title: Functional analysis of a highly conserved abundant larval transcript-2 (alt-2) intron 2 repeat region of lymphatic filarial parasites.
abstract: The filarial-specific protein abundant larval transcript-2 (ALT-2) is expressed exclusively in the infective larval stage (L3) and is a crucial protein for establishing immunopathogenesis in human hosts. The alt-2 gene has a conserved minisatellite repeat (29 or 27bp) in intron 2 (IR2) whose significance within lymphatic filarial species is unknown. Here, we report the role of IR2 in the regulation of alt-2 gene expression using an in vitro model. Using electrophoretic mobility shift assays, we identified the presence of a putative nuclear protein binding region within IR2. Subsequent transient expression experiments in eukaryotic cell lines demonstrated that the IR2 downregulated the expression of a downstream luciferase reporter gene, which was further validated with RT-PCR. We therefore identify IR2 as a suppressor element that regulates L3 stage-specific expression of alt-2.
authors.affiliation: Centre for Biotechnology, Anna University, Chennai, Tamil Nadu 600 025, India.
authors.forename: Moorthy
authors.initials: M
authors.lastname: Sakthidevi
authors.affiliation: Vector Control Research Centre, Puducherry 605 006, India.
authors.forename: Sugeerappa Laxmanappa
authors.initials: SL
authors.lastname: Hoti
authors.affiliation: Centre for Biotechnology, Anna University, Chennai, Tamil Nadu 600 025, India. Electronic address: pk.kaliraj@gmail.com.
authors.forename: Perumal
authors.initials: P
authors.lastname: Kaliraj
mesh_terms: D000818:Animals; D000947:Antigens, Helminth / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D017178:Brugia malayi / Q000235:genetics* / Q000276:immunology; D016466:CHO Cells; D002460:Cell Line; D003412:Cricetulus; D018509:DNA, Helminth / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D004605:Elephantiasis, Filarial / Q000469:parasitology*; D005786:Gene Expression Regulation; D017930:Genes, Reporter / Q000235:genetics; D015801:Helminth Proteins / Q000235:genetics; D056945:Hep G2 Cells; D006801:Humans; D057131:Immune Evasion; D007438:Introns / Q000235:genetics*; D007814:Larva / Q000235:genetics; D008156:Luciferases / Q000235:genetics; D018598:Minisatellite Repeats / Q000235:genetics; D009687:Nuclear Proteins / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D061987:Sf9 Cells; D037941:Silencer Elements, Transcriptional / Q000235:genetics; D018411:Spodoptera; D014958:Wuchereria bancrofti / Q000235:genetics* / Q000276:immunology
chemical_list: C521555:ALT-2 protein, Brugia malayi; D000947:Antigens, Helminth; D018509:DNA, Helminth; D015801:Helminth Proteins; D009687:Nuclear Proteins; D011994:Recombinant Proteins; D008156:Luciferases
keywords: Evolution; Gene regulation; Intron DNA repeat; Silencer; Transcription