Cellular and molecular effects of nonreciprocal chromosome translocations in Saccharomyces cerevisiae

Abstract

Saccharomyces cerevisiae strains harboring a nonreciprocal, bridgeinduced translocation (BIT) between chromosomes VIII and XV exhibited an abnormal phenotype comprising elongated buds and multibudded, unevenly nucleated pseudohyphae. In these cells, we found evidence of molecular effects elicited by the translocation event and specific for its particular genomic location. Expression of genes flanking both translocation breakpoints increased up to five times, correlating with an increased RNA polymerase II binding to their promoters and with their histone acetylation pattern. Microarray data, CHEF, and quantitative PCR confirmed the data on the dosage of genes present on the chromosomal regions involved in the translocation, indicating that telomeric fragments were either duplicated or integrated mostly on chromosome XI. FACS analysis revealed that the majority of translocant cells were blocked in G1 phase and a few of them in G2. Some cells showed a posttranslational decrease of cyclin B1, in agreement with elongated buds diagnostic of a G2/M phase arrest. The actin1 protein was in some cases modified, possibly explaining the abnormal morphology of the cells. Together with the decrease in Rad53p and the lack of its phosphorylation, these results indicate that these cells have undergone adaptation after checkpointmediated G2/M arrest after chromosome translocation. These BIT translocants could serve as model systems to understand further the cellular and molecular effects of chromosome translocation and provide fundamental information on its etiology of neoplastic transformation in mammals.DNA integration ͉ yeast ͉ genome expression ͉ genome dynamics ͉ transcription

Different Aneuploidies Arise From the Same Bridge-Induced Chromosomal Translocation Event in Saccharomyces cerevisiae

“…Figure 6 shows a few examples of the abnormal cell morphologies observed under the microscope, such as germination tube formation, pseudohyphal growth, multi-budded cells, and cells with elongated buds, all indicative of cell cycle defects. It is worth noting that the percentage of all these defects is much lower than that found in other translocants previously described, in which they were between 10 and 20% of the cell population (Nikitin et al 2008). As in previous translocants obtained with the BIT system (Nikitin et al 2008), defects in karyokinesis are also evident in most of the SUSU strains (Figure 6).…”

“…It is worth noting that the percentage of all these defects is much lower than that found in other translocants previously described, in which they were between 10 and 20% of the cell population (Nikitin et al 2008). As in previous translocants obtained with the BIT system (Nikitin et al 2008), defects in karyokinesis are also evident in most of the SUSU strains (Figure 6). …”

“…After 16, 40, or 72 hr growth, 1 ml of the culture was harvested and cells were stained with DAPI as described (Nikitin et al 2008). A Leica DMLB fluorescence microscope was used to monitor DAPI-stained images of yeast cells.…”

“…cDNA synthesis and semiquantitative PCR based on limiting the number of cycles (20-23 cycles) were performed as described (Nikitin et al 2008).…”

“…Recently, our group described a method that allows the generation and subsequent selection of chromo-somal translocations between any two chromosomal loci in diploid yeast strains. This was done by transformation of whole yeast cells with a linear DNA construct, carrying the KAN R selectable marker flanked by DNA sequences homologous to two different chromosomal loci (Tosato et al 2005(Tosato et al , 2009Nikitin et al 2008).…”