ABSTRACT
Phenotypic variation occurs through genome rearrangements and mutations in certain responsible genes; however, systematic gene identification methodologies based on genome rearrangements have not been fully established. Here, we explored the loci responsible for the given phenotype using the TAQing system and compared it with a conventional mutagenesis-based method. Two yeast strains with different genetic backgrounds and flocculation phenotypes were fused and genomic rearrangements were induced by transient DNA breaks. Then, selection pressure was applied and multiple mutants were generated, showing different flocculation abilities. We also raised mutants with altered cohesiveness due to spontaneous mutations during long-term recursive passages of haploid strains without TAQing treatment. Comparative genomic analysis of the TAQed mutants revealed three chromosomal regions harboring pivotal flocculation genes, whereas conventional mutagenesis generated a more diverse list of candidate loci after prolonged selection. The combined use of these approaches will accelerate the identification of genes involved in complex phenotypes.
Subject(s)
Gene Rearrangement , Genome , Chromosome Mapping , Comparative Genomic Hybridization , Genomics/methods , Saccharomyces cerevisiae/geneticsABSTRACT
BACKGROUND: Valproic acid (VPA) has been used to treat epilepsy and bipolar disorder. Several reports have demonstrated that VPA functions as a histone deacetylase (HDAC) inhibitor. While VPA is known to cause teratogenic changes in the embryonic zebrafish brain, its effects on neural stem cells (NSCs) in both the embryonic and adult zebrafish are not well understood. RESULTS: In this study, we observed a proliferative effect of VPA on NSCs in the embryonic hindbrain. In contrast, VPA reduced cell proliferation in the adult zebrafish optic tectum. Treatment with HDAC inhibitors showed a similar inhibitory effect on cell proliferation in the adult zebrafish optic tectum, suggesting that VPA reduces cell proliferation through HDAC inhibition. Cell cycle progression was also suppressed in the optic tectum of the adult zebrafish brain because of HDAC inhibition. Recent studies have demonstrated that HDAC inhibits the Notch signaling pathway; hence, adult zebrafish were treated with a Notch inhibitor. This increased the number of proliferating cells in the adult zebrafish optic tectum with down-regulated expression of her4, a target of Notch signaling. CONCLUSIONS: These results suggest that VPA inhibits HDAC activity and upregulates Notch signaling to reduce cell proliferation in the optic tectum of adult zebrafish.
Subject(s)
Cell Proliferation/drug effects , Histone Deacetylase Inhibitors/pharmacology , Neural Stem Cells/drug effects , Superior Colliculi/cytology , Valproic Acid/pharmacology , Zebrafish/physiology , Animals , Basic Helix-Loop-Helix Transcription Factors/metabolism , Blotting, Western , Bromodeoxyuridine , Cell Cycle/drug effects , Cell Cycle/physiology , Cell Proliferation/physiology , DNA Primers/genetics , Gene Expression Regulation/drug effects , Immunohistochemistry , In Situ Hybridization, Fluorescence , In Situ Nick-End Labeling , Real-Time Polymerase Chain Reaction , Receptors, Notch/metabolism , Signal Transduction/drug effects , Superior Colliculi/drug effects , Zebrafish/metabolism , Zebrafish Proteins/metabolismABSTRACT
PR/SET domain containing 9 (Prdm9) mediates histone modifications such as H3K4me3 and marks hotspots of meiotic recombination. In many mammalian species, the Prdm9 gene is highly polymorphic. Prdm9 polymorphism is assumed to play two critical roles in evolution: to diversify the spectrum of meiotic recombination hotspots and to cause male hybrid sterility, leading to reproductive isolation and speciation. Nevertheless, information about Prdm9 sequences in natural populations is very limited. In this study, we conducted a comprehensive population survey on Prdm9 polymorphism in the house mouse, Mus musculus. Overall M. musculus Prdm9 displays an extraordinarily high level of polymorphism, particularly in regions encoding zinc finger repeats, which recognize recombination hotspots. Prdm9 alleles specific to various M. musculus subspecies dominate in subspecies territories. Moreover, introgression into other subspecies territories was found for highly divergent Prdm9 alleles associated with t-haplotype. The results of our phylogeographical analysis suggest that the requirement for hotspot diversity depends on geographical range and time span in mouse evolution, and that Prdm9 polymorphism has not been maintained by a simple balanced selection in the population of each subspecies.
