title: Expression profiling, gene silencing and transcriptional networking of metzincin metalloproteases in the cattle tick, Rhipicephalus (Boophilus) microplus.
abstract: Tick proteins functioning in vital physiological processes such as blood meal uptake, digestion and reproduction are potential targets for anti-tick vaccines, since vaccination could inhibit these essential functions and ultimately affect tick survival. In this study we identified metzincin metalloproteases from Rhipicephalus microplus as potential vaccine candidates since they are implicated as essential to blood-cavity formation, bloodmeal digestion and reproduction in ixodid ticks. Eight transcripts encoding proteins that contain the characteristic metzincin zinc-binding motif HEXXHXXG/NXXH/D and a unique methionine containing "methionine-turn" were identified from native and in-house assembled R. microplus expressed sequence tag (EST) databases. These were representative of five reprolysin-like and three astacin-like metzincin metalloproteases. Reverse transcription-PCR analysis indicated that the reprolysins were most abundantly expressed in the salivary glands, whereas the astacins were most abundant in the midgut and ovaries. In vivo gene silencing was performed to assess a possible phenotype of these metalloproteases during adult female R. microplus blood feeding and reproduction. RNA interference (RNAi) against two of the reprolysins and one of the astacins significantly affected the average egg weight and oviposition rate. Evidently, this reverse genetic approach enabled the evaluation of the overall vital impact of tick proteins. Finally, integrated real time-PCR studies also revealed an extensive cross organ network between the R. microplus metzincin transcripts, supporting the use of a combinatorial metzincin-based anti- R. microplus vaccine.
authors.affiliation: Department of Biochemistry, Faculty of Natural and Agricultural Sciences, University of Pretoria, Pretoria, 0002, South Africa.
authors.forename: Annette-Christi
authors.initials: AC
authors.lastname: Barnard
authors.forename: Ard M
authors.initials: AM
authors.lastname: Nijhof
authors.forename: Anabella R M
authors.initials: AR
authors.lastname: Gaspar
authors.forename: Albert W H
authors.initials: AW
authors.lastname: Neitz
authors.forename: Frans
authors.initials: F
authors.lastname: Jongejan
authors.forename: Christine
authors.initials: C
authors.lastname: Maritz-Olivier
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D060829:Arthropod Proteins / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D002418:Cattle Diseases / Q000469:parasitology; D005260:Female; D020869:Gene Expression Profiling; D020868:Gene Silencing; D008297:Male; D045726:Metalloproteases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010802:Phylogeny; D012330:RNA, Double-Stranded; D048494:Rhipicephalus / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology*
chemical_list: D060829:Arthropod Proteins; D012330:RNA, Double-Stranded; D045726:Metalloproteases