Collagen Glycosylation, Maturation and Mineralization

说明(申请人提供)：I型胶原纤维是矿化组织(牙釉质除外)中的主要有机成分，为控制矿物晶体的沉积和堆积提供了稳定的模板。现已清楚的是，胶原蛋白翻译后修饰的模式是最终矿化的功能性纤维形成的关键因素。胶原的O-糖基化是由羟基半乳糖基转移酶(GT)和半乳糖基葡萄糖转移酶(GGT)催化的特定羟赖氨酸残基上的一种独特的修饰。虽然胶原糖基化的改变与胶原成熟和各种骨骼疾病有关，但这种修饰在骨骼中的确切机制和功能还不是很清楚。最近，我们证明了赖氨酰羟基酶3作为GGT的功能，并可能是糖基转移酶25家族成员1(GLT25D1)作为骨I型胶原的GT。基于这些和初步的数据，我们推测，通过调控编码GLT25D1的基因在成骨细胞中的表达，可以合成糖基化程度较低或过高的I型胶原，该系统可用于阐明糖基化在骨胶原成熟和矿化中的作用。为了验证这一假设，提出了以下具体目标：目的1.在成骨细胞中产生不同水平的糖基化的I型胶原，并确定其在体外胶原成熟和矿化中的作用，目的2.在体内移植系统中确定胶原糖基化对胶原成熟和矿化的影响。这项拟议的研究将为深入了解糖基化在胶原成熟和矿化中的作用，从而参与骨形成和病理，以及它在评估骨质量方面的潜在用途。

DESCRIPTION (provided by applicant): Fibrillar type I collagen is the predominant organic component in the mineralized tissues, except tooth enamel, providing a stable template to spatially control the deposition and packing of the mineral crystals. It has become clear that the pattern of collagen post-translational modifications is a critical factor for the formation of functional fibrils that eventually get mineralized. O-glycosylation of collagen is one of such unique modifications occurring at the specific hydroxylysine residues catalyzed sequentially by hydroxylysyl galactosyltransferase (GT) and, then, galactosylhydroxylysyl glucosyltransferase (GGT). Though alterations in collagen glycosylatioon have been implicated in collagen maturation and various bone disorders, the precise mechanisms and the functions of this modification in bone are not well understood. Most recently we have demonstrated that lysyl hydroxylase 3 functions as GGT and, possibly, glycosyltransferase 25 family member 1 (GLT25D1) as GT for bone type I collagen. Based on these and preliminary data, we hypothesized that by manipulating the expression of gene encoding GLT25D1 in osteoblastic cells, under- or overglycosylated type I collagen can be synthesized, and that this system can be used to elucidate the function of glycosylation in collagen maturation and mineralization in bone. To test the hypothesis, the following specific aims are proposed: Aim 1. To generate type I collagen with varied levels of glycosylation in osteoblastic cells and to define its role in collagen maturation and mineralization in vitro, Aim 2. To determine the effects of collagen glycosylation on collagen maturation and mineralization in an in vivo transplantation system. The proposed study should provide insights into the functions of glycosylation in collagen maturation and mineralization, thus, its involvement in bone formation and pathology, and its potential use for the assessment of bone quality.