Rab7 and estrogen-ER as B cell-intrinsic mediators of auto/antibody responses

Rab7 和雌激素-ER 作为自身/抗体反应的 B 细胞内在介质

• 批准号: 8825268
• 负责人: Paolo Casali
• 金额: $ 44.4万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-12-15 至 2019-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8825268
• 关键词: 3' Untranslated Regions; Address; Affinity; Antibodies; Antibody Response; Antigens; Autoantibodies; Autoimmunity; Autophagocytosis; B cell differentiation; B-Lymphocytes; Binding; Binding Sites; Biogenesis; Bioinformatics; Cell physiology; Cells; Chemosensitization; Complex; Data; Defect; Down-Regulation; Elements; Estradiol; Estrogen Receptor alpha; Estrogen Receptors; Estrogens; Female; Fulvestrant; Generations; Genetic Recombination; Genetic Transcription; Health; Human; Human body; IgE; Immunoglobulin A; Immunoglobulin Class Switching; Immunoglobulin G; Immunoglobulin M; Immunoglobulin Somatic Hypermutation; Incidence; Indium; Infection; Lead; Light; Lupus; Mediating; Mediator of activation protein; Messenger RNA; MicroRNAs; Molecular; Monomeric GTP-Binding Proteins; Mus; Mutate; Mutation; Organ; Pathway interactions; Play; Process; Production; Regulation; Research; Role; Shapes; Staging; TNFRSF5 gene; TNFSF5 gene; Testing; Therapeutic; Time; Tissues; Transcript; Translations; Up-Regulation; Vaccines; Viral; Work; activation-induced cytidine deaminase; ds-DNA; homeodomain; human DICER1 protein; immunopathology; inhibitor/antagonist; lupus prone mice; microbial; mutant; novel; pathogen; promoter; research study; response; sperm cell; therapeutic target; tool; transcription factor; transcriptome sequencing; tumor

DESCRIPTION (provided by applicant): Rab7 and estrogen-ER as B cell-intrinsic mediators of auto/antibody responses. Our work of the first four years of R01 AI079705 has shown that estrogen promotes production of class- switched and mutated antibodies/autoantibodies, such as anti-dsDNA IgG in systemic lupus; estrogen potentiates induction of AID (critical for CSR and SHM) through upregulation of the HoxC4 homeodomain transcription factor; estrogen receptors (ERs) induce HoxC4 by binding to three conserved ER responsive elements (EREs) we identified in the HoxC4 promoter; HoxC4 directly binds to the AICDA/Aicda promoter to induce AID expression; and, HoxC4 and AID are highly expressed in lupus B cells; their KO blunts class- switched/mutated antibodies in normal mice, and autoantibodies/autoimmunity in lupus-prone mice. Thus, our significant findings on the B cell-intrinsic role of estrogen in CSR/SHM are highly relevant to heightened antibody/autoantibody responses and higher incidence of autoimmunity, particularly lupus, in females. This competitive renewal will test our novel hypothesis that Rab7 mediates antibody/autoantibody responses and is modulated in this function by estrogen (ß-estradiol)-ER (E2-ER). Upon induction in B cells, the Rab7 small GTPase would activate NF-κB (to increase HoxC4/Aicda transcription) and downregulate microRNAs (to relieve HoxC4 and Aicda transcripts from silencing), thereby inducing HoxC4/AID and CSR/SHM. This Rab7-dependent pathway would be enhanced by E2-ER, further upregulating CSR/SHM. Our hypotheses are supported by compelling preliminary data, including: demonstration that Rab7 plays a B cell-intrinsic role in inducing AID and CSR in T-independent and T-dependent antibody responses; evidence that Rab7 decreases Dicer, which is crucial to microRNA biogenesis; identification of multiple EREs in Rab7 promoter; upregulation of Rab7 induction by estrogen in normal B cells and its expression in lupus B cells. Aim 1 will address the roles of Rab7 and E2-ER in autoantibody responses and lupus immunopathology by using a Rab7-inhibitor (CID 1067700), an ER-antagonist (fulvestrant), and MRL/Faslpr/lpr mice deleting Rab7 [Tg(Aicda-cre)Rab7fl/fl] or ERα [Tg(Aicda-cre)ERαfl/fl] in "induced" B cells. Aim 2 will address the B cell- intrinsic functions of Rab7 in HoxC4/Aicda induction, CSR/SHM and antibody responses, underlying mechanisms (induction of NF-κB) and potentiation by E2-ER, by using B cells deleting Rab7, enforcing NF-κB activation through expression of a constitutively active IKKß mutant in these B cells, and by using B cells deleting ERα. Aim 3 will address Rab7 and estrogen downregulation of miR-23b, miR-26, miR-214 (silencing HoxC4 mRNA), miR-181b, miR-155 and miR-361 (silencing Aicda mRNA), and define the role of Rab7 in this process: possibly as a mediator of Dicer degradation through promotion of autophagy (-like) processes. By bringing the three research fields of autophagy, estrogen and microRNAs together to address the regulation of autoantibody production, our proposal provides an integrated approach to understand the complex problem of lupus and contributes to the definition of new lupus therapeutic targets.

