Inducible gene expression of moricin, a unique antibacterial peptide from the silkworm (Bombyx mori)
Inducible gene expression of moricin, a unique antibacterial peptide from the silkworm (Bombyx mori)
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DOI：
10.1042/0264-6021:3400265
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发表时间：
1999-05-15
影响因子：
4.1
通讯作者：
Yamakawa, M
中科院分区：
文献类型：
--
作者：
Furukawa, S;Tanaka, H;Yamakawa, M
利用家蚕脂肪体cDNA文库克隆了家蚕抗菌肽moricin的cDNA。将编码moricin的部分核苷酸序列的逆转录PCR产物用作探针。四个阳性克隆的核苷酸测序揭示了两种类型的命名为moricin 1和2的moricin cDNA。moricin 1和moricin 2的cDNA核苷酸序列同源性为97.2%。尽管推测的信号肽中的moricin I的一个氨基酸残基（Phe（6））被moricin 2中的Lys（6）取代，但它们的成熟部分的氨基酸序列是相同的。B中Moricin基因表达。在脂肪体、血细胞和马氏管中观察到注射大肠杆菌的家蚕幼虫，但在其他组织如中肠和丝腺中没有观察到。用E.大肠杆菌在注射后8h达到最高水平，并持续至48 h。结果表明，大肠杆菌的细胞壁成分脂多糖（LPS）和脂质A（lipid A）在大肠杆菌中的表达量明显高于大肠杆菌。coli中，引发moricin基因的表达。用特异性引物对moricin 1和moricin 2基因进行PCR比较，结果表明moricin 2基因的表达量高于moricin 1基因。从B中筛选编码moricin 2的基因组克隆。以桑菌素cDNA为探针，构建了桑细菌基因组文库。基因表达调控基序如核因子-κ B结合位点样序列（KB位点）和核因子-白细胞介素-6结合位点样序列（NF-IL-6位点）位于5 '上游调控区。细胞迁移率改变分析表明，存在细菌LPS诱导的核蛋白，其可以结合到调节区的κ B位点和其他位点。
Molecular cloning of cDNAs encoding moricin, a novel antibacterial peptide from the silkworm (Bombyx mori), was performed using a fat-body cDNA library. A reverse-transcription PCR product encoding a partial nucleotide sequence of moricin was used as a probe. Nucleotide sequencing of four positive clones revealed two types of moricin cDNAs designated moricin 1 and 2. cDNAs for moricin 1 and 2 shared 97.2% identity in their nucleotide sequences. Although one amino acid residue (Phe(6)) of moricin I in the putative signal peptide was replaced with Lys(6) in moricin 2, amino acid sequences of their mature portions were identical. Moricin gene expression in B. mori larvae injected with Escherichia coli was observed in fat-bodies, haemocytes and the Malpighian tubule, but not in other tissues like the midgut and silk glands. Accumulation of moricin gene transcripts induced by E. coli reached a maximum level 8 h after injection and persisted up to 48 h. It was confirmed that lipopolysaccharide (LPS) and lipid A, which are cell-wall components of E. coli, triggered moricin gene expression. Comparison of gene expression between moricin 1 and 2 by PCR using specific primers indicated that moricin 2 gene was more strongly expressed than moricin 1 gene. A genomic clone encoding moricin 2 was screened from a B. mori genomic library using a moricin cDNA as a probe. Regulatory motifs for gene expression such as nuclear-factor-kappa B-binding-site-like sequence (KB site) and nuclear-factor-interleukin-6-binding-site-like sequence (NF-IL-6 site) were found in the 5'-upstream regulatory region. An electrophoretic-mobility-shift assay revealed that there are bacterial LPS-inducible nuclear proteins that can bind to the kappa B Site and other sites in the regulatory region.