Preferential binding of insulin-like growth factor-II (IGF-II) to a putative alpha 2 beta 2 IGF-II receptor type in C2 myoblasts.
Preferential binding of insulin-like growth factor-II (IGF-II) to a putative alpha 2 beta 2 IGF-II receptor type in C2 myoblasts.
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胰岛素样生长因子-II (IGF-II) 优先结合 C2 成肌细胞中假定的 α2β2 IGF-II 受体类型。
DOI：
10.1111/j.1432-1033.1992.tb17183.x
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发表时间：
1992
期刊：
影响因子：
--
通讯作者：
Barenton,B
中科院分区：
文献类型：
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作者：
Domeyne,A;Pinset,C;Montarras,D;Garandel,V;Rosenfeld,RG;Barenton,B
我们研究了胰岛素样生长因子（IGF）在C2肌源性细胞系的两个亚克隆中的结合。在允许的亲本亚克隆中，成肌细胞在补充有胎牛血清的培养基中自发分化成肌管。与许可型成肌细胞不同，诱导型成肌细胞需要高浓度的胰岛素（1.6 μM）或低浓度的IGF-I（25 nM）来分化，并且MyoD 1的表达不是组成型的。在增殖和静止的成肌细胞和肌管的微粒体膜中研究了IGF受体。还在转染MyoD 1 cDNA的诱导型成肌细胞（克隆EP 5）中研究了IGF-II结合。诱导型和允许型细胞均显示出一类对IGF-I具有相似亲和力的结合位点（Kd 0.8 -1.2 nM）。在还原条件下，[125 I] IGF-I与微粒体膜的亲和交联揭示了在允许细胞中表观分子量为130 kDa和在诱导细胞中表观分子量为140 kDa的结合部分，其对应于IGF-I受体的α亚基。线性Scatchard图表明，[125 I]IGF-II结合至与IGF-II/M6 P受体结合相容的单一类结合位点（Kd 0.6 nM）。亲和交联实验证实了这一点，在还原和非还原条件下研究时，标记复合物的表观分子量分别为260 kDa和220 kDa。相反，竞争性抑制[125 I]IGF-II与可诱导静止成肌细胞的结合产生曲线Scatchard图，可分解为两类结合位点。其中一个对应于IGF-II/M6 P受体（Kd 0.2 nM），如交联实验所证明。第二个是亲和力最高的结合位点（Kd 0.04 nM），其受IGF-I的抑制低于IGF-II，且不受胰岛素抑制。在SDS/PAGE中，在还原条件下迁移至相当于140 kDa分子量的位置，在非还原条件下迁移至约300 kDa。这种非典型结合部分的标记不受抗（IGF-II/M6 P-受体）免疫球蛋白的抑制。在增殖期的诱导型成肌细胞和许可型成肌细胞中也有表达。它是不存在的允许静止成肌细胞和允许和诱导肌管。在诱导型细胞（EP 5细胞）中强制表达MyoD 1可显著降低[125 I]IGF-II与这种非典型受体的结合。从这些实验中可以看出，C2细胞表达一种推定的α2β2 IGF-II受体，其结构与胰岛素/IGF-I受体家族相关。它存在于成肌细胞中，但不在肌管中。这种受体的表达和表达的MyoD 1以及成肌细胞分化之间的可能关系进行了讨论。
We have studied insulin‐like‐growth‐factor (IGF) binding in two subclones of the C2 myogenic cell line. In the permissive parental subclone, myoblasts differentiate spontaneously into myotubes in medium supplemented with fetal calf serum. Unlike permissive myoblasts, inducible myoblasts require high concentrations of insulin (1.6 μM) or lower concentrations of IGF‐I (25 nM) to differentiate, and expression of MyoD1 is not constitutive. IGF receptors were studied in microsomal membranes of proliferating and quiescent myoblasts and myotubes. IGF‐II binding was also studied in inducible myoblasts transfected with the MyoD1 cDNA (clone EP5).Both inducible and permissive cells exhibited a single class of binding sites with similar affinity for IGF‐I (Kd0.8–1.2 nM). Affinity cross‐linking of [125I]IGF‐I to microsomal membranes, under reducing conditions, revealed a binding moiety with an apparent molecular mass of 130 kDa in permissive cells and 140 kDa in inducible cells, which corresponded to the α subunit of the IGF‐I receptor.In permissive quiescent myoblasts, linear Scatchard plots suggested that [125I]IGF‐II bound to a single class of binding sites (Kd0.6 nM) compatible with binding to the IGF‐II/M6P receptor. This was confirmed by affinity cross‐linking experiments showing a labeled complex with an apparent molecular mass of 260 kDa and 220 kDa when studied under reducing and non‐reducing conditions, respectively.In contrast, competitive inhibition of [125I]IGF‐II binding to inducible quiescent myoblasts generated curvilinear Scatchard plots which could be resolved into two single classes of binding sites. One of them corresponded to the IGF‐II/M6P receptor (Kd0.2 nM) as evidenced by cross‐linking experiments. The second was the binding site of highest affinity (Kd0.04 nM) which was less inhibited by IGF‐I than by IGF‐II and was not inhibited by insulin. It migrated in SDS/PAGE at a position equivalent a molecular mass of 140 kDa, under reducing conditions, and at approximately 300 kDa, under non‐reducing conditions. The labeling of this atypical binding moiety was not inhibited by anti(IGF‐II/M6P‐receptor) immunoglobulin. It was also observed in permissive and inducible myoblasts at proliferating stage. It was absent for permissive quiescent myoblasts and from permissive and inducible myotubes. Forced expression of MyoD1 in inducible cells (EP5 cells) dramatically reduced [125I]IGF‐II binding to this atypical receptor.It emerges from these experiments that C2 cells express a putative α2β2 IGF‐II receptor structurally related to the insulin/IGF‐I receptor family. It is present in myoblasts but not in myotubes. The possible relationship between expression of this receptor and expression of MyoD1 as well as myoblast differentiation is discussed.