018
Citation Information: J Clin Invest. 2018;128(9):4163-4178. https://doi.org/10.1172/JCI99597.
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αv Integrins regulate germinal center B cell responses through noncanonical autophagy

Abstract

Germinal centers (GCs) are major sites of clonal B cell expansion and generation of long-lived, high-affinity antibody responses to pathogens. Signaling through TLRs on B cells promotes many aspects of GC B cell responses, including affinity maturation, class switching, and differentiation into long-lived memory and plasma cells. A major challenge for effective vaccination is identifying strategies to specifically promote GC B cell responses. Here, we have identified a mechanism of regulation of GC B cell TLR signaling, mediated by αv integrins and noncanonical autophagy. Using B cell–specific αv-KO mice, we show that loss of αv-mediated TLR regulation increased GC B cell expansion, somatic hypermutation, class switching, and generation of long-lived plasma cells after immunization with virus-like particles (VLPs) or antigens associated with TLR ligand adjuvants. Furthermore, targeting αv-mediated regulation increased the magnitude and breadth of antibody responses to influenza virus vaccination. These data therefore identify a mechanism of regulation of GC B cells that can be targeted to enhance antibody responses to vaccination.

Authors

Fiona Raso, Sara Sagadiev, Samuel Du, Emily Gage, Tanvi Arkatkar, Genita Metzler, Lynda M. Stuart, Mark T. Orr, David J. Rawlings, Shaun W. Jackson, Adam Lacy-Hulbert, Mridu Acharya

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Figure 4

αv Regulates TLR signaling in GC B cells through autophagy proteins.

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αv Regulates TLR signaling in GC B cells through autophagy proteins. (A ...
(A and B) GC B cells were sorted from spleens of αv-CD19 and control mice 14 days after immunization with VLP and restimulated in vitro with CpG DNA or VLP for the indicated times (minutes). Western blots show NF-κB p65 and IRF7 in nuclear fractions (A) or p62 and LC3b in whole-cell lysates (B). Also shown is staining of LSD1 or actin to confirm equivalent protein loading in nuclear fraction and whole-cell lysates, respectively. These were performed on the same blot, except in the case of LC3-II, for which actin staining was from aliquots of the same samples run on parallel gels contemporaneously. (C) Confocal images of sorted primary GC B cells (as in A and B) treated with CpG DNA, R848, or VLP for 2 hours and stained for LC3 and Hoescht. Scale bar: 2.9 µm. (D) Quantification of LC3 reorganization after stimulation with indicated TLR ligands. Data are based on analysis of at least 30 cells/condition. P values of less than 0.01 are shown (Pearson’s χ2 test). *P < 0.05. (E and F) FACS-sorted non-GC follicular B cells from spleens of VLP-immunized αv-CD19 and control mice analyzed as in A and B. (G) Expression of indicated genes measured by quantitative reverse-transcriptase PCR (QRT-PCR) in RNA from FACS-sorted VLP-specific GC cells, isolated 5 or 14 days after immunization with 2 μg VLPs. Data from individual mice are shown, with mean ± SD. P values of less than 0.05 are shown (2-tailed Student’s t test). *P < 0.05; **P < 0.005. Data are all from 1 representative experiment of at least 3 independent experiments (2 experiments for p62 blots) in which similar results were seen.