Centromeric Localization of Dispersed Pol III Genes in Fission Yeast

The eukaryotic genome is a complex three-dimensional entity residing in the nucleus. We present evidence that Pol III–transcribed genes such as tRNA and 5S rRNA genes can localize to centromeres and contribute to a global genome organization. Furthermore, we find that ectopic insertion of Pol III genes into a non-Pol III gene locus results in the centromeric localization of the locus. We show that the centromeric localization of Pol III genes is mediated by condensin, which interacts with the Pol III transcription machinery, and that transcription levels of the Pol III genes are negatively correlated with the centromeric localization of Pol III genes. This centromeric localization of Pol III genes initially observed in interphase becomes prominent during mitosis, when chromosomes are condensed. Remarkably, defective mitotic chromosome condensation by a condensin mutation, cut3-477, which reduces the centromeric localization of Pol III genes, is suppressed by a mutation in the sfc3 gene encoding the Pol III transcription factor TFIIIC subunit, sfc3-1. The sfc3-1 mutation promotes the centromeric localization of Pol III genes. Our study suggests there are functional links between the process of the centromeric localization of dispersed Pol III genes, their transcription, and the assembly of condensed mitotic chromosomes.

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Long Tandem Arrays of Cassandra Retroelements and Their Role in Genome Dynamics in Plants

International Journal of Molecular Sciences ◽

10.3390/ijms21082931 ◽

2020 ◽

Vol 21 (8) ◽

pp. 2931 ◽

Cited By ~ 5

Author(s):

Ruslan Kalendar ◽

Olga Raskina ◽

Alexander Belyayev ◽

Alan H. Schulman

Keyword(s):

Copy Number ◽

5S Rdna ◽

5S Rrna ◽

Rrna Genes ◽

Gene Copy ◽

Rrna Gene ◽

Long Terminal Repeats ◽

5S Rrna Genes ◽

Pol Iii ◽

Tandem Arrays

Retrotransposable elements are widely distributed and diverse in eukaryotes. Their copy number increases through reverse-transcription-mediated propagation, while they can be lost through recombinational processes, generating genomic rearrangements. We previously identified extensive structurally uniform retrotransposon groups in which no member contains the gag, pol, or env internal domains. Because of the lack of protein-coding capacity, these groups are non-autonomous in replication, even if transcriptionally active. The Cassandra element belongs to the non-autonomous group called terminal-repeat retrotransposons in miniature (TRIM). It carries 5S RNA sequences with conserved RNA polymerase (pol) III promoters and terminators in its long terminal repeats (LTRs). Here, we identified multiple extended tandem arrays of Cassandra retrotransposons within different plant species, including ferns. At least 12 copies of repeated LTRs (as the tandem unit) and internal domain (as a spacer), giving a pattern that resembles the cellular 5S rRNA genes, were identified. A cytogenetic analysis revealed the specific chromosomal pattern of the Cassandra retrotransposon with prominent clustering at and around 5S rDNA loci. The secondary structure of the Cassandra retroelement RNA is predicted to form super-loops, in which the two LTRs are complementary to each other and can initiate local recombination, leading to the tandem arrays of Cassandra elements. The array structures are conserved for Cassandra retroelements of different species. We speculate that recombination events similar to those of 5S rRNA genes may explain the wide variation in Cassandra copy number. Likewise, the organization of 5S rRNA gene sequences is very variable in flowering plants; part of what is taken for 5S gene copy variation may be variation in Cassandra number. The role of the Cassandra 5S sequences remains to be established.

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pEg7, a New Xenopus Protein Required for Mitotic Chromosome Condensation in Egg Extracts

The Journal of Cell Biology ◽

10.1083/jcb.143.6.1437 ◽

1998 ◽

Vol 143 (6) ◽

pp. 1437-1446 ◽

Cited By ~ 28

Author(s):

Fabien Cubizolles ◽

Vincent Legagneux ◽

René Le Guellec ◽

Isabelle Chartrain ◽

Rustem Uzbekov ◽

...

