Vacherot F, Delbé J, Heroult M, Barritault D, Fernig D G, Courty J
Laboratoire de Recherche sur la Croissance Cellulaire, la Réparation et la Régénération Tissulaires (CRRET), Unité Propre de Recherche de l'Enseignement Supérieur Associées an CNRS CNRS 7053, Université Paris XII-Val de Marne, France.
J Biol Chem. 1999 Mar 19;274(12):7741-7. doi: 10.1074/jbc.274.12.7741.
Heparin affin regulatory peptide (HARP) is a polypeptide belonging to a family of heparin binding growth/differentiation factors. The high affinity of HARP for heparin suggests that this secreted polypeptide should also bind to heparan sulfate proteoglycans derived from cell surface and extracellular matrix defined as extracellular compartments. Using Western blot analysis, we detected HARP bound to heparan sulfate proteoglycans in the extracellular compartments of MDA-MB 231 and MC 3T3-E1 as well as NIH3T3 cells overexpressing HARP protein. Heparitinase treatment of BEL cells inhibited HARP-induced cell proliferation, and the biological activity of HARP in this system was restored by the addition of heparin. We report that heparan sulfate, dermatan sulfate, and to a lesser extent, chondroitin sulfate A, displaced HARP bound to the extracellular compartment. Binding analyses with a biosensor showed that HARP bound heparin with fast association and dissociation kinetics (kass = 1.6 x 10(6) M-1 s-1; kdiss = 0.02 s-1), yielding a Kd value of 13 nM; the interaction between HARP and dermatan sulfate was characterized by slower association kinetics (kass = 0.68 x 10(6) M-1 s-1) and a lower affinity (Kd = 51 nM). Exogenous heparin, heparan sulfate, and dermatan sulfate potentiated the growth-stimulatory activity of HARP, suggesting that corresponding proteoglycans could be involved in the regulation of the mitogenic activity of HARP.
肝素亲和调节肽（HARP）是一种属于肝素结合生长/分化因子家族的多肽。HARP对肝素的高亲和力表明，这种分泌型多肽也应与源自细胞表面和细胞外基质（定义为细胞外区室）的硫酸乙酰肝素蛋白聚糖结合。通过蛋白质印迹分析，我们在MDA-MB 231和MC 3T3-E1的细胞外区室以及过表达HARP蛋白的NIH3T3细胞中检测到与硫酸乙酰肝素蛋白聚糖结合的HARP。用肝素酶处理BEL细胞可抑制HARP诱导的细胞增殖，并且通过添加肝素可恢复该系统中HARP的生物学活性。我们报告称，硫酸乙酰肝素、硫酸皮肤素以及程度较轻的硫酸软骨素A可取代与细胞外区室结合的HARP。用生物传感器进行的结合分析表明，HARP与肝素结合的结合和解离动力学较快（kass = 1.6 x 10(6) M-1 s-1；kdiss = 0.02 s-1），Kd值为13 nM；HARP与硫酸皮肤素之间的相互作用的特征是结合动力学较慢（kass = 0.68 x 10(6) M-1 s-1）且亲和力较低（Kd = 51 nM）。外源性肝素、硫酸乙酰肝素和硫酸皮肤素增强了HARP的生长刺激活性，这表明相应的蛋白聚糖可能参与了HARP促有丝分裂活性的调节。
