Monzat V, Ratovo G, Estival A, Fanjul M, Bertrand C, Clément B, Vaysse N, Hollande E, Clemente F
Laboratoire de Biologie Cellulaire, Université Paul Sabatier, Toulouse, France.
Eur J Cell Biol. 1996 Apr;69(4):316-26.
Fibroblast growth factor (FGF-2) is a multifunctional growth factor. In cells producing this factor, FGF-2 is synthesized as different molecular weight isoforms lacking the signal peptide sequence for secretion. All forms are highly concentrated in cells. The presence of a nuclearization signal sequence in some isoforms suggests the involvement of these isoforms in cell functions bypassing the cell surface receptors. Our aims were to better define the intracellular localizations of the FGF-2 isoforms by immunocytochemistry and confocal microscopy and to analyze whether these isoforms were involved in the expression of extracellular matrix (ECM) components. We chose the pancreatic acinar cell line AR4-2J since it does not synthesize FGF-2. These cells were retrovirally transfected by point-mutated FGF-2 cDNAs. The cell lines obtained produced either the 18 kDa form (A5 cells) or the 22.5 kDa form (A3 cells). In A5 cells, the 18 kDa form was found in the cytoplasm, on the cell surface reflecting its secretion, and in the nucleoli. Parental AR4-2J cells treated with exogenous FGF-2 exhibited identical localizations, suggesting that in A5 cells the 18 kDa form followed the same translocation pathways than the exogenous FGF-2. By contrast, in A3 cells the 22.5 kDa form was predominantly localized in the nucleoplasm but was undetectable on the cell surface, suggesting its direct translocation to the nucleus. Northern and Western blot analysis showed that cells expressing the high molecular weight form exhibited a decrease of laminin B1 protein level and mRNA stability. In contrast, collagen IV and fibronectin expressions were unmodified either in FGF-2-transfected cells or in parental cells treated by exogenous FGF-2. Thus, these data indicate that: 1) 18 and 22.5 kDa FGF-2 are preferentially localized in different nuclear compartments and 2) the high molecular weight form plays a role on the expression of some ECM components.
成纤维细胞生长因子（FGF - 2）是一种多功能生长因子。在产生这种因子的细胞中，FGF - 2被合成为不同分子量的异构体，这些异构体缺乏用于分泌的信号肽序列。所有形式都高度浓缩在细胞内。一些异构体中存在核定位信号序列，这表明这些异构体可能通过绕过细胞表面受体参与细胞功能。我们的目的是通过免疫细胞化学和共聚焦显微镜更好地确定FGF - 2异构体的细胞内定位，并分析这些异构体是否参与细胞外基质（ECM）成分的表达。我们选择了胰腺腺泡细胞系AR4 - 2J，因为它不合成FGF - 2。这些细胞通过点突变的FGF - 2 cDNA进行逆转录病毒转染。获得的细胞系分别产生18 kDa形式（A5细胞）或22.5 kDa形式（A3细胞）。在A5细胞中，18 kDa形式存在于细胞质、反映其分泌情况的细胞表面以及核仁中。用外源性FGF - 2处理的亲本AR4 - 2J细胞表现出相同的定位，这表明在A5细胞中，18 kDa形式遵循与外源性FGF - 2相同的转运途径。相比之下，在A3细胞中，22.5 kDa形式主要定位于核质中，但在细胞表面未检测到，这表明它直接转运到细胞核。Northern和Western印迹分析表明，表达高分子量形式的细胞中，层粘连蛋白B1蛋白水平和mRNA稳定性降低。相反，在FGF - 2转染的细胞或用外源性FGF - 2处理的亲本细胞中，IV型胶原和纤连蛋白的表达没有改变。因此，这些数据表明：1）18 kDa和22.5 kDa的FGF - 2优先定位于不同的核区室；2）高分子量形式对某些ECM成分的表达起作用。
