Molecular characterization of a myosin alkali light chain-like protein, a "concealed" antigen from the hard tick Haemaphysalis qinghaiensis.
There should be some differences between antibodies generated by feeding ticks on animals and those derived by immunizing animals with tick extracts. Here, we found serum collected from sheep immunized with Haemaphysalis qinghaiensis salivary gland extracts could detect two more protein bands with molecular weights of 22 and 37 kDa (P22 and P37) on Western blots of extracts of tick salivary glands than serum from tick infected animals. Rabbit anti-H. qinghaiensis differential protein immune serum was then generated from P22 and P37 and was used to immunoscreen a cDNA library constructed from salivary glands, Malpighian tubules and ovaries of partially engorged H. qinghaiensis. A cDNA contains an open reading frame of 483 bp that codes for 160 amino acid residues with a coding capacity of 18 kDa was cloned and designated Hq02. Expression analysis by RT-PCR showed that this gene is expressed in salivary glands, midguts, other organs and different developmental stages of H. qinghaiensis. The predicted amino acid sequence of the Hq02 gene had high homology to some known myosin alkali light chain (MLC) proteins. A fusion protein consisting of 130 amino acids of Hq02 protein and 335 amino acids of T7 gene 10 protein was expressed in Escherichia coli and used to immunize sheep. Western blot showed that only rabbit anti-H. qinghaiensis differential protein immune serum could recognize the expressed Hq02 protein, while rabbit anti-H. qinghaiensis saliva immune could not. This proved Hq02 protein was a "concealed" antigen. Immunization with the recombinant Hq02 conferred a 21.8% reduction of engorgement weight for adult female ticks that fed on the immunized sheep. This is the first report of tick myosin alkali light chain and the function of this protein is discussed.
Gao
Jinliang
J
Key Laboratory of Veterinary Parasitology of Gansu Province, State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China.
Luo
Jianxun
J
Fan
Ruiquan
R
Guan
Guiquan
G
Ren
Qiaoyun
Q
Ma
Miling
M
Sugimoto
Chihiro
C
Bai
Qi
Q
Yin
Hong
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D020869:Gene Expression Profiling / Q000662:veterinary; D018507:Gene Expression Regulation, Developmental; D015723:Gene Library; D026863:Ixodidae / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D018994:Myosin Light Chains / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000276:immunology; D012469:Salivary Glands / Q000737:chemistry / Q000276:immunology; D016415:Sequence Alignment / Q000662:veterinary; D017422:Sequence Analysis, DNA; D012756:Sheep; D012757:Sheep Diseases / Q000276:immunology*; D013984:Tick Infestations / Q000276:immunology / Q000662:veterinary*; D014614:Vaccines, Synthetic / Q000276:immunology
D018994:Myosin Light Chains; D011993:Recombinant Fusion Proteins; D014614:Vaccines, Synthetic