Gaubert F, Escaffit F, Bertrand C, Korc M, Pradayrol L, Clemente F, Estival A
INSERM U 531, Institut Louis Bugnard, CHU Rangueil Bat L 3, 31403 Toulouse Cedex 4, France.
J Biol Chem. 2001 Jan 12;276(2):1545-54. doi: 10.1074/jbc.M001184200.
The high molecular weight (HMW) fibroblast growth factor (FGF)-2 isoform of 210 amino acids initiated at a CUG start codon possesses a nuclear localization sequence and is not secreted. In contrast, the low molecular weight (LMW) isoform of 155 amino acids initiated at the AUG start codon can be secreted and activates the cell surface FGF receptors. The two isoforms possess different biological properties; however, little is known about the intracrine regulatory mechanisms involved in the biological effects of the HMW FGF-2 isoform. Using pancreatic cells stably transfected with cDNAs leading to the expression of either the HMW FGF-2 (A3 cells) or the LMW form (A5 cells), we provide evidence that the two FGF-2 isoforms differentially modulate PKC levels. The LMW FGF-2 up-regulated the PKC epsilon levels by 1.6-fold; by contrast the HMW isoform down-regulated the level of this PKC isotype by about 3-fold and increased the amount of PKC delta by 1.7-fold. PKC mRNAs were also modified, suggesting that PKC expression was regulated at a pretranslational level. Additionally, expression of different levels of the HMW FGF-2 with an inducible expression system confirmed the role of this isoform on PKC delta and epsilon expressions. Increased activation of ERK-1 and -2 was also observed in cells expressing the HMW FGF-2. By using different PKC inhibitors and a dominant negative PKC delta, it was found that ERK activation was PKC delta-dependent. These data indicate that expression of HMW FGF-2 can modify PKC levels by acting at the intracellular level and that the overexpression of PKC delta induces ERK-1/2 activation. The expression of a dominant negative FGFR1 did not reduce ERK-1/2 activation by the HMW FGF-2, suggesting that ERK activation does not require FGFR activity. The signaling cascade downstream of ERK might be involved in the known mitogenic effect exerted by this FGF-2 isoform.
由210个氨基酸组成、起始于CUG起始密码子的高分子量（HMW）成纤维细胞生长因子（FGF）-2亚型具有一个核定位序列，且不分泌。相比之下，由155个氨基酸组成、起始于AUG起始密码子的低分子量（LMW）亚型可以分泌，并激活细胞表面的FGF受体。这两种亚型具有不同的生物学特性；然而，对于HMW FGF-2亚型生物学效应所涉及的胞内分泌调节机制却知之甚少。利用稳定转染了导致HMW FGF-2（A3细胞）或LMW形式（A5细胞）表达的cDNA的胰腺细胞，我们提供证据表明这两种FGF-2亚型对蛋白激酶C（PKC）水平有不同的调节作用。LMW FGF-2使PKCε水平上调了1.6倍；相比之下，HMW亚型使这种PKC同工型的水平下调了约3倍，并使PKCδ的量增加了1.7倍。PKC的mRNA也发生了改变，这表明PKC的表达在翻译前水平受到调节。此外，利用诱导表达系统表达不同水平的HMW FGF-2证实了该亚型对PKCδ和ε表达的作用。在表达HMW FGF-2的细胞中还观察到细胞外信号调节激酶（ERK）-1和-2的激活增加。通过使用不同的PKC抑制剂和显性负性PKCδ，发现ERK激活依赖于PKCδ。这些数据表明，HMW FGF-2的表达可通过在细胞内发挥作用来改变PKC水平，并且PKCδ的过表达诱导ERK-1/2激活。显性负性FGFR-1的表达并没有降低HMW FGF-2对ERK-1/2的激活作用，这表明ERK激活不需要FGFR活性。ERK下游的信号级联可能参与了这种FGF-2亚型已知的促有丝分裂作用。
