Key Points lncRNA BCALM (AC099524.1) is B cell specific and highly expressed in human lymphomas. BCALM is necessary for the interaction of signal transduction proteins PLD1 and AKAP9. BCALM promotes negative feedback that downmodulates BCR-stimulated Ca+ signaling. Of the thousands of long noncoding RNAs (lncRNA) identified in lymphocytes, very few have defined functions. In this study, we report the discovery and functional elucidation of a human B cell–specific lncRNA with high levels of expression in three types of B cell cancer and normal B cells. The AC099524.1 gene is upstream of the gene encoding the B cell–specific phospholipase C γ 2 (PLCG2), a B cell–specific enzyme that stimulates intracellular Ca2+ signaling in response to BCR activation. AC099524.1 (B cell–associated lncRNA modulator of BCR-mediated Ca+ signaling [BCALM]) transcripts are localized in the cytoplasm and, as expected, CRISPR/Cas9 knockout of AC099524.1 did not affect PLCG2 mRNA or protein expression. lncRNA interactome, RNA immunoprecipitation, and coimmunoprecipitation studies identified BCALM-interacting proteins in B cells, including phospholipase D 1 (PLD1), and kinase adaptor proteins AKAP9 (AKAP450) and AKAP13 (AKAP-Lbc). These two AKAP proteins form signaling complexes containing protein kinases A and C, which phosphorylate and activate PLD1 to produce phosphatidic acid (PA). BCR stimulation of BCALM-deficient B cells resulted in decreased PLD1 phosphorylation and increased intracellular Ca+ flux relative to wild-type cells. These results suggest that BCALM promotes negative feedback that downmodulates BCR-mediated Ca+ signaling by promoting phosphorylation of PLD1 by AKAP-associated kinases, enhancing production of PA. PA activates SHP-1, which negatively regulates BCR signaling. We propose the name BCALM for B-Cell Associated LncRNA Modulator of BCR-mediated Ca+ signaling. Our findings suggest a new, to our knowledge, paradigm for lncRNA-mediated modulation of lymphocyte activation and signaling, with implications for B cell immune response and BCR-dependent cancers.

中文翻译：

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BCALM (AC099524.1) 是一种人 B 淋巴细胞特异性长非编码 RNA，可调节 B 细胞受体介导的钙信号转导


要点 lncRNA BCALM (AC099524.1) 是 B 细胞特异性的，在人类淋巴瘤中高度表达。 BCALM 对于信号转导蛋白 PLD1 和 AKAP9 的相互作用是必需的。 BCALM 促进负反馈，下调 BCR 刺激的 Ca+ 信号传导。在淋巴细胞中发现的数千个长非编码 RNA (lncRNA) 中，很少有具有明确的功能。在这项研究中，我们报告了人类 B 细胞特异性 lncRNA 的发现和功能阐明，该 lncRNA 在三种类型的 B 细胞癌和正常 B 细胞中高水平表达。 AC099524.1 基因位于编码 B 细胞特异性磷脂酶 C γ 2 (PLCG2) 的基因的上游，PLCG2 是一种 B 细胞特异性酶，可刺激细胞内 Ca2+ 信号传导以响应 BCR 激活。 AC099524.1（BCR 介导的 Ca+ 信号转导的 B 细胞相关 lncRNA 调节剂 [BCALM]）转录物位于细胞质中，正如预期的那样，AC099524.1 的 CRISPR/Cas9 敲除不会影响 PLCG2 mRNA 或蛋白质表达。 lncRNA 相互作用组、RNA 免疫沉淀和免疫共沉淀研究鉴定了 B 细胞中的 BCALM 相互作用蛋白，包括磷脂酶 D 1 (PLD1) 以及激酶接头蛋白 AKAP9 (AKAP450) 和 AKAP13 (AKAP-Lbc)。这两种 AKAP 蛋白形成含有蛋白激酶 A 和 C 的信号复合物，可磷酸化并激活 PLD1 以产生磷脂酸 (PA)。相对于野生型细胞，BCALM 缺陷 B 细胞的 BCR 刺激导致 PLD1 磷酸化降低和细胞内 Ca+ 通量增加。这些结果表明，BCALM 促进负反馈，通过 AKAP 相关激酶促进 PLD1 磷酸化，从而下调 BCR 介导的 Ca+ 信号传导，从而增强 PA 的产生。 PA 激活 SHP-1，从而负向调节 BCR 信号传导。 我们建议将 BCALM 命名为 BCR 介导的 Ca+ 信号传导的 B 细胞相关 LncRNA 调节剂。据我们所知，我们的研究结果提出了一种新的 lncRNA 介导的淋巴细胞激活和信号传导调节范例，对 B 细胞免疫反应和 BCR 依赖性癌症具有影响。