Keyword(s):

Mitotic Chromosome ◽

Cultured Cells ◽

Sperm Chromatin ◽

Mitotic Chromosomes ◽

Maternal Mrna ◽

Early Embryos ◽

Egg Extracts ◽

Cytostatic Factor ◽

Cell Expression ◽

Mitotic Chromosome Condensation

We have isolated a cDNA, Eg7, corresponding to a Xenopus maternal mRNA, which is polyadenylated in mature oocytes and deadenylated in early embryos. This maternal mRNA encodes a protein, pEg7, whose expression is strongly increased during oocyte maturation. The tissue and cell expression pattern of pEg7 indicates that this protein is only readily detected in cultured cells and germ cells. Immunolocalization in Xenopus cultured cells indicates that pEg7 concentrates onto chromosomes during mitosis. A similar localization of pEg7 is observed when sperm chromatin is allowed to form mitotic chromosomes in cytostatic factor-arrested egg extracts. Incubating these extracts with antibodies directed against two distinct parts of pEg7 provokes a strong inhibition of the condensation and resolution of mitotic chromosomes. Biochemical experiments show that pEg7 associates with Xenopus chromosome-associated polypeptides C and E, two components of the 13S condensin.

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Topoisomerase IIα is essential for maintenance of mitotic chromosome structure

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2001760117 ◽

2020 ◽

Vol 117 (22) ◽

pp. 12131-12142 ◽

Cited By ~ 2

Author(s):

Christian F. Nielsen ◽

Tao Zhang ◽

Marin Barisic ◽

Paul Kalitsis ◽

Damien F. Hudson

Keyword(s):

Cell Cycle ◽

Chromosome Segregation ◽

Mitotic Chromosome ◽

Chromosome Structure ◽

Chromatin Condensation ◽

Chromosome Condensation ◽

Mitotic Chromosomes ◽

Topoisomerase Iiα ◽

Mitotic Chromosome Condensation

Topoisomerase IIα (TOP2A) is a core component of mitotic chromosomes and important for establishing mitotic chromosome condensation. The primary roles of TOP2A in mitosis have been difficult to decipher due to its multiple functions across the cell cycle. To more precisely understand the role of TOP2A in mitosis, we used the auxin-inducible degron (AID) system to rapidly degrade the protein at different stages of the human cell cycle. Removal of TOP2A prior to mitosis does not affect prophase timing or the initiation of chromosome condensation. Instead, it prevents chromatin condensation in prometaphase, extends the length of prometaphase, and ultimately causes cells to exit mitosis without chromosome segregation occurring. Surprisingly, we find that removal of TOP2A from cells arrested in prometaphase or metaphase cause dramatic loss of compacted mitotic chromosome structure and conclude that TOP2A is crucial for maintenance of mitotic chromosomes. Treatments with drugs used to poison/inhibit TOP2A function, such as etoposide and ICRF-193, do not phenocopy the effects on chromosome structure of TOP2A degradation by AID. Our data point to a role for TOP2A as a structural chromosome maintenance enzyme locking in condensation states once sufficient compaction is achieved.

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A Human Condensin Complex Containing hCAP-C–hCAP-E and CNAP1, a Homolog of Xenopus XCAP-D2, Colocalizes with Phosphorylated Histone H3 during the Early Stage of Mitotic Chromosome Condensation

Molecular and Cellular Biology ◽

10.1128/mcb.20.18.6996-7006.2000 ◽

2000 ◽

Vol 20 (18) ◽

pp. 6996-7006 ◽

Cited By ~ 70

Author(s):

John A. Schmiesing ◽

Heather C. Gregson ◽

Sharleen Zhou ◽

Kyoko Yokomori

Keyword(s):

Mammalian Cells ◽

Structural Changes ◽

Mitotic Chromosome ◽

Early Stage ◽

Histone H3 ◽

Chromosome Condensation ◽

Early Prophase ◽

Mitotic Chromosomes ◽

Condensin Complex ◽

Mitotic Chromosome Condensation

ABSTRACT Structural maintenance of chromosomes (SMC) family proteins play critical roles in structural changes of chromosomes. Previously, we identified two human SMC family proteins, hCAP-C and hCAP-E, which form a heterodimeric complex (hCAP-C–hCAP-E) in the cell. Based on the sequence conservation and mitotic chromosome localization, hCAP-C–hCAP-E was determined to be the human ortholog of theXenopus SMC complex, XCAP-C–XCAP-E. XCAP-C–XCAP-E is a component of the multiprotein complex termed condensin, required for mitotic chromosome condensation in vitro. However, presence of such a complex has not been demonstrated in mammalian cells. Coimmunoprecipitation of the endogenous hCAP-C–hCAP-E complex from HeLa extracts identified a 155-kDa protein interacting with hCAP-C–hCAP-E, termed condensation-related SMC-associated protein 1 (CNAP1). CNAP1 associates with mitotic chromosomes and is homologous toXenopus condensin component XCAP-D2, indicating the presence of a condensin complex in human cells. Chromosome association of human condensin is mitosis specific, and the majority of condensin dissociates from chromosomes and is sequestered in the cytoplasm throughout interphase. However, a subpopulation of the complex was found to remain on chromosomes as foci in the interphase nucleus. During late G2/early prophase, the larger nuclear condensin foci colocalize with phosphorylated histone H3 clusters on partially condensed regions of chromosomes. These results suggest that mitosis-specific function of human condensin may be regulated by cell cycle-specific subcellular localization of the complex, and the nuclear condensin that associates with interphase chromosomes is involved in the reinitiation of mitotic chromosome condensation in conjunction with phosphorylation of histone H3.

