The Role of GCM2 in Parathyroid Gland Homeostasis

描述（由申请人提供）：该重新提交的资助提案是2年ARRA R 01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Gcm 2（神经胶质细胞缺失2）在甲状旁腺早期胚胎发育后的关键作用，以及该蛋白质如何在成熟甲状旁腺中发挥作用，以控制甲状旁腺细胞的功能和存活。长期目标是利用Gcm 2作用的知识来开发甲状旁腺功能亢进患者药物治疗的新策略。本申请的目的是确定 Gcm 2的降低导致基因工程小鼠模型中甲状旁腺细胞的存活减少。核心假设是，在甲状旁腺中Gcm 2的表达是维持甲状旁腺细胞质量所必需的，并且缺乏Gcm 2将诱导甲状旁腺细胞死亡。我们建议使用我们已经开发的小鼠模型，以时间控制的方式有条件地遗传删除Gcm 2基因，以鉴定调节Gcm 2基因表达的基因。 Gcm 2以及由Gcm 2作用控制的那些，并揭示Gcm 2缺失对成熟甲状旁腺细胞的影响。具有条件性Gcm 2等位基因的小鼠也将使我们能够确定在生命后期消融Gcm 2可以“拯救”患有复制人类甲状旁腺功能亢进的甲状旁腺疾病的小鼠的程度。在强有力的初步数据的指导下，这一假设将通过追求两个具体目标进行测试：1）确定Sonic hedgehog信号传导功能和其他转录因子途径对GCM 2基因转录的作用。基于我们的初步数据，我们预期Sonic hedgehog在抑制非甲状旁腺细胞中Gcm 2的表达中起主要作用;和2）确定Gcm 2的条件性缺失对正常甲状旁腺的转录体以及对功能亢进的小鼠甲状旁腺的功能和大小的影响。我们预期正常甲状旁腺中Gcm 2的缺失将揭示受Gcm 2调控的基因，并将导致甲状旁腺萎缩。此外，使用良好建立的原发性甲状旁腺功能亢进的小鼠模型，我们预期在增生性或腺瘤性甲状旁腺中定时缺失Gcm 2将导致甲状旁腺细胞存活率降低和甲状旁腺功能亢进消退。试剂和小鼠模型已在手，并且技术已在申请人的实验室中建立为可行的。该方法是创新的，因为它利用了新的小鼠模型，应用了新技术，如增强的酵母单杂交筛选，以揭示调控Gcm 2表达的基因，并使用RNA-seq开发了Gcm 2依赖性转录体的全面目录。这项拟议的研究意义重大，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第三性甲状旁腺功能亢进症开发新的医学治疗方法。

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.