Areshkov P. CHI3L1 and CHI3L2 as antagonistic effectors of cell viability and signal pathways activators

Mammalian genomes incode a set of chitinase-like proteins, and chitinase-3-like protein 1 (CHI3L1) is the most highly investigated among them. Overexpression of CHI3L1 gene was found in different tumors, particularly in glioblastomas. As it was shown in our previous investigations, CHI3L2 also highly overexpressed in gliomas (Kavsan et al., 2008) but in contrast to CHI3L1 mitogen, the CHI3L2 treatment of serum-starved 293 or U-251 cells lead to cell growth arrest. Suppression of mitogenesis and cell viability was accompanied by sustained ERK1/2 phosphorylation, whereas CHI3L1 induced more transient kinases activation. Localization analysis of activated ERK1/2 showed that unstimulated 293 cells did not contain phospho-ERK1/2 but after addition of CHI3L1 or CHI3L2 proteins to medium cells were characterized by nuclear accumulation of kinases. However, in contrast to incubation with CHI3L1 that resulted in phospho-ERK1/2 nuclear accumulation no longer than 30 min, CHI3L2-induced localization of kinases in nuclei maintained at least to 4 hrs. In U-251 cells, the kinetics of ERK1/2-phosphorylation by CHI3L1 and CHI3L2 were quite similar to that in 293 cells but localization of phospho-ERK1/2 was found mainly in the cytoplasm. Antagonistic effect of CHI3L1 and CHI3L2 on 293 or U-251 cell viability was accompanied with ERK1/2 phosphorylation that may indicate the difference of MAPK signaling pathway activation by chitinase-like proteins during its initiation stage. It was found that Chlorpromazine which arrest ligand-receptor complexes clustering and formation of clathrin-coated pits resulted in both CHI3L1- and CHI3L2-induced ERK1/2 phosphorylation suppression. This apparently indicated that initial steps of receptor-mediated endocytosis was obligatory for CHI3L1- and CHI3L2-accociated MAPK signaling in 293 cells. At the same time, Dynasore Hydrate, specific inhibitor of dynamin-mediated clathrin-coated vesicle formation, blocked only CHI3L1-induced ERK1/2 phosphorylation, showing that in contrast to CHI3L2, CHI3L1-accociated MAPK signaling in 293 cells required coated vesicle formation. Same as 293 and U-251 cell medium supplementation by obtained from E. coli recombinant protein, secretion of CHI3L2 after lentivirus-mediated transduction of corresponding cDNA into 293 cells resulted in cell growth arrest under serum-free cultivation. However, under serum-supplemented culturing, analysis of CHI3L2-ectopically expressing 293 cells revealed increasing of cell viability and capacity for substrate-independent growth similarly to CHI3L1-oncogene transduction. Thus, it was shown that in contrast to CHI3L1 oncogene, CHI3L2 gene has oncogenic and tumor-suppression properties dependending on microenvironment conditions. Key words: CHI3L1, CHI3L2, malignant transformation, cell viability, signaling pathways activation.