The Role of GCM2 in Parathyroid Gland Homeostasis

描述(由申请人提供)：本重新提交的资金申请是ARRA R01两年资助的结果，该资助产生了新的数据，证明了甲状旁腺细胞特异性转录因子Gcm2(胶质细胞缺失2)在甲状旁腺早期胚胎发育后的关键作用，以及该蛋白如何在成熟的甲状旁腺中发挥作用，控制甲状旁腺细胞的功能和生存。长期目标是利用对Gcm2作用的了解来开发甲状旁腺功能亢进症患者的医疗新策略。此应用程序的目标是确定耗尽的程度 在基因工程小鼠模型中，Gcm2的缺失导致甲状旁腺细胞存活率降低。中心假说是，Gcm2在甲状旁腺中的表达是维持甲状旁腺细胞质量所必需的，缺乏Gcm2会导致甲状旁腺细胞死亡。我们建议使用我们建立的小鼠模型，以时间控制的方式有条件地从基因上删除Gcm2基因，以确定调控Gcm2基因表达的基因 Gcm2以及那些受Gcm2作用控制的基因，并揭示Gcm2缺失对成熟甲状旁腺细胞的影响。携带条件性Gcm2等位基因的小鼠也将使我们能够确定在晚年切除Gcm2可以在多大程度上拯救患有复制人类甲状旁腺功能亢进症的甲状旁腺疾病的小鼠。在强大的初步数据的指导下，这一假说将通过追求两个特定的目标来验证：1)确定Sonic hedgehog信号功能和其他转录因子途径在GCM2基因转录中的作用。根据我们的初步数据，我们预计Sonic hedgehog在抑制Gcm2在非甲状旁腺细胞中的表达方面发挥主要作用；以及2)确定Gcm2条件缺失对正常甲状旁腺转录体和功能亢进小鼠甲状旁腺功能和大小的影响。我们预计，正常甲状旁腺中Gcm2基因的缺失将揭示受Gcm2调控的基因，并将导致甲状旁腺萎缩。此外，利用已建立的原发性甲状旁腺功能亢进症小鼠模型，我们预计，在增生性或腺瘤性甲状旁腺中适时缺失Gcm2将导致甲状旁腺细胞存活率降低和甲状旁腺功能亢进症的消退。试剂和老鼠模型已经在手中，技术已经在申请者的实验室中被确立为可行。这种方法是创新的，因为它利用了新的小鼠模型，应用了新的技术，如增强的酵母1-杂交筛选来发现调控Gcm2表达的基因，并使用RNA-seq来开发依赖Gcm2的转录体的全面目录。这项拟议的研究具有重要意义，因为它有望促进我们对甲状旁腺细胞生物学的理解，并最终确定分子靶点，从而为人类原发性和三级甲状旁腺功能亢进症开发新的药物治疗方法。

DESCRIPTION (provided by applicant): This resubmitted proposal for funding is the result of 2 years of ARRA R01 funding that generated novel data demonstrating a critical role for the parathyroid cell-specific transcription factor Gcm2 (glial cell missing 2) after early embryologica development of the parathyroid glands as well how this protein functions in mature parathyroid glands to control function and survival of parathyroid cells. The long-term goal is use knowledge of the role of Gcm2 to develop novel strategies for medical treatment of patients with hyperparathyroidism. The objective in this application is to identify the extent to which depletion of Gcm2 leads to reduced survival of parathyroid cells in genetically engineered mouse models. The central hypothesis is that expression of Gcm2 in the parathyroid is necessary throughout life to maintain parathyroid cell mass, and that lack of Gcm2 will induce parathyroid cell death. We propose to use mouse models that we have developed to genetically delete the Gcm2 gene conditionally, in a temporally controlled manner, to identify the genes that regulate expression of Gcm2 as well as those that are controlled by Gcm2 action, and to uncover the effect of loss of Gcm2 on mature parathyroid cells. Mice with conditional Gcm2 alleles will also allow us to determine the extent to which ablation of Gcm2 late in life can "rescue" mice that have parathyroid disorders that replicate human hyperparathyroidism. Guided by strong preliminary data, this hypothesis will be tested by pursuing two specific aims: 1) determine the role of Sonic hedgehog signaling function and other transcription factor pathway on transcription of the GCM2 gene. Based on our preliminary data, we expect Sonic hedgehog plays a major role in repressing expression of Gcm2 in non-parathyroid cells; and 2) determine the effect of conditional deletion of Gcm2 on the transcriptosome of normal parathyroid glands and on function and size of hyperfunctioning mouse parathyroid glands. We expect that deletion of Gcm2 in normal parathyroid glands will reveal genes that are regulated by Gcm2 and will result in parathyroid atrophy. Moreover, using well-established murine models of primary hyperparathyroidism, we expect that timed deletion of Gcm2 in hyperplastic or adenomatous parathyroid glands will result in decreased survival of parathyroid cells and regression of hyperparathyroidism. The reagents and mouse models are in hand, and the techniques have been established as feasible in the applicants' labs. The approach is innovative because it utilizes novel mouse models, applies new techniques such as enhanced yeast 1-hybrid screening to uncover genes that regulate Gcm2 expression, and uses RNA-seq to develop a comprehensive catalog of the Gcm2-dependent transcriptosome. The proposed research is significant because it is expected to advance our understanding of parathyroid cell biology, and ultimately, to identify molecular targets that will allow development of new medical treatments for primary and tertiary hyperparathyroidism in humans.