Department of Cell and Systems Biology, University of Toronto, Toronto, ON, Canada.
Department of Biological Sciences, University of Toronto Scarborough, Toronto, ON, Canada.
Front Immunol. 2024 Apr 17;15:1386719. doi: 10.3389/fimmu.2024.1386719. eCollection 2024.
B-cell activation triggers the release of endoplasmic reticulum calcium stores through the store-operated calcium entry (SOCE) pathway resulting in calcium influx by calcium release-activated calcium (CRAC) channels on the plasma membrane. B-cell-specific murine knockouts of SOCE do not impact humoral immunity suggesting that alternative channels may be important.
We identified a member of the calcium-permeable transient receptor potential (TRP) ion channel family, TRPV5, as a candidate channel expressed in B cells by a quantitative polymerase chain reaction (qPCR) screen. To further investigate the role of TRPV5 in B-cell responses, we generated a murine TRPV5 knockout (KO) by CRISPR-Cas9.
We found TRPV5 polarized to B-cell receptor (BCR) clusters upon stimulation in a PI3K-RhoA-dependent manner. TRPV5 KO mice have normal B-cell development and mature B-cell numbers. Surprisingly, calcium influx upon BCR stimulation in primary TRPV5 KO B cells was not impaired; however, differential expression of other calcium-regulating proteins, such as ORAI1, may contribute to a compensatory mechanism for calcium signaling in these cells. We demonstrate that TRPV5 KO B cells have impaired spreading and contraction in response to membrane-bound antigen. Consistent with this, TRPV5 KO B cells have reduced BCR signaling measured through phospho-tyrosine residues. Lastly, we also found that TRPV5 is important for early T-dependent antigen specific responses post-immunization.
Thus, our findings identify a role for TRPV5 in BCR signaling and B-cell activation.
B 细胞激活通过内质网钙库通过储存操作钙进入 (SOCE) 途径触发钙库释放，导致质膜上的钙释放激活钙 (CRAC) 通道的钙内流。B 细胞特异性 SOCE 敲除小鼠不会影响体液免疫，这表明替代通道可能很重要。
我们通过定量聚合酶链反应 (qPCR) 筛选鉴定了钙通透瞬时受体电位 (TRP) 离子通道家族的成员 TRPV5，作为在 B 细胞中表达的候选通道。为了进一步研究 TRPV5 在 B 细胞反应中的作用，我们通过 CRISPR-Cas9 生成了 TRPV5 敲除 (KO) 小鼠。
我们发现 TRPV5 在刺激时以 PI3K-RhoA 依赖性方式极化到 B 细胞受体 (BCR) 簇。TRPV5 KO 小鼠具有正常的 B 细胞发育和成熟 B 细胞数量。令人惊讶的是，初级 TRPV5 KO B 细胞中 BCR 刺激引起的钙内流没有受损；然而，其他钙调节蛋白（如 ORAI1）的差异表达可能有助于这些细胞中钙信号的补偿机制。我们证明 TRPV5 KO B 细胞在响应膜结合抗原时的扩散和收缩受损。与此一致，TRPV5 KO B 细胞中 BCR 信号通过磷酸酪氨酸残基测量时降低。最后，我们还发现 TRPV5 对免疫后早期 T 依赖性抗原特异性反应很重要。
因此，我们的发现确定了 TRPV5 在 BCR 信号转导和 B 细胞激活中的作用。
