Nucleolar structure: Yeast rDNA feels the heat

Condensins are highly conserved protein complexes that regulate chromosome organization, affecting transcription and preparing chromosomes for mitosis. Experiments conducted in budding yeast cells established a foundation for the identification of factors that mediate condensation and for characterizing the activities of condensin complexes. Genetic screens in budding yeast cells first identified SMC (“structural maintenance of chromosomes”) proteins that regulate chromosome stability; initial assignment of these factors to specific activities in chromosome cohesion and condensation relied on the development of DNA FISH methods, coupled to the examination of candidate factors using temperature sensitive alleles. The yeast rDNA locus has proven to be a convenient target for condensation assays, initially assessed using FISH probes to the rDNA, and subsequently by monitoring rDNA condensation state indirectly via observing the fluorescent signal produced by the nucleolar factor Net1 fused to the GFP protein. These experiments defined a series of morphological transitions in the rDNA as chromosomes enter mitosis that likely correspond to increasingly condensed states. Chromosome condensation has also been assessed on non-rDNA sequences by measuring the distance between two points on a chromosome arm in G1 phase cells versus cells blocked in metaphase. This has been determined by two-probe FISH (Guacci et al 1994), or by integrating two lac operator arrays on a chromosome arm and expressing a lacI-GFP fusion protein (D’Ambrosio et al 2008). These assays established that factors shown to be necessary for maintaining condensation of the rDNA also are responsible for condensation of non-rDNA sequences. In this issue of Cell Cycle Shen and Skibbens demonstrate that rDNA structure corresponding to differentially condensed states can be observed directly via DNA staining with DAPI. This simple, refined approach allows researchers to evaluate condensation in any yeast strain, without expression of GFP fusion proteins or use of the technically demanding FISH approach. Using this sensitive method, the authors make the unexpected discovery that rDNA structure is highly sensitive to temperature in yeast cells; yeast allowed to progress into mitosis at the elevated temperature of 37C adopt a novel rDNA morphology that they interpret as a hypercondensed state. Elevated temperature did not lead to increased condensation at a different chromosomal location. The new work identifies unexpected dynamic transitions in the structure of the rDNA and leads to several new questions. It is not clear if the transition to the hypercondensed state is caused by intrinsic responses of the rDNA chromatin architecture to increased temperature, or if it is a regulated response. Phosphorylation of condensin proteins is known to influence late steps in chromosome condensation, thus temperature-regulated alteration in post-translations modifications is a candidate regulatory mechanism. Alternatively, the change in structure could be mediated by a condensin-independent mechanism, possibly mediated by an arm of yeast’s well-defined heat shock response. The rDNA is subject to active suppression of homologous recombination and tight control of transcription, but it’s not yet clear if the newly described state is an adaptive response to high temperature that affects rDNA function. The authors demonstrate that growth rate in their strains is increased at 37C, making it unlikely that the hypercondensed state correlates with reduced rDNA transcription. Identification of mutants that are specifically defective for the transition to the hypercondensed state will likely aid the resolution of these questions.