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A mechanism for asymmetric segregation of age during yeast budding

Abstract

Ageing and the mortality that ensues are sustainable for the species only if age is reset in newborns. In budding yeast, buds are made young whereas ageing factors, such as carbonylated proteins and DNA circles, remain confined to the ageing mother cell. The mechanisms of this confinement and their relevance are poorly understood. Here we show that a septin-dependent, lateral diffusion barrier forms in the nuclear envelope and limits the translocation of pre-existing nuclear pores into the bud. The retention of DNA circles within the mother cell depends on the presence of the diffusion barrier and on the anchorage of the circles to pores mediated by the nuclear basket. In accordance with the diffusion barrier ensuring the asymmetric segregation of nuclear age-determinants, the barrier mutant bud6Δ fails to properly reset age in buds. Our data involve septin-dependent diffusion barriers in the confinement of ageing factors to one daughter cell during asymmetric cell division.

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Figure 1: Restriction of lateral diffusion through the bud neck in the outer membrane of yeast anaphase nuclei.
Figure 2: A diffusion barrier in the nuclear envelope retains pre-existing nuclear pores in the mother cell.
Figure 3: Nuclear diffusion barrier is septin dependent and Bud6 dependent.
Figure 4: Subnuclear distribution of the non-centromeric plasmid pPCM14.
Figure 5: Bud neck defects and plasmid segregation.
Figure 6: Effect of diffusion barrier defects on age asymmetry.

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Acknowledgements

We thank D. Barral and J. Sasse, P. Megee, D. Koschland, T. Lithgow, L. Guarente, V. Doye, E. Hurt and M. Winey for technical help and sharing reagents, D. Gerlich and C. Weirich for reading the manuscript, and the Barral, Meraldi and Gerlich labs for discussions. We acknowledge G. Csucs, J. Kusch and the Light Microscopy Center (LMC, ETH Zurich) for support with microscopy equipment and techniques. This work was supported by the ETH Zurich, and by a grant from the Swiss National Foundation to Y.B. and G.G.

Author Contributions Z.S. did most of the experiments and contributed to the data analysis and to the writing of the paper; S.B. contributed to the ageing experiments; S.B.F. contributed to the FLIP experiments; G.G. did the in silico modelling; and Y.B. had the idea for the project, contributed to the data analysis and wrote the paper.

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Correspondence to Yves Barral.

Supplementary information

Supplementary Information 1

The file contains Figures S1-S7 and Legends; Supplementary Methods; Supplementary Movie legends and Supplementary Notes with additional references. (PDF 818 kb)

nature07121-s2.mov

The file contains Supplementary Movie 1 which shows a representative FLIP experiment on a Nup49-GFP cell. Note that none of the movies was treated to account for the photobleaching due to picture acquisition. Please always keep an eye on the control cell. (MOV 963 kb)

nature07121-s3.mov

The file contains Supplementary Movie 2 which shows FLIP experiment on a Nsg1-GFP cell. (MOV 1032 kb)

nature07121-s4.mov

The file contains Supplementary Movie 3 which shows FLIP experiment on a Sec61-GFP cell. (MOV 1346 kb)

nature07121-s5.mov

The file contains Supplementary Movie 4 which shows FLIP experiment on a NLS-GFP cell. (MOV 210 kb)

nature07121-s6.mov

The file contains Supplementary Movie 5 which shows FLIP experiment on a Prm3-GFP cell. (MOV 433 kb)

nature07121-s7.mov

The file contains Supplementary Movie 6 which shows FLIP experiment on a GFP-HDEL cell. (MOV 869 kb)

nature07121-s8.mov

The file contains Supplementary Movie 7 which shows FRAP experiment on a Nup49-GFP cell. (MOV 1467 kb)

nature07121-s9.mov

The file contains Supplementary Movie 8 which shows FLIP experiment on a cdc12-6 Nsg1-GFP cell. (MOV 1280 kb)

nature07121-s10.mov

The file contains Supplementary Movie 9 which shows FLIP experiment on a bud6Δ Nsg1-GFP cell. (MOV 940 kb)

nature07121-s11.mov

The file contains Supplementary Movie 10 which shows CEN- pPCM14 plasmids codiffusing with nuclear pore clusters in nup133Δ cells. (MOV 57 kb)

nature07121-s12.mov

The file contains Supplementary Movie 11 which shows movement of CEN- pPCM14 plasmids in a kar9Δ cell. Note that the plasmids can move freely between the two lobes of anaphase nucleus until the mitotic spindle realigns with the mother-bud axis and passes through the bud neck. (MOV 105 kb)

nature07121-s13.mov

The file contains Supplementary Movie 12 which shows movement of CEN- pPCM14 plasmids in a wild type cell. None of the two plasmids crosses the bud neck. (MOV 31 kb)

nature07121-s14.mov

The file contains Supplementary Movie 13 which shows movement of CEN- pPCM14 plasmids in a cdc12-6 cell. In this cell, 2 out of 4 plasmids segregate towards the bud. (MOV 70 kb)

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Shcheprova, Z., Baldi, S., Frei, S. et al. A mechanism for asymmetric segregation of age during yeast budding. Nature 454, 728–734 (2008). https://doi.org/10.1038/nature07212

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