Comparative Analysis (Hippotragini versus Caprini, Bovidae) of X-Chromosome's Constitutive Heterochromatin by in situ Restriction Endonuclease Digestion: X-Chromosome Constitutive Heterochromatin Evolution

Abstract

The Bovidae X-chromosome shows a considerable variation, in contrast to the preservative autosomal conservatism. The X-chromosome variation is mostly a consequence of the constitutive heterochromatin (CH) variation; in what respect to its amount and position. This is especially common among the non-Bovinae subfamilies and tribes. In order to characterize the X-chromosome CH in non-Bovinae species -Hippotragini and Caprini tribes -we have used restriction endonuclease digestion on fixed chromosomes and sequential C-banding. With these techniques we were able to distinguish between the two X-chromosome types (Hippotragini and Caprini) CH, in what respect to its position and molecular nature. Moreover, we define at least, six subclasses of CH in both X-chromosome types analyzed. Evolutionary considerations were draw based on the results obtained. The technology used here for the analysis of the Bovidae X-chromosome CH showed to be more evolutionary informative than the classical approaches.Abbreviations: BAC -bacterial artificial chromosomes; CBP-banding -C-bands by barium hydroxide using propidium iodide; CH -constitutive heterochromatin; Chr -chromosome; DAPI -4¢-6-diamidino-2-phenylindole; DNA -deoxyribonucleic acid; FISH -fluorescent in situ hybridization; GTD-banding -G-bands by trypsin with DAPI; MYBP -million years ago before present; NAA -number of autosomal arms; PBS -phosphate-buffered saline; RE-banding -restriction endonuclease banding; RE-bands -restriction endonuclease bands; RE -restriction endonuclease; REs -restriction endonucleases; SSC -saline sodium citrate.

Chromosomal evolution and phylogenetic analyses in Tayassu pecari and Pecari tajacu (Tayassuidae): tales from constitutive heterochromatin

“…When we applied REs and sequential C-banding to the chromosomes, it was possible to verify the C-band heterogeneity ( figure 1), similarly to what has been observed in other species (Chaves et al 2004;Adega et al 2005).…”

“…These bands were not disclosed by classical C-banding procedures. These C-positive bands, only disclosed by previous RE treatments, could be explained by the fact that RE digestion might increase the capacity of the stain (in the present work, propidium iodide) to bind to a specific chromosome region (Gosálvez et al 1997;Nieddu et al 1999;Chaves et al 2004). Most of these bands are interstitial.…”

“…The higher number of Cpositive heterochromatin classes/subclasses on P. tajacu autosomes suggests that this karyotype is more primitive, that is it did not go through a high level of homogenization due to concerted evolution of chromosomes. This karyotype resembles the ancestral form, displaying a high diversity of CH, which is thought to facilitate the occurrence of chromosomal rearrangements (Yunis and Yasmineh 1971;Peacock et al 1982;John 1988;Chaves et al 2004).…”

“…Based on these two works, and on the assumption that the pres-ence of CH facilitates the occurrence of chromosomal rearrangements (Yunis and Yasmineh 1971;Peacock et al 1982;John 1988;Chaves et al 2004), we made a com-parative analysis between T. pecari and P. tajacu CH lo-calization and the breakpoints in each chromosome (figure 3). In figure 3, the corresponding breakpoints (assigned by the pig paint probes) in T. pecari and P. tajacu were named with letters in the G-banded karyotype.…”

“…molecular nature of CH is the central concern. Among these, use of restriction endonucleases (which cleave DNA at specific sequences) and use of base-specific fluorochromes (which delimit chromosome regions rich in different types of DNA bases) are techniques that have proven to be very useful in improving understanding of the mechanisms involved in the evolution of CH in different genomes (reviewed in Gosálvez et al 1997;Pieczarka et al 1998), In addition, restriction enzyme treatment followed by C-banding is useful in demonstrating C-heterochromatin heterogeneity (Rocco et al 2002;Schmid et al 2002;Chaves et al 2004;Adega et al 2005).…”