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Specific activation of ERK pathways by chitin oligosaccharides in embryonic zebrafish cell lines
Boguslawa E Snaar-Jagalska1, Simon F G Krens, Inmaculada Robina
1Institute of Biology, Leiden University, Clusius Laboratory, Wassenaarseweg 64, 2333 AL Leiden, The Netherlands. jagalska@rulbim.leidenuniv.nl
Abstract:
Chitin oligosaccharides (COs) play a role in plant development and are presumed to affect body plan formation during vertebrate embryogenesis. The mechanisms of COs recognition and cellular processes underlying embryonic development are still not understood. We analyze the possible link with the mitogen-activated protein kinase pathway that is conserved in evolution through the plant and animal kingdom and has been implicated in diverse cellular processes, including cell growth, proliferation, differentiation, survival, and vertebrate development. We show that in vivo stimulation of embryonic zebrafish cells ZF13 and ZF29 with chitin tetrasaccharides at 10-9 M concentration transiently induced activation/phosphorylation of extracellular regulated kinases (ERKs), with a maximum after 15 min. Furthermore the biological specificity of chitin tetrasaccharides and various derivatives was examined. The replacement of one or two GlcNAc residues of the chitin backbone by glucose and fucosylation of chitin tetrasaccharides at the reducing terminus caused a complete loss of their activity. We also tested a chitin tetrasaccharide analogue in which the oxygen atoms in glycosidic linkages were replaced by sulfur atoms. This analog, which could not be enzymatically hydrolyzed, was as potent an inducer as chitin tetrasaccharide. These results suggest that the observed activation of ERKs is chitin tetrasaccharide-specific and does not require further enzymatic processing. We examined possible signaling pathways leading to ERK activation by COs by use of phosphospecific antibodies and inhibitors. We conclude that a high-affinity CO receptor system exists that links to the Raf, MEK, and ERK pathway in zebrafish cells.
Insights
Chitin oligosaccharides (COs) activate extracellular regulated kinases (ERKs) in zebrafish embryos, suggesting a conserved signaling pathway for embryonic development. This chitin tetrasaccharide-specific activation does not require enzymatic processing.
Area of Science:
- Developmental Biology
- Molecular Biology
- Biochemistry
Background:
- Chitin oligosaccharides (COs) are known to influence plant development.
- Their role in vertebrate embryogenesis, particularly in body plan formation, is largely unknown.
- Understanding COs recognition and cellular signaling is crucial for developmental studies.
Purpose of the Study:
- To investigate the role of chitin oligosaccharides in vertebrate embryonic development.
- To explore the potential link between COs and the conserved mitogen-activated protein kinase (MAPK) pathway.
- To elucidate the specific mechanisms of COs recognition and signaling in zebrafish embryos.
Main Methods:
- Stimulation of embryonic zebrafish cell lines (ZF13 and ZF29) with chitin tetrasaccharides and derivatives.
- Analysis of extracellular regulated kinases (ERK) activation/phosphorylation using phosphospecific antibodies and inhibitors.
- Examination of signaling pathways including Raf, MEK, and ERK.
Main Results:
- Chitin tetrasaccharides at 10⁻⁹ M transiently induced ERK activation in zebrafish cells within 15 minutes.
- Specific structural modifications (glucose substitution, fucosylation) abolished activity, indicating biological specificity.
- A non-hydrolyzable sulfur-containing analog retained activity, suggesting no enzymatic processing is required.
- ERK activation was linked to the Raf, MEK, and ERK pathway.
Conclusions:
- A high-affinity chitin oligosaccharide receptor system exists in zebrafish cells.
- This receptor system is linked to the Raf, MEK, and ERK signaling pathway.
- COs play a role in vertebrate embryonic development through specific, non-enzymatically processed signaling.