OSTEOBLAST NUCLEAR MATRIX REGULATION OF COLLAGEN

间歇剂量的甲状旁腺激素(PTH)通过选择性地刺激骨形成来增加骨量。调节这种合成代谢作用的分子机制尚不清楚。使用体外模型来证明这一现象是有问题的，因为甲状旁腺素减弱了成骨细胞培养中胶原的合成。细胞结构影响胶原的合成，并可能导致骨和培养的成骨细胞对甲状旁腺素的相反反应。我们认为胶原蛋白的表达通过组织基质、细胞外基质的连接蛋白、细胞骨架和核基质与细胞结构偶联。此外，我们认为这种结构途径的最终结果是核基质结构转录因子，即通过弯曲启动子DNA来改变基因活性的蛋白质，以响应组织基质组织的变化。因此，甲状旁腺素诱导的组织基质蛋白的变化可能通过“拖拽”基因和改变启动子的几何形状来改变COL1A1的表达。我们已经鉴定出一个新的核基质结构转录因子家族NP/NMP4，它与大鼠I型胶原α1(I)多肽链(COL1A1)启动子序列特异性结合。这些独特的锌指蛋白与COL1A1启动子结合，使其弯曲，并包含基本的启动子活性。PTH调节NP/NMP4-COL1A1结合活性和NP/NMP4mRNA表达。激素诱导的NP/NMP4-COL1A1结合的变化在骨和培养中不同。我们的目标是研究NP/NMP4在介导基础和PTH调节的COL1A1转录中的功能作用。第一项研究将在体外和体内验证COL1A1启动子NP/NMP4结合元件参与成骨细胞基础转录和PTH调节转录的假设。第二项研究将确定NP和NMP4在介导基础和PTH调节的COL1A1转录中的不同功能。第三项研究将确定NP/NMP4蛋白的功能结构域。这一信息将确定甲状旁腺素在成骨细胞中作用的结构途径，并进一步阐明这种激素对骨的合成代谢作用的细胞基础。

Intermittent doses of parathyroid hormone (PTH) increase bone mass by selectively stimulating bone formation. The molecular mechanisms mediating this anabolic effect are unknown. Demonstrating this phenomenon is problematic using in vitro models because PTH attenuates collagen synthesis in osteoblast culture. Cell structure influences collagen synthesis and likely contributes to the opposing responses of bone and cultured osteoblasts to PTH. We propose that collagen expression is coupled to cell structure via the tissue matrix, the interlinking proteins of the extracellular matrix, cytoskeleton, and nuclear matrix. Furthermore, we propose that this structural pathway culminates with nuclear matrix architectural transcription factors, proteins that alter gene activity by bending promoter DNA in response to changes in tissue matrix organization. Therefore, PTH-induced changes in tissue matrix proteins may alter COL1A1 expression by "tugging" at the gene and altering promoter geometry. We have identified a novel family of nuclear matrix architectural transcription factors, NP/NMP4, that bind with sequence specificity to the promoter of the rat alpha1 (I) polypeptide chain (COL1A1) of type I collagen. These unique zinc finger proteins bind to the COL1A1 promoter, bend it, and contain basal promoter activity. PTH regulates NP/NMP4-COL1A1 binding activity and NP/NMP4 mRNA expression. Hormone-induced alterations in NP/NMP4-COL1A1 binding differ between bone and culture. Our goal is to investigate the functional role of NP/NMP4 in mediating basal and PTH-regulated COL1A1 transcription. The first study will test the hypothesis that the COL1A1 promoter NP/NMP4 binding elements contribute to osteoblast basal and PTH-modulated transcription in vitro and in vivo. The second study will determine the distinct functions of NP and NMP4 in mediating basal and PTH-regulated COL1A1 transcription. The third study will determine the functional domains of the NP/NMP4 proteins. This information will define a structural pathway for PTH action in osteoblasts and further clarify the cellular basis for the anabolic action of this hormone on bone.