Preferential translation of Hsp83 in Leishmania requires a thermosensitive polypyrimidine-rich element in the 3′ UTR and involves scanning of the 5′ UTR

Abstract

Heat shock proteins (HSPs) provide a useful system for studying developmental patterns in the digenetic Leishmania parasites, since their expression is induced in the mammalian life form. Translation regulation plays a key role in control of protein coding genes in trypanosomatids, and is directed exclusively by elements in the 39 untranslated region (UTR). Using sequential deletions of the Leishmania Hsp83 39 UTR (888 nucleotides [nt]), we mapped a region of 150 nt that was required, but not sufficient for preferential translation of a reporter gene at mammalian-like temperatures, suggesting that changes in RNA structure could be involved. An advanced bioinformatics package for prediction of RNA folding (UNAfold) marked the regulatory region on a highly probable structural arm that includes a polypyrimidine tract (PPT). Mutagenesis of this PPT abrogated completely preferential translation of the fused reporter gene. Furthermore, temperature elevation caused the regulatory region to melt more extensively than the same region that lacked the PPT. We propose that at elevated temperatures the regulatory element in the 39 UTR is more accessible to mediators that promote its interaction with the basal translation components at the 59 end during mRNA circularization. Translation initiation of Hsp83 at all temperatures appears to proceed via scanning of the 59 UTR, since a hairpin structure abolishes expression of a fused reporter gene.

Gene duplication in trypanosomatids – Two DED1 paralogs are functionally redundant and differentially expressed during the life cycle

“…The product was cloned into the pH-CAT-H cassette [43] through its BamHI and XbaI sites (list of primers in Table A.1) generating pH-LeishIF4G3-FLAG-H.…”

Two related trypanosomatid eIF4G homologues have functional differences compatible with distinct roles during translation initiation

“…6 Selective regulation of gene expression acts on mRNA levels and efficiency of translation. [7][8][9] There are several examples of regulation of translation in trypanosomatids [10][11][12][13][14] and many others are likely to emerge, but the molecular mechanisms associated with this regulation are not well understood. In higher eukaryotes most translation regulation events occurs at the level of translation initiation when selection of the mRNAs by the protein synthesis apparatus takes place.…”

Post-Transcriptional Regulation of the Trypanosome Heat Shock Response by a Zinc Finger Protein

“…The multiple copies of the HSP83 genes (encoding the major Kinetoplastid HSP90 homologue) are also in a tandem array. The 3′-UTR of Leishmania HSP83 mRNA is important for both mRNA stability and increased translation during heat-shock [26], and it was proposed that temperature-induced changes in RNA secondary structure might play a role in regulation [27]. Post-transcriptional mechanisms are also responsible for heat-induced increases in Leishmania HSP100 mRNA [28], [29].…”