Inheritance of DNA Transferred from American Trypanosomes to Human Hosts
Figure 3
LkDT in somatic cells suggests mechanism of kDNA integration by homologous recombination.
(A) Representative PCR amplifications of T. cruzi kDNA and nDNA. kDNA and nDNA signatures indicate active infections and LkDTs, while only kDNA indicates LkDT. Amplification products obtained with kDNA (S35/S36) and nDNA (Tcz1/Tcz2) primer sets hybridized with specific internal probes S67 and Tcz3, respectively, on blots of 1% agarose gels. The numbers above each lane indicates the study case. Tc, T. cruzi. Patients 10, 34, 51, 61 and 71, showing nDNA and kDNA signatures, harbor active T. cruzi infections. Patients 9, 36, 50 and 60, with only the kDNA signature, may carry inherited integration events. (B) Southern hybridization of integrated kDNA revealed by a minicircle (kCR) probe. A 0.8% agarose gel was used to analize EcoRI-digested DNA from Chagas patient blood mononuclear cells. Top numbers indicate family founders who had the kDNA integrations in LINE-1 on chromosome X (Table S3). DNA from uninfected human donors (C2 to C5) and from T. cruzi were included as controls. (C) Schematic representation of microhomology-mediated end-joining kDNA minicircle integration into retrotransposon LINE-1. Patients 71 (top) and 24 (bottom) show AC-rich intermediates involved in recombination. Each patient showing nDNA and/or kDNA footprints yielded at least one chimeric sequence.