OGT Overexpression in Women with Lupus

描述（由申请人提供）：狼疮折磨女性的次数是男性的9倍。雌激素有助于狼疮的严重程度，但不能完全解释风险增加。受损的DNA甲基化，一种抑制性表观遗传修饰，导致T细胞基因过度表达，导致狼疮。DNA甲基化也会使女性的一条X染色体沉默。CD 40 LG是一种X连锁基因，已知在狼疮中过表达，并有助于自身抗体的产生。我们发现，CD 40 LG上的非活性X去甲基化和过度表达的女性，但不是男性狼疮，易患女性狼疮。其他X连锁基因也可能脱甲基，诱发自身免疫。我们调查了甲基化敏感的T细胞基因，并确定O-连接的N-乙酰葡糖胺转移酶（OGT）作为另一个X-连锁基因在去甲基化的女性T细胞中过表达。OGT将N-乙酰葡糖胺（GlcNAc）偶联至多种蛋白质（包括信号分子）中的丝氨酸和苏氨酸，以类似于磷酸化的方式修饰功能，并被称为己糖胺信号通路（HSP）。HSP异常与糖尿病和神经退行性变有关，但关于其在T细胞中的作用知之甚少。我们推测，OGT的去甲基化的失活X的结果在妇女中的过度表达，改变HSP信号和致病性T细胞功能的修改信号。我们将通过以下方式检验这一假设：1）将来自健康男性和女性的对照和去甲基化的CD 4+和CD 8 + T细胞中的OGT mRNA和蛋白质水平与来自患有非活动性狼疮、活动性狼疮和疾病/年龄对照的男性和女性的CD 4+和CD 8 + T细胞中的水平进行比较，并通过亚硫酸氢盐测序确认OGT去甲基化，JNK和/或p38途径信号传导并改变T细胞基因表达模式，和3）使用具有T细胞特异性诱导型OGT转基因的转基因小鼠确定T细胞OGT过表达对自身免疫发展的功能意义。这些研究将描述一种调节T细胞基因表达的新途径，并描述该途径的异常如何使女性易患自身免疫。 公共卫生相关性：狼疮是一种自身免疫性疾病，主要影响女性，导致残疾，有时甚至死亡。女性易患狼疮的原因尚不清楚。我们的研究小组发现，一种调节基因表达的机制，称为DNA甲基化，在狼疮患者的T淋巴细胞中存在缺陷，导致基因过度表达，使细胞攻击身体。这有助于狼疮在那些有适当的基因组成。女性有两条X染色体，而男性只有一条。女性的一条X染色体因DNA甲基化而失活，以防止她们表达的基因是男性的两倍。然而，狼疮女性患者的第二条X染色体上的基因发生去甲基化，导致可能导致其疾病的基因过度表达。我们已经发现，当DNA甲基化被抑制时，在调节细胞功能中重要的X染色体基因OGT在女性T淋巴细胞中的表达加倍。这表明OGT也可能在患有狼疮的女性中过度表达，从而导致其疾病。本申请中描述的研究将确定患有狼疮的女性是否过度表达OGT，并确定功能后果。这些结果可能为女性自身免疫机制提供新的见解，并提出治疗这种疾病的新方法。

DESCRIPTION (provided by applicant): Lupus afflicts women 9 times more often than men. Estrogen contributes to lupus severity, but does not completely explain the increased risk. Impaired DNA methylation, a repressive epigenetic modification, causes overexpression of T cell genes that contribute to lupus. DNA methylation also silences one X chromosome in women. CD40LG is an X-linked gene known to be overexpressed in lupus and contribute to autoantibody production. We found that CD40LG on the inactive X demethylates and is overexpressed in women but not men with lupus, predisposing women to lupus. Other X-linked genes may also demethylate, predisposing to autoimmunity. We surveyed methylation sensitive T cell genes, and identified O-linked N-acetylglucosamine transferase (OGT) as another X-linked gene overexpressed in demethylated female T cells. OGT couples N-acetylglucosamine (GlcNAc) to serines and threonines in a variety of proteins including signaling molecules, modifying function in a manner analogous to phosphorylation and referred to as the hexosamine signaling pathway (HSP). HSP abnormalities are implicated in diabetes and neurodegeneration, but little is known regarding its role in T cells. We hypothesize that demethylation of OGT on the inactive X results in overexpression in women, altering HSP signaling and contributing to pathogenic T cell function by modifying signaling. We will test this hypothesis by: 1) Comparing OGT mRNA and protein levels in control and demethylated CD4+ and CD8+ T cells from healthy men and women with levels in CD4+ and CD8+ T cells from men and women with inactive lupus, active lupus, and disease/age controls, and confirming OGT demethylation by bisulfite sequencing, 2) Determining if OGT overexpression impairs T cell ERK, JNK and/or p38 pathway signaling and modifies T cell gene expression patterns, and 3) Determining the functional significance of T cell OGT overexpression on the development of autoimmunity using transgenic mice with a T cell specific inducible OGT transgene. These studies will characterize a new pathway regulating T cell gene expression, and characterize how abnormalities in the pathway may predispose women to autoimmunity. PUBLIC HEALTH RELEVANCE: Lupus is an autoimmune disease primarily affecting women, causing disability and sometimes death. The reason women are predisposed to lupus is unknown. Our group has found that a mechanism regulating gene expression, called DNA methylation, is defective in T lymphocytes from lupus patients, causing overexpression of genes that make the cells attack the body. This contributes to lupus in those with the appropriate genetic makeup. Women have 2 X chromosomes, while men have only one. One X chromosome is inactivated by DNA methylation in women to prevent them from expressing the genes twice as much as men. However, genes on the second X chromosome demethylate in women with lupus, causing overexpression of genes that may contribute to their disease. We have found that OGT, an X chromosome gene important in regulating cellular function, doubles expression in T lymphocytes from women when DNA methylation is inhibited. This suggests that OGT may also be overexpressed in women with lupus, contributing to their disease. The studies described in this application will determine if women with lupus overexpress OGT, and determine the functional consequences. The results may provide new insights into mechanisms causing autoimmunity in women, and suggest new ways to treat the disease.