The kinetoplast ultrastructural organization of endosymbiont-bearing trypanosomatids as revealed by deep-etching, cytochemical and immunocytochemical analysis.
; Thiry, Marc ; et al
in Histochemistry & Cell Biology (2008), 130(6), 1177-85
The endosymbiont-bearing trypanosomatids present a typical kDNA arrangement, which is not well characterized. In the majority of trypanosomatids, the kinetoplast forms a bar-like structure containing ... [more ▼]
The endosymbiont-bearing trypanosomatids present a typical kDNA arrangement, which is not well characterized. In the majority of trypanosomatids, the kinetoplast forms a bar-like structure containing tightly packed kDNA fibers. On the contrary, in trypanosomatids that harbor an endosymbiotic bacterium, the kDNA fibers are disposed in a looser arrangement that fills the kinetoplast matrix. In order to shed light on the kinetoplast structural organization in these protozoa, we used cytochemical and immunocytological approaches. Our results showed that in endosymbiont-containing species, DNA and basic proteins are distributed not only in the kDNA network, but also in the kinetoflagellar zone (KFZ), which corresponds to the region between the kDNA and the inner mitochondrial membrane nearest the flagellum. The presence of DNA in the KFZ is in accordance with the actual model of kDNA replication, whereas the detection of basic proteins in this region may be related to the basic character of the intramitochondrial filaments found in this area, which are part of the complex that connects the kDNA to the basal body. The kinetoplast structural organization of Bodo sp. was also analyzed, since this protozoan lacks the highly ordered kDNA-packaging characteristic of trypanosomatid and represents an evolutionary ancestral of the Trypanosomatidae family. [less ▲]Detailed reference viewed: 39 (0 ULg)
Immunocytochemical detection of DNA and RNA in endosymbiont-bearing trypanosomatids.
; ; Thiry, Marc
in FEMS Microbiology Letters (2003), 221(1), 17-23
Research about the kinetoplast of trypanosomatids has yielded valuable information about the organization of extranuclear structure. However, the ultrastructural localization of nucleic acids within these ... [more ▼]
Research about the kinetoplast of trypanosomatids has yielded valuable information about the organization of extranuclear structure. However, the ultrastructural localization of nucleic acids within these protozoa remains uncertain. We have applied cytochemical and immunocytochemical approaches to precisely identify DNA and RNA in lower endosymbiont-bearing trypanosomatids. Using the Terminal deoxynucleotidyl Transferase (TdT) immunogold technique, we showed that nuclear DNA is seen associated with the nuclear envelope during the trypanosomatid cell cycle. By combining the TdT technique with the acetylation method, which improves the contrast between structures containing fibrils and granules, we have demonstrated that the nucleolus of endosymbiont-bearing trypanosomatids is composed of two constituents: a granular component and a DNA-positive fibrillar zone. Moreover, we revealed that DNA of endosymbiotic bacteria consisted of electron-dense filaments which are usually in close contact with the prokaryote envelope. Using a Lowicryl post-embedding immunogold labeling procedure with anti-RNA antibodies, we showed the presence of RNA not only over the cytoplasm, the interchromatin spaces and the nucleolus, but also over the kinetoplast and virus-like particles present in Crithidia desouzai. [less ▲]Detailed reference viewed: 5 (0 ULg)
Chromosome localization changes in the Trypanosoma cruzi nucleus.
; ; et al
in Eukaryotic Cell (2002), 1(6), 944-53
Chromosome localization in the interphase nuclei of eukaryotes depends on gene replication and transcription. Little is known about chromosome localization in protozoan parasites such as trypanosomes ... [more ▼]
Chromosome localization in the interphase nuclei of eukaryotes depends on gene replication and transcription. Little is known about chromosome localization in protozoan parasites such as trypanosomes, which have unique mechanisms for the control of gene expression, with most genes being posttranscriptionally regulated. In the present study, we examined where the chromosomes are replicated in Trypanosoma cruzi, the agent of Chagas' disease. The replication sites, identified by the incorporation of 5-bromodeoxyuridine, are located at the nuclear periphery in proliferating epimastigote forms in the early S phase of the cell cycle. When the S phase ends and cells progress through the cell cycle, 5-bromodeoxyuridine labeling is observed in the nuclear interior, suggesting that chromosomes move. We next monitored chromosome locations in different stages of the cell cycle by using a satellite DNA sequence as a probe in a fluorescence in situ hybridization assay. We found two distinct labeling patterns according to the cell cycle stage. The first one is seen in the G(1) phase, in hydroxyurea-arrested epimastigotes or in trypomastigotes, which are differentiated nondividing forms. In all of these forms the satellite DNA is found in dots randomly dispersed in the nucleus. The other pattern is found in cells from the S phase to the G(2) phase. In these cells, the satellite DNA is found preferentially at the nuclear periphery. The labeling at the nuclear periphery disappears only after mitosis. Also, DNA detected with terminal deoxynucleotidyl transferase is found distributed throughout the nuclear space in the G(1) phase but concentrated at the nuclear periphery in the S phase to the G(2) phase. These results strongly suggest that T. cruzi chromosomes move and, after entering the S phase, become constrained at the nuclear periphery, where replication occurs. [less ▲]Detailed reference viewed: 7 (0 ULg)