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Identification of a Chromosome-Targeting Domain in the Human Condensin Subunit CNAP1/hCAP-D2/Eg7

Molecular and Cellular Biology ◽

10.1128/mcb.22.16.5769-5781.2002 ◽

2002 ◽

Vol 22 (16) ◽

pp. 5769-5781 ◽

Cited By ~ 33

Author(s):

Alexander R. Ball, ◽

John A. Schmiesing ◽

Changcheng Zhou ◽

Heather C. Gregson ◽

Yoshiaki Okada ◽

...

Keyword(s):

Mitotic Chromosome ◽

Mitotic Chromosomes ◽

Family Protein ◽

Localization Signal ◽

The Individual ◽

Mitotic Chromosome Condensation ◽

Structural Maintenance Of Chromosomes

ABSTRACT CNAP1 (hCAP-D2/Eg7) is an essential component of the human condensin complex required for mitotic chromosome condensation. This conserved complex contains a structural maintenance of chromosomes (SMC) family protein heterodimer and three non-SMC subunits. The mechanism underlying condensin targeting to mitotic chromosomes and the role played by the individual condensin components, particularly the non-SMC subunits, are not well understood. We report here characterization of the non-SMC condensin component CNAP1. CNAP1 contains two separate domains required for its stable incorporation into the complex. We found that the carboxyl terminus of CNAP1 possesses a mitotic chromosome-targeting domain that does not require the other condensin components. The same region also contains a functional bipartite nuclear localization signal. A mutant CNAP1 missing this domain, although still incorporated into condensin, was unable to associate with mitotic chromosomes. Successful chromosome targeting of deletion mutants correlated with their ability to directly bind to histones H1 and H3 in vitro. The H3 interaction appears to be mediated through the H3 histone tail, and a subfragment containing the targeting domain was found to interact with histone H3 in vivo. Thus, the CNAP1 C-terminal region defines a novel histone-binding domain that is responsible for targeting CNAP1, and possibly condensin, to mitotic chromosomes.

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Axial contraction and short-range compaction of chromatin synergistically promote mitotic chromosome condensation

eLife ◽

10.7554/elife.10396 ◽

2015 ◽

Vol 4 ◽

Cited By ~ 13

Author(s):

Tom Kruitwagen ◽

Annina Denoth-Lippuner ◽

Bryan J Wilkins ◽

Heinz Neumann ◽

Yves Barral

Keyword(s):

Short Range ◽

Mitotic Chromosome ◽

Chromosome Condensation ◽

Yeast Cells ◽

Histone H4 ◽

Mitotic Chromosomes ◽

Chromatin Compaction ◽

Early Anaphase ◽

Local Compaction ◽

Mitotic Chromosome Condensation

The segregation of eukaryotic chromosomes during mitosis requires their extensive folding into units of manageable size for the mitotic spindle. Here, we report on how phosphorylation at serine 10 of histone H3 (H3 S10) contributes to this process. Using a fluorescence-based assay to study local compaction of the chromatin fiber in living yeast cells, we show that chromosome condensation entails two temporally and mechanistically distinct processes. Initially, nucleosome-nucleosome interaction triggered by H3 S10 phosphorylation and deacetylation of histone H4 promote short-range compaction of chromatin during early anaphase. Independently, condensin mediates the axial contraction of chromosome arms, a process peaking later in anaphase. Whereas defects in chromatin compaction have no observable effect on axial contraction and condensin inactivation does not affect short-range chromatin compaction, inactivation of both pathways causes synergistic defects in chromosome segregation and cell viability. Furthermore, both pathways rely at least partially on the deacetylase Hst2, suggesting that this protein helps coordinating chromatin compaction and axial contraction to properly shape mitotic chromosomes.

