Live imaging of telomeres: yKu and Sir proteins define redundant telomere-anchoring pathways in yeast.
BACKGROUND
The positioning of chromosomal domains within interphase nuclei is thought to facilitate transcriptional repression in yeast. Although this is particularly well characterized for telomeres, the molecular basis of their specific subnuclear organization is poorly understood. The use of live fluorescence imaging overcomes limitations of in situ staining on fixed cells and permits the analysis of chromatin dynamics in relation to stages of the cell cycle.


RESULTS
We have characterized the dynamics of yeast telomeres and their associated domains of silent chromatin by using rapid time-lapse microscopy. In interphase, native telomeres are highly dynamic but remain within a restricted volume adjacent to the nuclear envelope. This constraint is lost during mitosis. A quantitative analysis of selected mutants shows that the yKu complex is necessary for anchoring some telomeres at the nuclear envelope (NE), whereas the myosin-like proteins Mlp1 and Mlp2 are not. We are able to correlate increased telomeric repression with increased anchoring and show that silent chromatin is tethered to the NE in a Sir-dependent manner in the absence of the yKu complex. Sir-mediated anchoring is S phase specific, while the yKu-mediated pathway functions throughout interphase. Subtelomeric elements of yeast telomere structure influence the relative importance of the yKu- and Sir-dependent mechanisms.


CONCLUSIONS
Interphase positioning of telomeres can be achieved through two partially redundant mechanisms. One requires the heterodimeric yKu complex, but not Mlp1 and Mlp2. The second requires Silent information regulators, correlates with transcriptional repression, and is specific to S phase.
Hediger
Florence
F
University of Geneva, Department of Molecular Biology, Quai Ernest-Ansermet 30, CH-1211 Geneva 4, Switzerland.
Neumann
Frank R
FR
Van Houwe
Griet
G
Dubrana
Karine
K
Gasser
Susan M
SM
D002467:Cell Nucleus / Q000235:genetics; D004256:DNA Polymerase I / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004259:DNA-Directed DNA Polymerase; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006655:Histone Deacetylases / Q000235:genetics / Q000378:metabolism; D007091:Image Processing, Computer-Assisted; D017404:In Situ Hybridization, Fluorescence; D007399:Interphase / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods; D008938:Mitosis / Q000235:genetics; D009154:Mutation; D016196:S Phase / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D056565:Sirtuin 2; D037761:Sirtuins / Q000235:genetics / Q000378:metabolism; D016615:Telomere / Q000502:physiology* / Q000648:ultrastructure; D015003:Yeasts / Q000235:genetics* / Q000378:metabolism
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D008164:Luminescent Proteins; C061573:SIR4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; C081647:high affinity DNA-binding factor, S cerevisiae; D049452:Green Fluorescent Proteins; D004256:DNA Polymerase I; D004259:DNA-Directed DNA Polymerase; C068352:MIP1 protein, S cerevisiae; C087778:SIR2 protein, S cerevisiae; D056565:Sirtuin 2; D037761:Sirtuins; D006655:Histone Deacetylases