描述（由申请人提供）：Rab7 和雌激素-ER 作为自身/抗体反应的 B 细胞内在介质。我们在 R01 AI079705 的前四年的工作表明，雌激素促进类别转换和突变抗体/自身抗体的产生，例如系统性狼疮中的抗 dsDNA IgG；雌激素通过上调 HoxC4 同源域转录因子来增强 AID 的诱导（对于 CSR 和 SHM 至关重要）；雌激素受体 (ER) 通过与我们在 HoxC4 启动子中鉴定的三个保守的 ER 反应元件 (ERE) 结合来诱导 HoxC4； HoxC4直接结合AICDA/Aicda启动子诱导AID表达；并且，HoxC4和AID在狼疮B细胞中高表达；他们的 KO 会削弱正常小鼠中的类别转换/突变抗体，以及易患狼疮的小鼠中的自身抗体/自身免疫。因此，我们关于雌激素在 CSR/SHM 中 B 细胞内在作用的重要发现与女性中抗体/自身抗体反应的增强和自身免疫（尤其是狼疮）的较高发病率高度相关。 这种竞争性更新将检验我们的新假设，即 Rab7 介导抗体/自身抗体反应，并在该功能中受到雌激素 (ß-雌二醇)-ER (E2-ER) 的调节。在 B 细胞中诱导后，Rab7 小 GTPase 将激活 NF-κB（以增加 HoxC4/Aicda 转录）并下调 microRNA（以解除 HoxC4 和 Aicda 转录本的沉默），从而诱导 HoxC4/AID 和 CSR/SHM。 E2-ER 会增强这种 Rab7 依赖性途径，进一步上调 CSR/SHM。我们的假设得到了令人信服的初步数据的支持，包括：证明 Rab7 在 T 依赖性和 T 依赖性抗体反应中诱导 AID 和 CSR 中发挥 B 细胞固有作用； Rab7 减少 Dicer 的证据，而 Dicer 对 microRNA 生物发生至关重要； Rab7 启动子中多个 ERE 的鉴定；雌激素对正常 B 细胞中 Rab7 诱导的上调及其在狼疮 B 细胞中的表达。 目标 1 将通过使用 Rab7 抑制剂 (CID 1067700)、ER 拮抗剂（氟维司群）和删除 Rab7 [Tg(Aicda-cre)Rab7fl/fl] 或 ERα 的 MRL/Faslpr/lpr 小鼠来探讨 Rab7 和 E2-ER 在自身抗体反应和狼疮免疫病理学中的作用[Tg(Aicda-cre)ERαfl/fl] 在“诱导的”B 细胞中。目标 2 将解决 Rab7 在 HoxC4/Aicda 诱导、CSR/SHM 和抗体反应中的 B 细胞内在功能、潜在机制（NF-κB 的诱导）和 E2-ER 的增强作用，方法是使用删除 Rab7 的 B 细胞，通过在这些 B 细胞中表达组成型活性 IKKß 突变体来强制 NF-κB 激活，以及使用删除 Rab7 的 B 细胞 ERα。目标 3 将解决 Rab7 和雌激素对 miR-23b、miR-26、miR-214（沉默 HoxC4 mRNA）、miR-181b、miR-155 和 miR-361（沉默 Aicda mRNA）的下调，并定义 Rab7 在此过程中的作用：可能通过促进自噬（类似）过程作为 Dicer 降解的介质。通过将自噬、雌激素和 microRNA 三个研究领域结合起来解决自身抗体产生的调节问题，我们的提案提供了一种综合方法来理解狼疮的复杂问题，并有助于定义新的狼疮治疗靶点。