CD44 MEDIATED CATABOLISM OF HYALURONAN BY CHONDROCYTES

CD44 介导软骨细胞对透明质酸的分解代谢

• 批准号: 7755215
• 负责人: Warren Knudson
• 金额: $ 2.44万
• 依托单位: EAST CAROLINA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-09-30 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7755215
• 关键词: ADAMTS; Actins; Affect; Binding; CD44 gene; Cartilage; Catabolism; Cattle; Cell membrane; Cells; Centrifugation; Chondrocytes; Co-Immunoprecipitations; Cytoplasmic Tail; Cytoskeleton; Data; Endocytosis; Enzymes; Event; Excision; Funding; Generations; Goals; Half-Life; Hyaluronan; Hyaluronidase; Inorganic Sulfates; Interleukin-1; Label; Link; Literature; Matrix Metalloproteinases; Measures; Mediating; Membrane; Membrane Microdomains; Metabolism; Methods; Monitor; Nature; One-Step dentin bonding system; Organ Culture Techniques; Peptide Hydrolases; Play; Process; Protein Isoforms; Proteins; Proteoglycan; Proteolytic Processing; Regulation; Research Personnel; Residual state; Resistance; Role; Slice; Staging; Techniques; Tertiary Protein Structure; Unspecified or Sulfate Ion Sulfates; Work; aggrecan; articular cartilage; base; bone morphogenetic protein 7; enzyme substrate; extracellular; link protein; monomer; mutant; palmitoylation; programs; receptor; receptor mediated endocytosis

DESCRIPTION (provided by applicant): The underlying new hypothesis of this proposal is that CD44 receptor-mediated endocytosis of hyaluronan (HA) provides a mechanism for chondrocytes to regulate local proteolytic events in the turnover of aggrecan proteoglycan (PG). During the current funding period we have determined that the local turnover of HA in cartilage occurs by CD44-mediated endocytosis, as there is no contribution of membrane-associated hyaluronidases to extracellular HA degradation. In addition we have determined that the ITEGE and DIPEN-containing aggrecan G1 domains-domains that remain bound to HA following proteolytic cleavage of PG, are co-internalized with HA by the same CD44-mediated mechanism. Furthermore, we have also shown that when metabolism is altered, and chondrocytes become more "catabolic" due to IL-1 treatment, there is an increase in CD44 expression by 6-8 fold at the protein level -- a change that increases the internalization of HA by 3 fold above control. Another critical observation that we made was that HA decorated with intact proteoglycan monomers cannot be internalized. Thus, for HA turnover to proceed, cleavage of PG within or near the G1-G2 domains is a prerequisite event. We propose that the presence of intact, highly-sulfated PG bound to HA establishes a resistance that inhibits CD44-mediated HA endocytosis. However the active cell-mediated processes that drive endocytosis also generate a focal clustering of PG aggregates, setting the stage for targeted PG degradation. Our new exciting data demonstrates that chondrocyte CD44 clusters into lipid raft microdomains as a prerequisite for endocytosis. We can now selectively manipulate CD44-mediated endocytosis of HA and determine the resultant effect on the rate of PG degradation. We will also determine whether CD44 serves as a raft-anchoring platform for MMP and ADAMTS proteinases- proteinases that recently have been shown to be present as active membrane-bound enzymes. Thus, the focus of this proposal is to determine the mechanistic link between HA endocytosis and PG degradation.

描述（由申请人提供）：该提案的潜在新假设是CD44受体介导的透明质酸（HA）内吞作用为软骨细胞提供了一种机制，以调节聚集蛋白聚糖蛋白聚糖（PG）周转中的局部蛋白水解事件。在当前资助期间，我们确定软骨中 HA 的局部周转是通过 CD44 介导的内吞作用发生的，因为膜相关透明质酸酶对细胞外 HA 降解没有贡献。此外，我们还确定，含有 ITEGE 和 DIPEN 的聚集蛋白聚糖 G1 结构域（在 PG 蛋白水解裂解后仍与 HA 结合的结构域）通过相同的 CD44 介导机制与 HA 共内化。此外，我们还表明，当新陈代谢发生改变，并且软骨细胞因 IL-1 处理而变得更加“分解代谢”时，CD44 表达在蛋白质水平上增加了 6-8 倍，这一变化使 HA 的内化比对照增加了 3 倍。我们进行的另一个关键观察是，用完整蛋白聚糖单体修饰的 HA 不能被内化。因此，为了进行 HA 周转，G1-G2 结构域内或附近的 PG 裂解是先决条件。我们认为，与 HA 结合的完整、高度硫酸化的 PG 的存在会产生抑制 CD44 介导的 HA 内吞作用的抵抗力。然而，驱动内吞作用的活跃细胞介导过程也会产生 PG 聚集体的焦点聚集，为目标 PG 降解奠定基础。我们令人兴奋的新数据表明，软骨细胞 CD44 聚集成脂筏微区是内吞作用的先决条件。我们现在可以选择性地操纵 CD44 介导的 HA 内吞作用，并确定由此产生的对 PG 降解速率的影响。我们还将确定 CD44 是否充当 MMP 和 ADAMTS 蛋白酶的筏锚定平台，这些蛋白酶最近已被证明作为活性膜结合酶存在。因此，该提案的重点是确定 HA 内吞作用和 PG 降解之间的机制联系。