A genetically-encoded crosslinker screen identifies SERBP1 as a PKCε substrate influencing translation and cell division

AbstractThe PKCε-regulated genome protective pathway provides transformed cells a failsafe to successfully complete mitosis. Despite the necessary role for Aurora B in this programme, it is unclear whether its requirement is sufficient or if other PKCε cell cycle targets are involved. To address this, we developed a trapping strategy using UV-photocrosslinkable amino acids encoded in the PKCε kinase domain. The validation of the mRNA binding protein SERBP1 as a PKCε substrate revealed a series of mitotic events controlled by the catalytic form of PKCε. PKCε represses protein translation, altering SERBP1 binding to the 40 S ribosomal subunit and promoting the assembly of ribonucleoprotein granules containing SERBP1, termed M-bodies. Independent of Aurora B, SERBP1 is shown to be necessary for chromosome segregation and successful cell division, correlating with M-body formation. This requirement for SERBP1 demonstrates that Aurora B acts in concert with translational regulation in the PKCε-controlled pathway exerting genome protection.

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Borealin–nucleosome interaction secures chromosome association of the chromosomal passenger complex

The Journal of Cell Biology ◽

10.1083/jcb.201905040 ◽

2019 ◽

Vol 218 (12) ◽

pp. 3912-3925 ◽

Cited By ~ 7

Author(s):

Maria A. Abad ◽

Jan G. Ruppert ◽

Lana Buzuk ◽

Martin Wear ◽

Juan Zou ◽

...

Keyword(s):

Cell Division ◽

Chromosome Segregation ◽

Chromosome Association ◽

Aurora B ◽

Master Regulator ◽

Multifaceted Approach ◽

Chromosomal Passenger Complex ◽

Chromosome Congression ◽

Chromosomal Passenger ◽

And Function

Chromosome association of the chromosomal passenger complex (CPC; consisting of Borealin, Survivin, INCENP, and the Aurora B kinase) is essential to achieve error-free chromosome segregation during cell division. Hence, understanding the mechanisms driving the chromosome association of the CPC is of paramount importance. Here using a multifaceted approach, we show that the CPC binds nucleosomes through a multivalent interaction predominantly involving Borealin. Strikingly, Survivin, previously suggested to target the CPC to centromeres, failed to bind nucleosomes on its own and requires Borealin and INCENP for its binding. Disrupting Borealin–nucleosome interactions excluded the CPC from chromosomes and caused chromosome congression defects. We also show that Borealin-mediated chromosome association of the CPC is critical for Haspin- and Bub1-mediated centromere enrichment of the CPC and works upstream of the latter. Our work thus establishes Borealin as a master regulator determining the chromosome association and function of the CPC.

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Translation inhibition from a distance: the small RNA SgrS interferes with a ribosomal protein S1-dependent enhancer

10.1101/808485 ◽

2019 ◽

Author(s):

Muhammad S. Azam ◽

Carin K. Vanderpool

Keyword(s):

Ribosomal Protein ◽

Translational Regulation ◽

Ribosomal Subunit ◽

Start Codon ◽

Binding Interaction ◽

Translation Inhibition ◽

Ribosomal Protein S1 ◽

Chaperone Protein ◽

Ribosomal Subunit Protein ◽

Mrna Binding

SummaryMany bacterial small RNAs (sRNAs) efficiently inhibit translation of target mRNAs by forming a duplex that sequesters the Shine-Dalgarno (SD) sequence or start codon and prevents formation of the translation initiation complex. There are a growing number of examples of sRNA-mRNA binding interactions distant from the SD region, but how these mediate translational regulation remains unclear. Our previous work in Escherichia coli and Salmonella identified a mechanism of translational repression of manY mRNA by the sRNA SgrS through a binding interaction upstream of the manY SD. Here, we report that SgrS forms a duplex with a uridine-rich translation-enhancing element in the manY 5’ untranslated region. Notably, we show that the enhancer is ribosome-dependent and that the small ribosomal subunit protein S1 interacts with the enhancer to promote translation of manY. In collaboration with the chaperone protein Hfq, SgrS interferes with the interaction between the translation enhancer and ribosomal protein S1 to repress translation of manY mRNA. Since bacterial translation is often modulated by enhancer-like elements upstream of the SD, sRNA-mediated enhancer silencing could be a common mode of gene regulation.

