Structural and functional evidence that a B chromosome in the characid fish Astyanax scabripinnis is an isochromosome.

Astyanax scabripinnis possesses a widespread polymorphism for metacentric B chromosomes as large as the largest chromosome pair in the A complement. On the basis of C-banding pattern, it was hypothesized that these B chromosomes are isochromosomes that have arisen by means of centromere misdivision and chromatid nondisjunction. In the present paper we test this hypothesis by analysing (i) the localization of a repetitive DNA sequence on both B chromosome arms, and (ii) synaptonemal complex formation, in order to test the functional homology of both arms. Genomic DNA digested with KpnI and analysed by gel electrophoresis showed fragments in a ladder-like pattern typical of tandemly repetitive DNA. These fragments were cloned and their tandem organization in the genome was confirmed. A 51-bp long consensus sequence, which was AT-rich (59%) and contained a variable region and two imperfect reverse sequences, was obtained. Fluorescence in situ hybridization (FISH) localized this repetitive DNA into noncentromeric constitutive heterochromatin which encompasses the terminal region of some acrocentric chromosomes, the NOR region, and interstitial polymorphic heterochromatin in chromosome 24. Most remarkably, tandem repeats were almost symmetrically placed in the two arms of the B chromosome, with the exception of two additional small clusters proximally located on the slightly longer arm. Synaptonemal complex (SC) analysis showed 26 completely paired SCs in males with 1B. The ring configuration of the B univalent persisting until metaphase I suggests that the two arms formed chiasmata. All these data provided strong support for the hypothesis that the B chromosome is an isochromosome.

Accentuated polymorphism of heterochromatin and nucleolar organizer regions in Astyanax scabripinnis (Pisces, Characidae): tools for understanding karyotypic evolution.

Astyanax scabripinnis has been considered a species complex because it presents high karyotypic and morphological variability among its populations. In this work, individuals of two A. scabripinnis populations from different streams in the same hydrographic basin were analyzed through C-banding and AgNOR. Although they present distinct diploid numbers, they show meta and submetacentric chromosome groups highly conserved (numerically and morphologically). Other chromosomal characteristics are also shared by both populations, as the pattern of constitutive heterochromatin distribution (large blocks in the telomeric regions of subtelocentric and acrocentric chromosomes) and some nucleolar chromosomes. Inter-individual variations both in the number and size of heterochromatic blocks, and in the number and localization of NORs were verified in the studied populations, characterizing them as polymorphics for these regions. The mechanisms involved in the dispersion of heterochromatin and NORs through the karyotypes, as well as the possible events related to the generation of polymorphism of those regions are discussed. Furthermore, relationships between these populations and within the context of the scabripinnis complex are also approached.

The X1X2Y sex chromosome system in the fish Hoplias malabaricus. II. Meiotic analyses.

The neotropical fish Hoplias malabaricus shows diversified cytotypes and may represent a group of distinct species. One of these cytotypes is characterized by 2n = 40 and 2n = 39 chromosomes in females and males, respectively, with a multiple sex chromosome system of the X1X1X2X2/X1X2Y type. The Y, representing a large chromosome in male karyotype, is derived from a translocation event between two biarmed chromosomes: one of them similar to X1 chromosome (no. 6) and another one similar to X2 chromosome (probably no. 20). Meiotic data (standard and synaptonemal complexes analyses) show 18 bivalents and one characteristic trivalent in pachytene and metaphase I spermatocytes, as well as two kinds of metaphase II cells with 19 and 20 chromosomes. The trivalent is formed by the Y, X1 and X2 chromosomes and usually presents a complete pairing in pachytene. However, trivalents with partially or fully asynapsed segments are also observed. These segments are assumed to be non-homologous regions of the X1 and X2 chromosomes without correspondence with the Y chromosome, which can heterosynapse. This behaviour of the sex trivalent leading to a fully paired structure, taken together with the close frequencies of the two spermatocyte types at metaphase II, suggests a normal pattern for male H. malabaricus meiosis, representing a stabilized multiple sex chromosome system in this species.

Chromosome banding and synaptonemal complexes in Leporinus lacustris (Pisces, Anostomidae): analysis of a sex system.

Leporinus lacustris had been studied previously, and shows 2n = 54 metacentric and submetacentric chromosomes, including an XX/XY system described on the basis of Giemsa-stained preparations. However, there was some doubt regarding the identification of the sex chromosomes, because of the relative homogeneity of this species karyotype. Thus, the main goal of the present study was to find new evidence of heteromorphic sex chromosomes in these fish through chromosome banding and synaptonemal complex analyses. In fact, the data obtained do not support the presence of sexual heteromorphism. The importance of these methodologies in the study of fish sex chromosomes is discussed.

Post-zygotic modifications and intra- and inter-individual nucleolar organizing region variations in fish: report of a case involving Leporinus friderici.

Silver nitrate staining, a rapid and efficient method, has proven to be excellent for nucleolar organizing region (NOR) studies in fish. Some fish appear to have only two NOR-bearing chromosomes in their karyotype, whereas others probably have several. In the present study we analyzed the NORs of Leporinus friderici, a species that, on the basis of previous studies, has been considered as representative of species with NORs carried by a single chromosome pair. The analyses were performed by a combination of three methods, i.e. silver nitrate staining, staining with the GC-specific fluorochrome chromomycin A3, and in situ hybridization with digoxigenin-labeled probes. The results showed that, although more frequent and conspicuous in a single chromosome pair, the NORs of this species are present in multiple chromosomes. Intra- and inter-individual variations observed by the three methods strongly suggest the occurrence of post-zygotic modifications involving NORs. NOR identification in fish, almost exclusively performed by the silver nitrate method, is currently being re-evaluated by methods such as chromomycin A3 staining and in situ hybridization, which may provide important information leading to a better understanding of chromosome evolution in these animals.

Clonal expansion of cells with trisomy of chromosomes 12 and X in an EBV-transformed lymphoblastoid cell line and establishment of a tumorigenic monoclonal cell line (48,XX,+X,+12).

Clonal expansion of cells with a certain abnormal chromosome constitution has been demonstrated in EBV-transformed lymphoblastoid cell lines, and a new monoclonal tumorigenic cell line (48,XX,+X,+12) has been established. Based on these data and our previous hypothesis that predisposition to tumor development could be associated with inherited susceptibility of certain chromosomes and chromosome regions to breakage, we emphasize that cytogenetic analysis of immortalized cell populations established from normal individuals and cancer patients might be of great importance for predicting development of different primary and secondary malignancies.

Heterochromatin and karyotype reorganization in fish of the family Anostomidae (Characiformes).

Mitotic chromosomes of four fish species of the family Anostomidae, belonging to the genera Leporinus, Leporellus, and Schizodon, were studied. With 2n = 54 meta- and sub-metacentric chromosomes, this family appears to be characterized by marked karyotypic stability. Although perceptible differences exist, mainly in the amount of constitutive heterochromatin present in the chromosomes of these species, these differences do not affect the structure and/or size of these chromosomes. Chromatin substitutions and/or modifications may have led, in one direction, to an increase in heterochromatin in some species and, in the opposite direction, to heterochromatin reduction in others. Whether these changes are accompanied by changes in the amount of euchromatin in the chromosomes is an open question. The nucleolar organizer regions, which may be located on different chromosomes in the various species, may also be indicators of reorganization of these karyotypes.