Spatial distribution of AT- and GC-rich DNA within interphase cell nuclei of Triatoma infestans Klug

• Published in: Micron (Oxford, England : 1993) England : 1993), 2011-08, Vol.42 (6), p.568-578
• Main Authors: Alvarenga, Elenice M., Mondin, Mateus, Martins, James A., Rodrigues, Vera L.C.C., Vidal, Benedicto C., Rincones, Johana, Carazzolle, Marcelo F., Andrade, Larissa Mara, Mello, Maria Luiza S.
• Format: Article
• Language: English
• Subjects: 4',6-diamidino-2-phenylindole; Animals; Base Composition; Bioinformatics; Cell Nucleus - chemistry; Chagas disease; Chromatin; chromocenters; chromosome banding; confocal microscopy; cytosine; differential staining; DNA; DNA - analysis; DNA Methylation; Epigenetics; Hemiptera; Heterochromatin; image analysis; Interphase; Microscopy; Restriction Mapping; Triatoma - chemistry; Triatoma - cytology; Triatoma infestans
• ISSN: 0968-4328; 1878-4291
• DOI: 10.1016/j.micron.2011.02.002

► Somatic cell nuclei of Triatoma infestans contain large heterochromatin bodies. ► We studied the spatial localization of AT- and GC-rich DNA in this model. ► Topochemistry, immuno assay, cytidine demethylation, and microphotometry were used. ► These methods plus bioinformatics revealed AT-richness mainly in the heterochromatin. ► DNA methylation is not responsible for the packing state of this heterochromatin type. Heterochromatin bodies in single- and multichromocentered interphase cell nuclei of Triatoma infestans, a vector of Chagas disease, have been suggested to contain AT-rich DNA, based on their positive response to Q-banding and Hoechst 33248 treatment. No information exists on whether GC-rich DNA is also present in these nuclei and whether it plays a role on chromatin condensation. Considering that methodologies more precise than those previously used to determine DNA base composition in situ are currently available, and that the spatial distribution of chromatin areas differing in composition in interphase cell nuclei of different species is a matter of interest, the localization of AT- and GC-rich DNA in T. infestans nuclei is revisited here. The methodologies used included DAPI/AMD and CMA 3/Distamycin differential staining, Feulgen-DNA image analysis following Msp I and Hpa II enzymatic digestion, 5-methylcytidine immunodetection, AgNOR response, confocal microscopy, and the 5-aza-2′-deoxycytidine (5-AZA) demethylation assay. The results identified the presence of AT-rich/GC-poor DNA in chromocenters and evenly distributed AT and GC sequences in euchromatin. A GC-rich DNA zone encircling the chromocenters was also found but it could not be associated with NOR regions. To corroborate the DNA AT-richness in T. infestans nuclei, bioinformatic analyses were also performed. Methylated cytosine was evident at some points of the chromocenters’ edge in single- and multichromocentered nuclei and at the euchromatin of multichromocentered nuclei and could be transiently affected by the 5-AZA treatment. The present results suggest that in the particular case of chromocenters of the hemipteran T. infestans, cytosine methylation is not a relevant factor involved in chromatin condensation.