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Prophase removal of chromosome-associated RNAs facilitates anaphase chromosome segregation

10.1101/813527 ◽

2019 ◽

Author(s):

Judith A. Sharp ◽

Wei Wang ◽

Michael D. Blower

Keyword(s):

Cell Division ◽

Chromosome Segregation ◽

Mitotic Chromosome ◽

Nuclear Protein ◽

Aurora B ◽

Dna Binding Domain ◽

Nucleic Acid Binding ◽

Mitotic Chromosomes ◽

Nuclear Envelope Breakdown ◽

Binding Domains

AbstractDuring mitosis, the genome is transformed from a decondensed, transcriptionally active state to a highly condensed, transcriptionally inactive state. Mitotic chromosome reorganization is marked by the general attenuation of transcription on chromosome arms, yet how the cell regulates nuclear and chromatin-associated RNAs after chromosome condensation and nuclear envelope breakdown is unknown. SAF-A/hnRNPU is an abundant nuclear protein with RNA-to-DNA tethering activity, coordinated by two spatially distinct nucleic acid binding domains. Here we show that RNA is evicted from prophase chromosomes through Aurora-B-dependent phosphorylation of the SAF-A DNA-binding domain; failure to execute this pathway leads to accumulation of SAF-A:RNA complexes on mitotic chromosomes and elevated rates of anaphase segregation defects. This work reveals a role for Aurora-B in removing chromatin-associated RNAs during prophase, and demonstrates that Aurora-B dependent relocalization of SAF-A during cell division contributes to the fidelity of chromosome segregation.

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Direct Nucleosome Binding of Borealin Secures Chromosome Association and Function of the Chromosomal Passenger complex

10.1101/620385 ◽

2019 ◽

Cited By ~ 1

Author(s):

M. A. Abad ◽

J. G. Ruppert ◽

L. Buzuk ◽

M. Wear ◽

J. Zou ◽

...

Keyword(s):

Cell Division ◽

Chromosome Segregation ◽

Chromosome Association ◽

Aurora B ◽

Multifaceted Approach ◽

Chromosomal Passenger Complex ◽

Chromosome Congression ◽

Chromosomal Passenger ◽

And Function ◽

Nucleosome Binding

SummaryChromosome association of the Chromosomal Passenger Complex (CPC; consisting of Borealin, Survivin, INCENP and the Aurora B kinase) is essential to achieve error-free chromosome segregation during cell division. Hence, understanding the mechanisms driving the chromosome association of the CPC is of paramount importance. Here using a multifaceted approach, we show that the CPC binds nucleosomes through a multivalent interaction predominantly involving Borealin. Strikingly, Survivin, previously suggested to target the CPC to centromeres [1–3] failed to bind nucleosomes on its own and requires Borealin and INCENP for its binding. Disrupting Borealin-nucleosome interactions excluded the CPC from chromosomes and caused chromosome congression defects. We also show that Borealin-mediated chromosome association of the CPC is critical for Haspin- and Bub1-mediated centromere enrichment of the CPC and works upstream of the latter. Our work thus establishes Borealin as a master regulator determining the chromosome association and function of the CPC.

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Nuclear Assembly of UGA Decoding Complexes on Selenoprotein mRNAs: a Mechanism for Eluding Nonsense-Mediated Decay?

Molecular and Cellular Biology ◽

10.1128/mcb.26.5.1795-1805.2006 ◽

2006 ◽

Vol 26 (5) ◽

pp. 1795-1805 ◽

Cited By ~ 39

Author(s):

Lucia A. de Jesus ◽

Peter R. Hoffmann ◽

Tanya Michaud ◽

Erin P. Forry ◽

Andrea Small-Howard ◽

...

Keyword(s):

Mammalian Cells ◽

Stop Codon ◽

Elongation Factor ◽

Protein Translation ◽

Nonsense Mediated Decay ◽

Nuclear Assembly ◽

Mrna Binding Protein ◽

Two Factors ◽

Mrna Binding ◽

Selenoprotein Mrnas

ABSTRACT Recoding of UGA from a stop codon to selenocysteine poses a dilemma for the protein translation machinery. In eukaryotes, two factors that are crucial to this recoding process are the mRNA binding protein of the Sec insertion sequence, SBP2, and the specialized elongation factor, EFsec. We sought to determine the subcellular localization of these selenoprotein synthesis factors in mammalian cells and thus gain insight into how selenoprotein mRNAs might circumvent nonsense-mediated decay. Intriguingly, both EFsec and SBP2 localization differed depending on the cell line but significant colocalization of the two proteins was observed in cells where SBP2 levels were detectable. We identify functional nuclear localization and export signals in both proteins, demonstrate that SBP2 undergoes nucleocytoplasmic shuttling, and provide evidence that SBP2 levels and localization may influence EFsec localization. Our results suggest a mechanism for the nuclear assembly of the selenocysteine incorporation machinery that could allow selenoprotein mRNAs to circumvent nonsense-mediated decay, thus providing new insights into the mechanism of selenoprotein translation.

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Cell division requires RNA eviction from condensing chromosomes

The Journal of Cell Biology ◽

10.1083/jcb.201910148 ◽

2020 ◽

Vol 219 (11) ◽

Cited By ~ 1

Author(s):

Judith A. Sharp ◽

Carlos Perea-Resa ◽

Wei Wang ◽

Michael D. Blower

Keyword(s):

Cell Division ◽

Chromosome Segregation ◽

Mitotic Chromosome ◽

Aurora B ◽

Nucleic Acid Binding ◽

Mitotic Chromosomes ◽

Nuclear Envelope Breakdown ◽

Binding Domains ◽

Chromosome Alignment ◽

Rna Complexes

During mitosis, the genome is transformed from a decondensed, transcriptionally active state to a highly condensed, transcriptionally inactive state. Mitotic chromosome reorganization is marked by the general attenuation of transcription on chromosome arms, yet how the cell regulates nuclear and chromatin-associated RNAs after chromosome condensation and nuclear envelope breakdown is unknown. SAF-A/hnRNPU is an abundant nuclear protein with RNA-to-DNA tethering activity, coordinated by two spatially distinct nucleic acid–binding domains. Here we show that RNA is evicted from prophase chromosomes through Aurora-B–dependent phosphorylation of the SAF-A DNA-binding domain; failure to execute this pathway leads to accumulation of SAF-A–RNA complexes on mitotic chromosomes, defects in metaphase chromosome alignment, and elevated rates of chromosome missegregation in anaphase. This work reveals a role for Aurora-B in removing chromatin-associated RNAs during prophase and demonstrates that Aurora-B–dependent relocalization of SAF-A during cell division contributes to the fidelity of chromosome segregation.

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Tension promotes kinetochore-microtubule release in response to Aurora B activity

10.1101/2020.06.01.127795 ◽

2020 ◽

Author(s):

Geng-Yuan Chen ◽

Fioranna Renda ◽

Huaiying Zhang ◽

Alper Gokden ◽

Daniel Z. Wu ◽

...

Keyword(s):

Cell Division ◽

Error Correction ◽

Chromosome Segregation ◽

Kinase Activity ◽

Aurora B ◽

Aurora B Kinase ◽

Spindle Poles ◽

Low Tension

AbstractAurora B kinase regulates kinetochore-microtubule interactions to ensure accurate chromosome segregation in cell division. Tension provides a signal to discriminate attachment errors from bi-oriented kinetochores with sisters correctly attached to opposite spindle poles. Current models focus on tension as an input to locally regulate Aurora B activity. Here we show that the outcome of Aurora B activity depends on tension. Using an optogenetic approach to manipulate Aurora B at individual kinetochores, we find that kinase activity promotes microtubule release when tension is high. Conversely, when tension is low, Aurora B activity promotes depolymerization of kinetochore-microtubule bundles while maintaining attachment. We propose that tension is a signal inducing distinct error-correction mechanisms, with release or depolymerization advantageous for typical errors characterized by high or low tension, respectively.

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