Quantitative kinetic analysis of nucleolar breakdown and reassembly during mitosis in live human cells

Leung, Anthony K.L.; Gerlich, Daniel W.; Miller, Gail; Lyon, Carol E.; Lam, Yun Wah; Llères, David; Daigle, Nathalie; Zomerdijk, Joost C.B.M.; Ellenberg, Jan; Lamond, Angus I.

doi:10.1083/jcb.200405013

Cited by 144 publications

(135 citation statements)

References 39 publications

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“…2,31 Interestingly, we found that after aclarubicin treatment a small number of early antephase cells proceed to the end of antephase where they arrested for hours with disassembled nucleoli (Mikhailov, Shinohara and Rieder, unpublished observations). This suggests that in these cells, there is sufficient CDK2 activity to drive chromatin condensation and nucleoli dispersion, 50 but not enough cyclin B/CDK1 activity to induce NEB. This could occur, e.g., if the activity of the Cdc25B phosphatase was limited.…”

Section: How Does the P38 Checkpoint Inhibit Progression Through Antementioning

confidence: 99%

The p38-Mediated Stress-Activated Checkpoint: A Rapid Response System for Delaying Progression through Antephase and Entry into Mitosis

Mikhailov¹,

Shinohara²,

Rieder³

2004

Cell Cycle

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Section: How Does the P38 Checkpoint Inhibit Progression Through Antementioning

confidence: 99%

The p38-Mediated Stress-Activated Checkpoint: A Rapid Response System for Delaying Progression through Antephase and Entry into Mitosis

Mikhailov¹,

Shinohara²,

Rieder³

2004

Cell Cycle

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“…In prophase, these nucleoli disassemble and their transcription becomes repressed to allow the stripped rDNA to segregate with the rest of the genome. Nucleolar transcription is downregulated by the CDK1-cyclin B kinase, whose phosphorylation of promoter selectivity factor SL1 prevents the formation of the PolI pre-initiation complex [63][64][65] . Similar to yeast Rio1, RIOK1 might just as well contribute to the transcriptional repression of PolI.…”

Section: Discussionmentioning

confidence: 99%

Rio1 promotes rDNA stability and downregulates RNA polymerase I to ensure rDNA segregation

et al. 2015

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The conserved protein kinase Rio1 localizes to the cytoplasm and nucleus of eukaryotic cells. While the roles of Rio1 in the cytoplasm are well characterized, its nuclear function remains unknown. Here we show that nuclear Rio1 promotes rDNA array stability and segregation in Saccharomyces cerevisiae. During rDNA replication in S phase, Rio1 downregulates RNA polymerase I (PolI) and recruits the histone deacetylase Sir2. Both interventions ensure rDNA copy-number homeostasis and prevent the formation of extrachromosomal rDNA circles, which are linked to accelerated ageing in yeast. During anaphase, Rio1 downregulates PolI by targeting its subunit Rpa43, causing PolI to dissociate from the rDNA. By stimulating the processing of PolI-generated transcripts at the rDNA, Rio1 allows for rDNA condensation and segregation in late anaphase. These events finalize the genome transmission process. We identify Rio1 as an essential nucleolar housekeeper that integrates rDNA replication and segregation with ribosome biogenesis. A t anaphase onset, the replicated chromosomes separate and then segregate along the mitotic spindle into the daughter cells. In the budding yeast Saccharomyces cerevisiae, the locus containing the genes that encode the ribosomal RNAs (rDNA) segregates after the rest of the genome, in late anaphase [1][2][3][4] . The rDNA locus exists as a tandem-repeat array comprising B150 rDNA units containing the 35S and 5S genes, which are transcribed by RNA polymerase I (PolI) and PolIII, respectively. Processing of the 35S pre-rRNA generates 5.8S, 18S and 25S rRNA that, together with the 5S rRNA, become the catalytic backbones of each ribosome 5,6 . Only in anaphase does yeast repress rDNA transcription 4 , which allows the sister rDNA loci to condensate and segregate. PolI downregulation in anaphase is mediated by the Cdc14 phosphatase acting on PolI subunit Rpa43 (ref. 4), resulting in PolI dissociating from the 35S rDNA. The removal of PolI and the local resolution of its transcripts allow the condensin complex to bind. The latter compacts the rDNA array and recruits the DNA decatenating enzyme topoisomerase II (refs 1,3,4,7) resulting in the physical separation and subsequent segregation of the sister rDNA loci.S. cerevisiae Rio1 belongs to the atypical RIO protein kinase family whose members lack the activation loop and substrate recognition domain present in canonical eukaryotic protein kinases [8][9][10][11] . Noteworthy, the RIO kinases may act especially as ATPases as they exhibit o0.1% kinase activity in vitro [12][13][14] . Cytoplasmic Rio1 contributes to pre-40S ribosome biogenesis by promoting 20S pre-rRNA maturation and by stimulating the recycling of trans-acting factors at the pre-40S subunit, both in yeast 12,[15][16][17][18] and human cells 19,20 . Roles in the nucleus are unknown for any RIO member, either in yeast or eukaryotes beyond. Using S. cerevisiae, we now describe the first activities of Rio1 in the nucleus. Foremost, Rio1 downregulates PolI transcription through the cell cycle. In G...

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“…HeLa and HeLa(E-GFP-Ago2) cell lines were generated as described previously (29) and grown with the supplement of 600 g/ml G418 for selection. Dicerϩ/Ϫ, DicerϪ/Ϫ (J.M.C.…”

Section: Methodsmentioning

confidence: 99%

Quantitative analysis of Argonaute protein reveals microRNA-dependent localization to stress granules

Leung

Calabrese

Sharp

2006

Proc. Natl. Acad. Sci. U.S.A.

351

342

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Argonaute proteins associate with microRNAs (miRNAs) that bind mRNAs through partial base-pairings to primarily repress translation in animals. A fraction of Argonaute proteins and miRNAs biochemically cosediment with polyribosomes, yet another fraction paradoxically accumulates in ribosome-free processing bodies (PBs) in the cytoplasm. In this report, we give a quantitative account of the Argonaute protein localization and dynamics in living cells in different cellular states. We find that the majority of Argonaute is distributed diffusely in the cytoplasm, and, when cells are subjected to stress, Argonaute proteins accumulate to newly assembled structures known as stress granules (SGs) in addition to PBs. Argonaute proteins displayed distinct kinetics at different structures: exchange faster at SGs and much slower at PBs. Further, miRNAs are required for the Argonaute protein localization to SGs but not PBs. These quantitative kinetic data provide insights into miRNA-mediated repression.fluorescence microscopy ͉ processing bodies ͉ post-transcriptional control ͉ dynamics

show abstract

Quantitative kinetic analysis of nucleolar breakdown and reassembly during mitosis in live human cells

Cited by 144 publications

References 39 publications

The p38-Mediated Stress-Activated Checkpoint: A Rapid Response System for Delaying Progression through Antephase and Entry into Mitosis

The p38-Mediated Stress-Activated Checkpoint: A Rapid Response System for Delaying Progression through Antephase and Entry into Mitosis

Rio1 promotes rDNA stability and downregulates RNA polymerase I to ensure rDNA segregation

Quantitative analysis of Argonaute protein reveals microRNA-dependent localization to stress granules

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