Cytogenetic Analysis of the Fish Genus Carassius Indicates Divergence, Fission, and Segmental Duplication as Drivers of Tandem Repeat and Microchromosome Evolution.

Fishes of the genus Carassius are useful experimental vertebrate models for the study of evolutionary biology and cytogenetics. Carassius demonstrates diverse biological characteristics, such as variation in ploidy levels and chromosome numbers, and presence of microchromosomes. Those Carassius polyploids with ≥150 chromosomes have microchromosomes, but the origin of microchromosomes, especially in European populations, is unknown. We used cytogenetics to study evolution of tandem repeats (U1 and U2 small nuclear DNAs and H3 histone) and microchromosomes in Carassius from the Czech Republic. We tested the hypotheses whether the number of tandem repeats was affected by polyploidization or divergence between species and what mechanism drives evolution of microchromosomes. Tandem repeats were found in tetraploid and hexaploid Carassius gibelio, and tetraploid Carassius auratus and Carassius carassius in conserved numbers, with the exception of U1 small nuclear DNA in C. auratus. This conservation indicates reduction and/or loss in the number of copies per locus in hexaploids and may have occurred by divergence rather than polyploidization. To study the evolution of microchromosomes, we used the whole microchromosome painting probe from hexaploid C. gibelio and hybridized it to tetraploid and hexaploid C. gibelio, and tetraploid C. auratus and C. carassius. Our results revealed variation in the number of microchromosomes in hexaploids and indicated that the evolution of the Carassius karyotype is governed by macrochromosome fissions followed by segmental duplication in pericentromeric areas. These are potential mechanisms responsible for the presence of microchromosomes in Carassius hexaploids. Differential efficacy of one or both of these mechanisms in different tetraploids could ensure variability in chromosome number in polyploids in general.

Consequences of polyploidy and divergence as revealed by cytogenetic mapping of tandem repeats in African clawed frogs (Xenopus, Pipidae).

Repetitive elements have been identified in several amphibian genomes using whole genome sequencing, but few studies have used cytogenetic mapping to visualize these elements in this vertebrate group. Here, we used fluorescence in situ hybridization and genomic data to map the U1 and U2 small nuclear RNAs and histone H3 in six species of African clawed frog (genus Xenopus), including, from subgenus Silurana, the diploid Xenopus tropicalis and its close allotetraploid relative X. calcaratus and, from subgenus Xenopus, the allotetraploid species X. pygmaeus, X. allofraseri, X. laevis, and X. muelleri. Results allowed us to qualitatively evaluate the relative roles of polyploidization and divergence in the evolution of repetitive elements because our focal species include allotetraploid species derived from two independent polyploidization events - one that is relatively young that gave rise to X. calcaratus and another that is older that gave rise to the other (older) allotetraploids. Our results demonstrated conserved loci number and position of signals in the species from subgenus Silurana; allotetraploid X. calcaratus has twice as many signals as diploid X. tropicalis. However, the content of repeats varied among the other allotetraploid species. We detected almost same number of signals in X. muelleri as in X. calcaratus and same number of signals in X. pygmaeus, X. allofraseri, X. laevis as in the diploid X. tropicalis. Overall, these results are consistent with the proposal that allopolyploidization duplicated these tandem repeats and that variation in their copy number was accumulated over time through reduction and expansion in a subset of the older allopolyploids.

Divergent subgenome evolution in the allotetraploid frog Xenopus calcaratus.

Allopolyploid genomes are divided into compartments called subgenomes that are derived from lower ploidy ancestors. In African clawed frogs of the subgenus Xenopus (genus Xenopus), allotetraploid species have two subgenomes (L and S) with morphologically distinct homoeologous chromosomes. In allotetraploid species of the sister subgenus Silurana, independently evolved subgenomes also exist, but their cytogenetics has not been investigated in detail. We used a diverse suite of cytogenetic and molecular FISH techniques on an allotetraploid species in Silurana-Xenopus calcaratus-to explore evolutionary dynamics of chromosome morphology and rearrangements. We find that the subgenomes of X. calcaratus have distinctive characteristics, with a more conserved a-subgenome resembling the closely related genome of the diploid species X. tropicalis, and a more rapidly evolving b-subgenome having more pronounced changes in chromosome structure, including diverged heterochromatic blocks, repetitive sequences, and deletion of a nucleolar secondary constriction. Based on these cytogenetic differences, we propose a chromosome nomenclature for X. calcaratus that may apply to other allotetraploids in subgenus Silurana, depending on as yet unresolved details of their evolutionary origins. These findings highlight the potential for large-scale asymmetry in subgenome evolution following allopolyploidization.