Isolation and characterization of immune-related genes from the fall webworm, Hyphantria cunea, using PCR-based differential display and subtractive cloning

• Published in: Insect biochemistry and molecular biology Vol. 28; no. 11; pp. 827 - 837
• Main Authors: Woon Shin, Sang, Park, Soon-Sik, Park, Doo-Sang, Gwang Kim, Mi, Sun Chang Kim, Brey, Paul T, Park, Ho-Yong
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.11.1998
• Subjects: Amino Acid Sequence; Animals; Arctiidae; Attacin; Bombyx mori; Cloning, Molecular; DNA, Complementary - chemistry; DNA, Complementary - isolation & purification; Escherichia coli - genetics; Extracellular copper; Genes, Insect; Gram negative binding protein (GNBP); Hemolin; Hyphantria cunea; Lectin; Lectins - genetics; Locusta migratoria; Molecular Sequence Data; Moths - genetics; Moths - immunology; Moths - microbiology; Polymerase Chain Reaction; RNA, Messenger - analysis; Sequence Alignment; Serine protease inhibitor (serpin); Serpins - genetics; Superoxide Dismutase - genetics; Zinc superoxide dismutase (Cu–Zn SOD); Zinc superoxide dismutase (Cu–Zn SOD); Lectin; Serine protease inhibitor (serpin); Gram negative binding protein (GNBP); Hemolin; Extracellular copper; Attacin
• ISSN: 0965-1748; 1879-0240
• DOI: 10.1016/S0965-1748(98)00077-0

Following injection of bacteria into the hemocoel of the fall webworm, Hyphantria cunea, several inducible genes were identified and characterized using PCR-based differential display (DD-PCR) and subtractive cloning. Ten immune-related cDNA clones (Hdd1, Hdd2, Hdd3, Hdd11, Hdd13, Hdd15, Hdd17, Hdd23, Hs106, Hs302) were isolated and characterized. The deduced amino acid sequence of Hdd2 was shown to be a member of the copper, zinc superoxide dismutase (Cu–Zn SOD) family. The H. cunea Cu–Zn SOD is novel in that it is up-regulated following a bacterial challenge and has a putative signal peptide suggesting its secretion and involvement in the insect immune response. Hdd3 was found to encode a new member of the serpin (serine protease inhibitor) family. The putative lectin corresponding to Hdd15 is of a different kind in that it has two lectin C domains in a single molecule. These two lectin C domains show significant homology to the lectin C domain of Periplaneta lipopolysaccharide binding protein (LPS-BP). Three cloned genes, Hdd17, Hs106 and Hs302, encode a homologue to Bombyx mori Gram negative binding protein, a hemolin-like protein and a attacin-like protein, respectively. The deduced amino acid sequences from Hdd11 showed weak homology with a Locusta migratoria hemolymph protein. On the contrary, Hdd1, Hdd13 and Hdd23 did not reveal any significant homology with known proteins. All of the 10 genes were clearly inducible by E. coli and M. luteus injection. Injection of distilled water only slightly induced mRNA levels. Comparison of temporal mRNA expression following E. coli injection showed three types of expression patterns.