SYK and ZAP70 kinases in lymphocyte selection

SYK 和 ZAP70 激酶在淋巴细胞选择中的作用

• 批准号: 10467215
• 负责人: Eric Meffre
• 金额: $ 61.73万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2027-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10467215
• 关键词: Ablation; Acute; Alleles; Antibodies; Autoantibodies; Autoimmune Diseases; B lymphoid malignancy; B-Cell Acute Lymphoblastic Leukemia; B-Cell Antigen Receptor; B-Cell Development; B-Cell Lymphomas; B-Cell Neoplasm; B-Lymphocytes; BLNK gene; Biological Markers; Calcium Signaling; Cell Death; Cell Proliferation; Cell Survival; Cells; Chronic Lymphocytic Leukemia; Clonal Deletion; Clone Cells; Development; Dominant-Negative Mutation; Engineering; Etiology; Evolution; Flare; Frequencies; Genetic; Genetic study; Human; Impairment; Knock-in; Lymphocyte; Malignant - descriptor; Mediating; Mus; Oncogenes; Oncogenic; Onset of illness; Output; Pathologic; Patients; Peripheral; Pharmacology; Phosphoric Monoester Hydrolases; Phosphotransferases; Preclinical Testing; Production; Protein Analysis; Receptor Cell; Receptor Signaling; Reporting; Reverse Transcriptase Polymerase Chain Reaction; Rheumatoid Arthritis; SYK gene; Signal Transduction; Structure of germinal center of lymph node; Systemic Lupus Erythematosus; T-Cell Development; T-Cell Lymphoma; T-Cell Receptor; T-Lymphocyte; Testing; ZAP-70 Gene; analog; anergy; autoreactive B cell; autoreactivity; base; cell transformation; genetic selection; humanized mouse; inducible gene expression; loss of function; mouse model; mutant; novel; optogenetics; patient derived xenograft model; phosphoproteomics; premalignant; pressure; prevent; promoter; remote control; segregation; single cell proteins; small molecule inhibitor; stoichiometry

ABSTRACT While B-cells are under intense selective pressure to eliminate autoreactive and pre-malignant clones, we identified SYK as central kinase to set the thresholds for negative selection. SYK and its highly homologous relative ZAP70 initiate B-cell receptor (BCR) and T-cell receptor (TCR) signaling in B- and T-lymphocytes, respectively. Even though the two kinases are almost identical and serve analogous functions, their expression in B- and T-cells is strictly segregated throughout evolution. Whereas the reason for separation of the two kinases is not known, aberrant coexpression of Syk and Zap70 was previously reported in B-cell chronic lymphocytic leukemia (CLL) and in peripheral T-cell lymphoma (PTCL). Our group recently demonstrated that aberrant ZAP70 expression is a common feature in pre- germinal center (GC) B-cell malignancies (B-ALL, CLL, MCL; Sadras et al., Mol Cell 2021). In genetic mouse models for B-ALL and B-CLL, inducible coexpression of Zap70 accelerated disease onset, while genetic deletion impaired malignant transformation. Likewise, inducible expression of Zap70 during early B-cell development subverted negative selection of autoreactive B-cells to promote pervasive autoantibody production. Mechanistically, ZAP70 competes with SYK and exerts a dominant-negative effect on SYK-dependent Ca2+-signaling. By occupying but not phosphorylating BLNK, BTK and PLC2 substrates upstream of calcium signaling, ZAP70 dramatically reduces the frequency of autonomous Ca2+-oscillations. Fast oscillations in the sole presence of SYK (4.5 mHz) are decoded by NFATC1 and result in anergy and cell death. In contrast, slow Ca2+-oscillations in the presence of ZAP70 (0.25 mHz) promote selective activation of NF-B and B-cell survival and proliferation. We test here the central hypothesis that B-cells sense pathological signaling downstream of an autoreactive BCR or a transforming oncogene through SYK-dependent high-frequency Ca2+- oscillations. At high frequencies, Ca2+-oscillations activate NFATC1 to initiate B-cell anergy and clonal deletion. Conversely, ZAP70 slows down Ca2+-oscillations to activate NF-B instead of NFATC1 and enables autoreactive and premalignant B-cell clones to persist and eventually give rise to autoimmune disease or pre-GC B-cell malignancies. We are proposing three Aims to (1) elucidate the mechanisms by which ZAP70 slows down SYK- dependent high-frequency Ca2+-oscillations, to (2) determine how coexpression of ZAP70 subverts negative selection of autoreactive B-cells in autoimmune disease and (3) to uncover how ZAP70 enables oncogenic signaling and overt malignant transformation of pre-malignant B-cells.

抽象的 虽然B细胞面临着强烈的选择压力，以消除自动反应性和恶性克隆，但 我们将SYK确定为中央激酶，以设定负选择的阈值。 Syk及其高度 同源相对ZAP70启动B细胞受体（BCR）和T细胞受体（TCR）信号传导 T淋巴细胞分别。即使这两个激酶几乎相同，并且具有类似的作用 在整个进化过程中，它们在B-和T细胞中的表达被严格隔离。而 两种激酶分离的原因尚不清楚，Syk和Zap70的异常共表达为 先前在B细胞慢性淋巴细胞性白血病（CLL）和周围T细胞淋巴瘤中报道 （PTCL）。我们的小组最近表明，异常的ZAP70表达是前pre- 生发中心（GC）B细胞恶性肿瘤（B-All，Cll，Mcl； Sadras等，Mol Cell 2021）。通用 B-all和B-CLL的小鼠模型，ZAP70加速疾病发作的诱导共表达，而 遗传缺失受损的恶性转化。同样，ZAP70在 早期的B细胞发育颠覆了自动反应性B细胞的负面选择，以促进普遍性 自身抗体生产。 从机械上讲，ZAP70与SYK竞争，并对SYK依赖性执行主导性影响 Ca2+信号。通过占据但不占磷酸化的blnk，BTK和PLC2底物的上游 钙信号传导，ZAP70大大降低了自主Ca2+振荡的频率。快速地 SYK（4.5 MHz）唯一存在的振荡被NFATC1解码，并导致反感 细胞死亡。相比之下，在ZAP70（0.25 MHz）存在下缓慢的Ca2+振荡促进选择性 NF-B和B细胞存活和增殖的激活。 我们在这里检验了中心假设，即B细胞在下游 自动反应性BCR或通过SYK依赖性高频Ca2+ - 转化的癌基因 - 振荡。在高频下，Ca2+振荡激活NFATC1以启动B细胞厌食和克隆 删除。相反，ZAP70减慢Ca2+振荡激活NF-B，而不是NFATC1和 启用自动反应性和前B细胞克隆，并最终引起自身免疫性 疾病或GC前B细胞Malignancys。 我们提出的三个目标是（1）阐明ZAP70减慢SYK-的机制 依赖性高频Ca2+振荡，至（2）确定ZAP70的共表达方式 自身免疫性疾病中自动反应性B细胞的阴性选择和（3）揭示了ZAP70如何 实现了恶性前B细胞的致癌信号传导和明显的恶性转化。