Autophagy-related protein Atg11 preserves asymmetric inheritance of spindle microtubule-organizing centers

2021 ◽  
Author(s):  
Md Hashim Reza ◽  
Jigyasa Verma ◽  
Ratul Chowdhury ◽  
Ravi Manjithaya ◽  
Kaustuv Sanyal
Keyword(s):  
Chromosome Segregation ◽  
Protein Interactions ◽  
Molecular Motors ◽  
Growth Dynamics ◽  
In Silico Analysis ◽  
Spindle Pole ◽  
High Rate ◽  
Chromosome Loss ◽  
Spindle Positioning

Asymmetric spindle pole body (SPB) inheritance requires a cascade of events that involve kinases, phosphatases and structural scaffold proteins including molecular motors and microtubule-associated proteins present in the nucleus and/or the cytoplasm. Higher levels of an SPB component Spc72 and the spindle positioning factor Kar9 at the old SPB, which migrates to the daughter cell, ensure asymmetric SPB inheritance. Timely SPB duplication followed by its asymmetric inheritance is a key to correct spindle alignment leading to high-fidelity chromosome segregation. By combining in silico analysis of known protein-protein interactions of autophagy (Atg)-related proteins with those that constitute the chromosome segregation machinery, and growth dynamics of 35 atg mutants in the presence of a microtubule poison, we identified Atg11 as a potential regulator of chromosome transmission. Cells lacking Atg11 did not show any kinetochore defects but displayed a high rate of chromosome loss and delayed anaphase onset. Atg11 positively interacted with Kar9 and Kip2 and negatively with Dyn1 and Kar3 in mediating proper chromosome segregation suggesting a role of Atg11 in spindle positioning. Indeed, atg11∆ cells displayed an inverted SPB inheritance. We further show that Atg11 promotes asymmetric localization of Spc72 and Kar9 on the old SPB. Atg11 physically interacted with Spc72 and transiently localized close to the old SPB during metaphase-to-anaphase progression. Taken together, our study uncovers an autophagy-independent role of Atg11 in spindle alignment and emphasizes the importance of unbiased screens to identify factors mediating the complex and intricate crosstalk between processes fundamental to genomic integrity.

scholarly journals The Role of Cdh1p in Maintaining Genomic Stability in Budding Yeast

Genetics ◽  
2003 ◽  
Vol 165 (2) ◽  
pp. 489-503 ◽  
Author(s):  
Karen E Ross ◽  
Orna Cohen-Fix
Keyword(s):  
Cell Cycle ◽  
Chromosome Segregation ◽  
Spindle Assembly Checkpoint ◽  
Chromosome Loss ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
Growth Defects ◽  
Genes Encoding ◽  
Specificity Factor

Abstract Cdh1p, a substrate specificity factor for the cell cycle-regulated ubiquitin ligase, the anaphase-promoting complex/cyclosome (APC/C), promotes exit from mitosis by directing the degradation of a number of proteins, including the mitotic cyclins. Here we present evidence that Cdh1p activity at the M/G1 transition is important not only for mitotic exit but also for high-fidelity chromosome segregation in the subsequent cell cycle. CDH1 showed genetic interactions with MAD2 and PDS1, genes encoding components of the mitotic spindle assembly checkpoint that acts at metaphase to prevent premature chromosome segregation. Unlike cdh1Δ and mad2Δ single mutants, the mad2Δ cdh1Δ double mutant grew slowly and exhibited high rates of chromosome and plasmid loss. Simultaneous deletion of PDS1 and CDH1 caused extensive chromosome missegregation and cell death. Our data suggest that at least part of the chromosome loss can be attributed to kinetochore/spindle problems. Our data further suggest that Cdh1p and Sic1p, a Cdc28p/Clb inhibitor, have overlapping as well as nonoverlapping roles in ensuring proper chromosome segregation. The severe growth defects of both mad2Δ cdh1Δ and pds1Δ cdh1Δ strains were rescued by overexpressing Swe1p, a G2/M inhibitor of the cyclin-dependent kinase, Cdc28p/Clb. We propose that the failure to degrade cyclins at the end of mitosis leaves cdh1Δ mutant strains with abnormal Cdc28p/Clb activity that interferes with proper chromosome segregation.

scholarly journals The half-bridge component Kar1 promotes centrosome separation and duplication during budding yeast meiosis

2018 ◽  
Vol 29 (15) ◽  
pp. 1798-1810
Author(s):  
Meenakshi Agarwal ◽  
Hui Jin ◽  
Melainia McClain ◽  
Jinbo Fan ◽  
Bailey A. Koch ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Budding Yeast ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Pole Body ◽  
Centrosome Separation ◽  
Chromatid Segregation ◽  
Yeast Meiosis ◽  
Unique Mechanism

The budding yeast centrosome, often called the spindle pole body (SPB), nucleates microtubules for chromosome segregation during cell division. An appendage, called the half bridge, attaches to one side of the SPB and regulates SPB duplication and separation. Like DNA, the SPB is duplicated only once per cell cycle. During meiosis, however, after one round of DNA replication, two rounds of SPB duplication and separation are coupled with homologue segregation in meiosis I and sister-chromatid segregation in meiosis II. How SPB duplication and separation are regulated during meiosis remains to be elucidated, and whether regulation in meiosis differs from that in mitosis is unclear. Here we show that overproduction of the half-bridge component Kar1 leads to premature SPB separation during meiosis. Furthermore, excessive Kar1 induces SPB overduplication to form supernumerary SPBs, leading to chromosome missegregation and erroneous ascospore formation. Kar1-­mediated SPB duplication bypasses the requirement of dephosphorylation of Sfi1, another half-bridge component previously identified as a licensing factor. Our results therefore reveal an unexpected role of Kar1 in licensing meiotic SPB duplication and suggest a unique mechanism of SPB regulation during budding yeast meiosis.

scholarly journals Loss of Kinesin-8 improves the robustness of the acentrosomal spindle

2020 ◽  
Author(s):  
Alberto Pineda-Santaella ◽  
Nazaret Fernández-Castillo ◽  
Ángela Sánchez-Gómez ◽  
Alfonso Fernández-Álvarez
Keyword(s):  
Chromosome Segregation ◽  
Molecular Basis ◽  
Spindle Pole ◽  
High Rate ◽  
Female Meiosis ◽  
Aneuploidy Rate ◽  
Microtubule Organizing Centres ◽  
Spindle Pole Bodies ◽  
Nucleation Mechanisms ◽  
Yeast Meiosis

Chromosome segregation in female meiosis is inherently error-prone, among other reasons, because the acentrosomal spindle assembles and segregates chromosomes without the major microtubule-organizing centres in eukaryotes, the centrosomes, which causes high rate of aneuploidy. The molecular basis underlying formation of acentrosomal spindles is not as well-understood as that of centrosomal spindles and, consequently, strategies to improve spindle robustness are difficult to address. Recently, we noticed during fission yeast meiosis the formation of unexpected microtubules arrays, independent of the spindle pole bodies (yeast centrosome equivalent), with ability to segregate chromosomes. Here, we establish such microtubules formation as bonafide self-assembled spindles that depend on the canonical microtubule crosslinker Ase1/PRC1, share similar structural polarity and harbour the microtubule polymerase Alp14/XMAP215, while being independent of conventional γ-tubulin-mediated nucleation mechanisms. Remarkably, acentrosomal spindle robustness was reinforced by deletion of the Klp6/Kinesin-8, which, consequently, led to a reduced meiotic aneuploidy rate. Our results enlighten the molecular basis of acentrosomal meiosis, a crucial event in understanding gametogenesis.

scholarly journals Cytoplasmic Microtubule Organizing Centers Regulate Meiotic Spindle Positioning in Mouse Oocyte

2020 ◽  
Author(s):  
Daniela Londono Vasquez ