Axial contraction and short-range compaction of chromatin synergistically promote mitotic chromosome condensation
- Eidgenössische Technische Hochschule Zürich, Switzerland
- Georg- August University Göttingen, Germany
Figures
Figure 1
Fluorescence intensity of TetO/TetR-mCherry as a read-out for chromatin compaction.
(A) Representative images of a cell in G1 and anaphase, containing a TetO array at the TRP1 locus and expressing TetR-mCherry (red). Fluorescence intensity of a focus is measured by determining the …
Figure 2
Fluorophore quenching causes changes of TetO/TetR intensity over the cell cycle.
(A) Two models to explain anaphase-specific decrease in fluorescence brightness (cleansing and quenching, see text for explanations). Shown are the consequences of each model in G1 and anaphase, in …
Figure 3
Dynamics of chromatin compaction and chromosome arm contraction.
(A) Example of a cell going from metaphase to the next G1 phase with TRP1 and LYS4 loci marked with TetR-mCherry and LacI-GFP, respectively. (B) Background normalized, mean GFP-intensity values of …
Figure 4 with 1 supplement
Condensin does not impact chromatin compaction.
(A) TetR-mCherry intensities in the mother cell for the indicated strains and cell cycle stages. To inactivate Smc2, cells were shifted to 37ºC for 90 min. One way ANOVA was performed to test …
Figure 4—figure supplement 1
FACS analysis of alpha-factor synchronized cells released into nocodazole (wild type or smc2-8).
The same samples were analysed by western blot in Figure 4B.
Figure 5
Chromatin compaction does not influence axial chromosome contraction.
(A) TRP1-LYS4 distances for the indicated strains, synchronized in G1 by alpha-factor treatment and released at the indicated temperatures. (B) TRP1-LYS4 distances were determined in the mother cell …
Figure 6 with 1 supplement
Condensin and HST2 are in the same genetic pathway that ensures proper chromosome segregation.
(A) TRP1-LYS4 distances were determined for the indicated strains in anaphase after shifting cells for 90 min to 37ºC. Box shows median value, whiskers all data points n>30. One way ANOVA was …
Figure 6—figure supplement 1
Plate growing spotting assay on YPD plates of wild type, H3 S10A, H3S10D and hst2Δ strains at different temperatures.
YPD, yeast extract peptone dextrose.
Figure 7
Model of chromosome condensation.
H3 S10 phosphorylation leads to chromatin fiber closure, whereas condensin is required for axial shortening. Hst2 is needed for both of these levels of condensation.
Tables
Table 1
Yeast strains used in this study
| yYB number | Mating type | genotype |
|---|---|---|
| N/A | a | his3Δ1 leu2Δ0 met15Δ0 ura3Δ0 smc2::smc2-8:kanMX |
| N/A | a | his3Δ1 leu2Δ0 met15Δ0 ura3Δ0 |
| 3476 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP |
| 3477 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP ipl1-321 |
| 4699 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP Spc42-GFP:hph hht1::hht1-S10A:KanMXloxP hht2::hht2-S10A:bleloxP ura3 ade1 leu2 |
| 8080 | a | Nup170-GFP:HIS3 ura3 ade1 leu2 |
| 9101 | a | his3Δ200 leu2Δ0 lys2Δ0 trp1Δ63 ura3Δ0 met15Δ0 can1::MFA1pr-HIS3 hht1-hhf1::NatMX4 hht2-hhf2::[HHTS-HHFS]-URA3 |
| 9332 | a | his3Δ200 leu2Δ0 lys2Δ0 trp1Δ63 ura3Δ0 met15Δ0 can1::MFA1pr-HIS3 hht1-hhf1::NatMX4 hht2-hhf2::[HHT-S10D-HHFS]-URA3 |
| 10818 | a | his3Δ200 leu2Δ0 lys2Δ0 trp1Δ63 ura3Δ0 met15Δ0 can1::MFA1pr-HIS3 hht1-hhf1::NatMX4 hht2-hhf2::[HHTS10AS]-URA3 |
| 11676 | a | smc2-8 hhf1-hht1::NatMX hst2::hphNT1 ura3 ade1 leu2 |
| 12002 | a | hst2::KanMX his3∆1 leu2∆0 ura3∆0 met15∆0 |
| 10116, 10117 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP Hhf1-Hht1::NatMX hht2-hhf2::[HHT-HHF∆9-16]-URA3 ura3 ade1 leu2 |
| 10122, 10123 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP Hhf1-Hht1::NatMX hht2-hhf2::[HHT-HHFK16R]-URA3 ura3 ade1 leu2 |
| 10331 | a | Nup170-GFP:HIS3 hst2::NatMX ura3 ade1 leu2 |
| 11518, 11519 | diploid | trp1::TetO:TRP1/TRP1 lys4::LacO:LEU2/LYS4 TetR-mRFP leu2/leu2 ura3/ura3 his3/his3 |
| 11520, 11521 | diploid | trp1::TetO:TRP1/TRP1 TetR-mRFP/ TetR-GFP:LEU2 leu2/leu2 ura3/ura3 his3/his3 |
| 11173, 11989 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP hht1::hht1-S10A:KanMXloxP hht2::hht2-S10A:bleloxP smc2-8 ura3 ade leu2 |
| 9782, 9783, 9784 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP Hhf1-Hht1::NatMX ura3 ade1 leu2 |
| 10006, 10007, 10008 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP Hhf1-Hht1::NatMX hst2::hphNT1 ura3 ade1 leu2 |
| 10274, 10276, 10277 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP smc2-8 |
| 10450, 10451, 10452 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP HHT2S10D:URA3 ura3 ade1 leu2 |
| 10584, 10585, 10586 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP HHT2S10D:URA3 smc2-8 |
| 10858, 10859, 10860 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP HHT2S10D:URA3 hst2::NatMX ura3 ade1 leu2 |
| 10861, 10862, 10863 | a | trp1::TetO:TRP1lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP HHT2S10D:URA3 smc2-8 hst2::NatMX |
| 11566, 11567, 11568 | alpha | his3Δ200 leu2Δ0 lys2Δ0 trp1Δ63 ura3Δ0 met15Δ0 can1::MFA1pr-HIS3 hht1-hhf1::NatMX4 hht2-hhf2::[HHTS10AS]-URA3 smc2-8 |
| 11767, 11768, 11769 | a, alpha | his3Δ200 leu2Δ0 lys2Δ0 trp1Δ63 ura3Δ0 met15Δ0 can1::MFA1pr-HIS3 hht1-hhf1::NatMX4 hht2-hhf2::[HHTS-HHFS]-URA3 smc2-8 |
| 12155, 12156, 12157 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP hst2::KanMX |
| 12731, 12732, 12733 | a | his3Δ200 leu2Δ0 lys2Δ0 trp1Δ63 ura3Δ0 met15Δ0 can1::MFA1pr-HIS3 hht1-hhf1::NatMX4 hht2-hhf2::[HHT-S10D-HHFS]-URA3 smc2-8 |
| 13100,13102 | a | trp1::TetO:TRP1 lys4::LacO:LEU2 his3::LacR-GFP:HIS3 TetR-mRFP Hhf1-Hht1::NatMX hht2-hhf2::[HHT-HHFK16R]-URA3 hst2::hphNT1 ura3 ade1 leu2 |
