分类: 生物学 >> 生物物理学 >> 生物物理、生物化学与分子生物学 提交时间: 2016-05-12
摘要: The integrins, a family of transmembrane proteins, function in cell-to-cell and cell-to-extracellular matrix (ECM) adhesive interactions, and influence cell signaling of cell growth and differentiation. Expression of integrin 6 in three bladder cancer cell lines, HCV29, KK47 and YST1 were quantitatively analyzed by LC-MS using stable isotope labeling by amino acids in cell culture (SILAC), a simple and powerful proteomic strategy. The results showed that the non-invasive bladder cancer cell line KK47 expressed the highest level of integrin alpha 6. The expression of integrin alpha 6 in invasive bladder cancer cell line YTS1 was also higher than in normal bladder epithelial cell line HCV29. Furthermore, these results were confirmed by Western blotting, qPCR, immunohistochemistry and flow cytometry. Clinical data of mRNA 1TGA6 expression pattern from open-access database (www.oncomine.org) showed the same result during bladder cancer progression. All these indicated that integrin alpha 6 is associated with the invasion progress of the bladder cancer. The preliminary data in this study may sparkle the fundamental role of integrin 6 in the research of bladder cancer.
分类: 生物学 >> 生物物理学 >> 生物物理、生物化学与分子生物学 提交时间: 2016-05-12
摘要: Peroxisome proliferator-activated receptor-gamma (PPAR gamma) is a ligand-activated nuclear receptor and plays an essential role in insulin signaling. Macrophage infiltration into adipose tissue is a character of metabolic inflammation and closely related to insulin resistance in type 2 diabetes. The mechanism by which pro-inflammatory macrophages cause insulin resistance remains to be elucidated. Here we showed that coculture with macrophages significantly suppressed the transcriptional activity of PPAR gamma on its target genes in 3T3-L1 preadipocytes and diabetic primary adipocytes, depending on inducible nitric oxide synthase (iNOS). We further showed that PPAR gamma underwent S-nitrosylation in response to nitrosative stress. Mass-spectrometry and site-directed mutagenesis revealed that S-nitrosylation at cysteine 168 was responsible for the impairment of PPAR gamma function. Extended exposure to NO instigated the proteasome-dependent degradation of PPAR gamma. Consistently, in vivo evidence revealed an association of the decreased PPAR gamma protein level with increased macrophage infiltration in visceral adipose tissue (VAT) of obese diabetic db/db mice. Together, our results demonstrated that pro-inflammatory macrophages suppressed PPAR gamma activity in adipocytes via S-nitrosylation, suggesting a novel mechanism linking metabolic inflammation with insulin resistance. (C) 2015 Elsevier Inc. All rights reserved.
分类: 生物学 >> 生物物理学 >> 生物物理、生物化学与分子生物学 提交时间: 2016-05-11
摘要: Glycosylation is one of the most common and important post-translational modifications of proteins. Identification of large-scale N-linked glycoprotein is a very important aspect in glycoproteomics research. The N-glycopeptide enrichment is a key step in high-throughput identification of N-glycosylation site. Lectin enrichment and hydrazide chemistry are the two widely used N-glycopeptides enrichment methods. Each method can only enrich certain types of glycopeptides. It has been reported that the two methods are highly complementary, but few studies compared overlaps of glycosties from the two methods. In this paper, using HepG2 cells, we systematically compared the performance of hydrazide chemistry and lectins enrichment methods. The results showed that although the hydrazide method with glycopeptides enrichment efficiency of 76.7%, far higher than the 54.6% lectin enrichment method, 825 glycoprotein and 1 879 N-glycosylation sites identified with the lectin method was significantly more than 522 glycoprotein and 1 014 glycosylation sites enriched by the hydrazide method. Moreover, the two methods did not show significant complementary, together, only 853 glycoproteins and 1 959 N-glycosylation sites were identified. The overlapping results of identified N-glycosylation sites and N-glycoproteins from the two methods show that lectins enrichment method was better than hydrazide chemistry method.