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Molecular characterization of 5S ribosomal RNA genes and transcripts in the protozoan parasite Leishmania major

Parasitology ◽

10.1017/s0031182016001712 ◽

2016 ◽

Vol 143 (14) ◽

pp. 1917-1929 ◽

Cited By ~ 3

Author(s):

RODRIGO MORENO-CAMPOS ◽

LUIS E. FLORENCIO-MARTÍNEZ ◽

TOMÁS NEPOMUCENO-MEJÍA ◽

SAÚL ROJAS-SÁNCHEZ ◽

DANIEL E. VÉLEZ-RAMÍREZ ◽

...

Keyword(s):

Leishmania Major ◽

Rna Polymerase Iii ◽

Nuclear Periphery ◽

Ribosomal Subunit ◽

Protozoan Parasite ◽

5S Rrna ◽

Rrna Genes ◽

Rrna Gene ◽

5S Rrna Genes ◽

Pol Iii

SUMMARYEukaryotic 5S rRNA, synthesized by RNA polymerase III (Pol III), is an essential component of the large ribosomal subunit. Most organisms contain hundreds of 5S rRNA genes organized into tandem arrays. However, the genome of the protozoan parasite Leishmania major contains only 11 copies of the 5S rRNA gene, which are interspersed and associated with other Pol III-transcribed genes. Here we report that, in general, the number and order of the 5S rRNA genes is conserved between different species of Leishmania. While in most organisms 5S rRNA genes are normally associated with the nucleolus, combined fluorescent in situ hybridization and indirect immunofluorescence experiments showed that 5S rRNA genes are mainly located at the nuclear periphery in L. major. Similarly, the tandemly repeated 5S rRNA genes in Trypanosoma cruzi are dispersed throughout the nucleus. In contrast, 5S rRNA transcripts in L. major were localized within the nucleolus, and scattered throughout the cytoplasm, where mature ribosomes are located. Unlike other rRNA species, stable antisense RNA complementary to 5S rRNA is not detected in L. major.

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Condensins under the microscope

The Journal of Cell Biology ◽

10.1083/jcb.201804078 ◽

2018 ◽

Vol 217 (7) ◽

pp. 2229-2231 ◽

Cited By ~ 1

Author(s):

Kazuhiro Maeshima ◽

Kayo Hibino ◽

Damien F. Hudson

Keyword(s):

Quantitative Data ◽

Mitotic Chromosome ◽

State Of The Art ◽

Imaging Techniques ◽

Mechanistic Model ◽

Chromosome Condensation ◽

Mitotic Chromosomes ◽

Cell Biol ◽

Key Players ◽

Mitotic Chromosome Condensation

Condensins are key players in mitotic chromosome condensation. Using an elegant combination of state-of-the-art imaging techniques, Walther et al. (2018. J. Cell Biol. https://doi.org/10.1083/jcb.201801048) counted the number of Condensins, examined their behaviors on human mitotic chromosomes, and integrated the quantitative data to propose a new mechanistic model for chromosome condensation.

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Mitotic Chromosome Condensation Driven by a Volume Phase Transition

10.1101/2021.07.30.454418 ◽

2021 ◽

Author(s):

Andrew J Beel ◽

Pierre-Jean Matteï ◽

Roger D Kornberg

Keyword(s):

Phase Transition ◽

Mitotic Chromosome ◽

Chromosome Morphology ◽

Central Axis ◽

Volume Phase Transition ◽

Mitotic Chromosomes ◽

Volume Phase ◽

Change Of State ◽

Physico Chemical ◽

Mitotic Chromosome Condensation

Procedures were devised for the reversible decondensation and recondensation of purified mitotic chromosomes. Computational methods were developed for the quantitative analysis of chromosome morphology in high throughput, enabling the recording of condensation behavior of thousands of individual chromosomes. Established physico-chemical theory for ionic hydrogels was modified for application to chromosomal material and shown to accurately predict the observed condensation behavior. The theory predicts a change of state (a "volume phase transition") in the course of condensation, and such a transition was shown to occur. These findings, together with classical cytology showing loops of chromatin, lead to the description of mitotic chromosome structure in terms of two simple principles: contraction of length of chromatin fibers by the formation of loops, radiating from a central axis; and condensation of the chromosomal material against the central axis through a volume phase transition.

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