Enhanced Membrane-tethered Mucin 3 (MUC3) Expression by a Tetrameric Branched Peptide with a Conserved TFLK Motif Inhibits Bacteria Adherence
Enhanced Membrane-tethered Mucin 3 (MUC3) Expression by a Tetrameric Branched Peptide with a Conserved TFLK Motif Inhibits Bacteria Adherence
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具有保守 TFLK 基序的四聚体分支肽增强膜束缚粘蛋白 3 (MUC3) 表达，抑制细菌粘附
DOI：
10.1074/jbc.m112.408245
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发表时间：
2013-02-22
影响因子：
4.8
通讯作者：
Wang, Rongquan
中科院分区：
文献类型：
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作者：
Pan, Qiong;Tian, Yin;Wang, Rongquan
我们研究了基于来自乳腺相关血清淀粉样蛋白A3（M-SAA 3）的保守TFLK基序的合成四聚体分支肽是否比单体肽更有效地上调MUC 3表达，并检查了该过程的可能机制和生物学意义。我们使用标准的固相方法来合成含有三赖氨酸核心的四聚体分支肽（序列GWLTFL-KAAG），称为含有TFLK的10聚体BP。使用转录因子蛋白/DNA阵列分析转录因子的异常表达。采用实时定量PCR和/或Western印迹法检测MUC 3及相关转录因子。荧光素酶测定、EMSA和ChIP测定用于分析人MUC 3启动子的活性。采用细菌粘附试验评价其对致病性大肠杆菌和肠出血性大肠杆菌的体外抑制作用。大肠杆菌血清型O 157：H7（EHEC O 157：H7）在用含TFLK的10-mer BP处理后粘附于HT-29-Gal细胞。在HT-29-Gal细胞中，含TFLK的10-mer BP比M-SAA 3衍生的N-末端10-mer单体肽诱导更高水平的MUC 3表达，并且MUC 3表达通过转录机制激活，包括诱导多个转录因子并进一步与MUC 3启动子内核苷酸-242和-62之间的顺式元件结合。有趣的是，含TFLK的10-mer BP显著抑制肠致病性E。大肠埃希菌和肠出血性大肠杆菌O 157：H7对HT-29-Gal细胞的粘附性。这一发现表明这种肽在预防胃肠道感染方面具有潜在的治疗用途。
We investigated whether a synthetic tetrameric branched peptide based on the conserved TFLK motif from mammary-associated serum amyloid A3 (M-SAA3) is more efficient than the monomeric peptide at up-regulating MUC3 expression and examined the possible mechanism(s) and biological significance of this process. We used standard solid-phase methods to synthesize a tetrameric branched peptide (sequence GWLTFL-KAAG) containing a trilysine core, termed the TFLK-containing 10-mer BP. The aberrant expression of transcription factors was analyzed using a transcription factor protein/DNA array. MUC3 and relevant transcription factors were detected using real-time PCR and/or Western blots. The luciferase assay, EMSA, and ChIP assays were used to analyze the activity of the human MUC3 promoter. The bacterial adherence assay was used to evaluate the in vitro inhibition of enteropathogenic Escherichia coli or enterohemorrhage E. coli serotype O157:H7 (EHEC O157:H7) adherence to HT-29-Gal cells after treatment with the TFLK-containing 10-mer BP. In HT-29-Gal cells, the TFLK-containing 10-mer BP induced higher levels of MUC3 expression than the M-SAA3-derived N-terminal 10-mer monomeric peptide, and MUC3 expression was activated through transcriptional mechanisms, including the induction of multiple transcription factors and further binding with their cis-elements between nucleotides -242 and -62 within MUC3 promoter. Interestingly, the TFLK-containing 10-mer BP dramatically inhibited enteropathogenic E. coli and EHEC O157:H7 adherence to the HT-29-Gal cells compared with the controls. This finding suggests a potential therapeutic use for this peptide to prevent gastrointestinal infection.