title: The stage-regulated expression of Leishmania mexicana CPB cysteine proteases is mediated by an intercistronic sequence element.
abstract: The tandemly arranged CPB genes of Leishmania mexicana are polycistronically transcribed and encode cysteine proteases that are differentially stage-specific; CPB1 and CPB2 are expressed predominantly in metacyclics, whereas CPB3-CPB18 are expressed mainly in amastigotes. The mechanisms responsible for this differential expression have been studied via gene analysis and re-integration of individual CPB genes, and variants thereof, into a CPB-deficient parasite mutant. Comparison of the nucleotide sequences of the repeat units of CPB1 and CPB2 with CPB2.8 (typical of CPB3-CPB18) revealed two major regions of divergence as follows: one of 258 base pairs (bp) corresponding to the C-terminal extension of CPB2.8; another, designated InS, of 120 bp, with insertions totaling 57 bp, localized to the intercistronic region downstream of CPB1 and CPB2. Cell lines expressing CPB2.8 or CPB2 with the 3'-untranslated region and intercistronic sequence of CPB2.8 showed up-regulation in amastigotes. Conversely, metacyclic-specific expression occurred with CPB2 or CPB2.8 with the 3'-untranslated region and intercistronic sequence of CPB2. Moreover, the InS down-regulated expression in amastigotes of a reporter gene integrated into the CPB locus. It is proposed that the InS mediates metacyclic-specific stage-regulated expression of CPB by affecting the maturation of polycistronic pre-mRNA. This is the first well defined cis-regulatory element implicated in post-transcriptional stage-specific gene expression in Leishmania.
authors.affiliation: Wellcome Centre for Molecular Parasitology, University of Glasgow, the Anderson College, 56 Dumbarton Road, Glasgow G11 6NU, Scotland, United Kingdom.
authors.forename: D R
authors.initials: DR
authors.lastname: Brooks
authors.forename: H
authors.initials: H
authors.lastname: Denise
authors.forename: G D
authors.initials: GD
authors.lastname: Westrop
authors.forename: G H
authors.initials: GH
authors.lastname: Coombs
authors.forename: J C
authors.initials: JC
authors.lastname: Mottram
mesh_terms: D020413:3' Untranslated Regions; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D015536:Down-Regulation; D005780:Gelatin / Q000737:chemistry; D005786:Gene Expression Regulation; D005796:Genes; D017930:Genes, Reporter; D007894:Leishmania mexicana / Q000201:enzymology*; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011061:Poly A / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014162:Transfection
chemical_list: D020413:3' Untranslated Regions; D012333:RNA, Messenger; D011061:Poly A; D005780:Gelatin; D015500:Chloramphenicol O-Acetyltransferase; D003546:Cysteine Endopeptidases