Affiliation:
1. Mendel University Brno 1 Laboratory of Molecular Embryology , , Czech Republic
2. Institute of Animal Physiology and Genetics 2 Developmental Biology Unit , , Zemědělská 1, 613 00 Brno, Czech Republic
3. Institute of Physical and Chemical Research 3 Division of Experimental Animal Research , , 3-1-1 Koyadai, Tsukuba, Ibaraki 305, Japan
Abstract
ABSTRACT
Basic fibroblast growth factor (FGF-2) functions as a natural inducer of mesoderm, regulator of cell differentiation and autocrine modulator of cell growth and transformation. The FGF-2 signals are transduced through receptors with intrinsic protein tyrosine kinase activity. However, receptor binding and activation is governed by extracellular matrix, cell surface or soluble proteoglycans. This paper focuses on the role of proteoglycans synthesized by embryonic cells, embryoglycans, in FGF-2 signaling via FGF receptor-1 (FGFR-1).
We found that embryoglycan ectodomain Lewis X, analog of developmentally regulated embryonic cell surface epitope TEC 1, promotes oligomerization of FGF-2 in the cell free chemical crosslinking. In vitro assays show that a large molar excess of extracellular Lewis X does not inhibit binding of FGF-2 to embryonic stem (ES) cells, but prevents the mitogenic effect of FGF-2. Western blot analysis of ES cells revealed the presence of abundant 52 kDa and trace amounts of 67 and 125 kDa isoforms of FGFR-1. However, none of these isoforms undergo any detectable changes in tyrosine phosphorylation under the conditions that modulate the mitogenic effect of FGF-2. Rather, a primary substrate of all receptor tyrosine kinases, phospholipase Cγ (PLCγ), is activated by both FGF-2 and Lewis X. The combination, FGF-2 plus Lewis X, leads to weak inhibition, when compared with the effects of FGF-2 and Lewis X, respectively. In accordance, the level of phosphorylation of non-receptor tyrosine kinase c-Src is reduced in a reversed pattern to PLCγ. Furthermore, in this particular cell type we show the presence of activated forms of extracellular signal-related kinase (ERK) in all nontreated and treated cells.
These findings demonstrate that embryoglycan ectodomains may act as negative regulators of FGF-2-induced ES cell proliferation, most likely through the FGFR-1-independent signaling pathway.
Publisher
The Company of Biologists