Hitchhiking Trypanosoma cruzi minicircle DNA affects gene expression in human host cells via LINE-1 retrotransposon

Abstract

Full-length RNA transcribed from the human LINE-1 (Li) element Li Homo sapiens (LlHs) has a 900-nt, G+C-rich, 5'-untranslated region (UTR). The 5' UTR is followed by two long open reading frames, ORF1 and ORF2, which are separated from each other by an inter-ORF region of 33 nt that includes two or three in-frame stop codons. We examine here the mechanism(s) by which the translation of LlHs ORF1 and ORF2 is initiated. A stable hairpin structure (AG =-74.8 kcal/mol), inserted at nt 661 of the 5' UTR, caused a 3-to 8-fold decrease in the in vitro and in vivo translation ofeither a lacZ reporter gene for ORF1 or the ORF1 polypeptide product, p40, but translation of a lacZ reporter gene in ORF2 was increased. The results are compatible with a model for ORF1 translation initiation in which the majority of ribosomes scan from a point 5' of nt 661 but suggest that ORF2 is not translated by attached ribosomes that reinitiate after the termination of ORF1 translation. Our data are compatible with a model whereby the translation of LlHs ORF2 is initiated internally. The human LINE-1 (Li) element (Li Homo sapiens; LiHs) is the only known transposable element that is endogenous to the human genome. The structural features of LlHs (sche-matically presented in Fig.

Pathogenesis of Chagas' Disease: Parasite Persistence and Autoimmunity

“…Primary data on the integration of parasite DNA into vertebrate genomes have taken on a controversial nature [259a], with justifications that are not reasonable to many in the scientific community. The original work has been supported subsequently by additional published studies [173,366,402], of humans, rabbits, and chickens, from the authors' laboratory. After six years, the editor did not show experimental data to refute the original observations of kDNA integration.…”

“…These findings suggest that the kDNA minicircle is integrated into the macrophage genome (412). The topology of the minicircle at the integration junction site is as yet unknown, but again, the chance of two integration events occurring at a short distance favors the PCR amplification of the host DNA flanked by the kDNAs at both ends (366). In this context, the amplification of the chimeric kDNA-host DNA is obtained with primer sets annealing to the flanking kDNAs at both ends.…”

Chagas Heart Disease Pathogenesis: One Mechanism or Many?

“…Using this highly sensitive method, the authors identified what they interpreted as an absolute correlation between parasite persistence and tissue inflammation [16]. Simões-Barbosa et al later found evidence that the horizontal transfer of T. cruzi mitochondrial kDNA to the genomes of naturally infected humans via integration into LINE-1 retrotransposons may play an important role in the pathogenesis of CHD [17]. Although these findings have provided the strongest evidence to date that parasite persistence is crucial for the protracted progression of CHD pathology, no distinction could be made between antigens present on intact T. cruzi in the tissue and lingering T. cruzi antigens or DNA that had been shed from parasites no longer present in the proximate tissue.…”

Evolution and pathology in Chagas disease: a review

“…At issue was the site of integration into the vertebrate genomes. We continue to challenge our conclusions experimentally (Simões-Barbosa et al 2006), and aim to resolve any questions to the satisfaction of the scientific community. (Margulis et al 2000).…”

“…An answer to the origin of autoimmunity in Chagas disease was sought in genomic DNA from the human macrophage line U937 that had been infected (ratio 1:5) with trypomastigoes of T. cruzi (Simões-Barbosa et al 2006). All samples assayed harbored parasite DNA as judged by NAT molecular markers to T. cruzi.…”