Inhibition of N-linked oligosaccharide processing in tumor cells is associated with enhanced tissue inhibitor of metalloproteinases (TIMP) gene expression

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Abstract
Previous studies on tumor-cell glycosylation mutants and drugs which inhibit oligosaccharide processing suggest that expression of sialylated and highly branched complex-type N-linked oligosaccharides is required for efficient tumor-cell metastasis. These observations prompted the present investigation, in order to determine whether loss of sialylated and highly branched complex-type oligosaccharide in cellular glycoproteins might affect the expression of genes, particularly of genes which can influence the malignant phenotypes. Swainsonine, an inhibitor of Golgi α-mannosidase II, has previously been shown to inhibit invasion in vitro and reduces solid tumor in vivo. Metastatic sub-lines of the SPI murine mammary carcinoma cells cultured in the presence of swainsonine for 48 hr showed approximately 3-fold enhancement of TIMP mRNA levels, while urokinase (uPA) transcripts remained unchanged. To determine whether swainsonine's effect on TIMP mRNA levels was related to inhibition of oligosaccharide processing, we examined somatic glycosylation mutants with processing defects which attenuate metastatic potential. The Golgi UDP-Gal transport defect in murine MDAY-D2 lymphoma cells, Chinese hamster ovary cells (CHO) and human MeWo melanoma cells (i.e., D35W25, Lec8, 355 cell lines, respectively) was associated with increased TIMP mRNA levels. A revertant of Lec8 showed a return to the wild-type levels of TIMP mRNA, consistent with a causal relationship between the glycosylation mutation and TIMP gene expression. Similarly, CHO and MDAY-D2 mutants defective in GlcNAc-TV (i.e., Lec4 and KBL-I respectively), which also reduces metastatic potential, showed increases in TIMP transcript levels. Nuclear run-on assays showed that transcription of the TIMP gene was increased in cells where N-linked oligosaccharide processing was inhibited either by swainsonine or by a glycosylation mutation. The results suggest that cell-specific patterns of glycoprotein glycosylation in human, murine and hamster cell lines affects the transcription of select genes, including TIMP, which may influence the invasive phenotype.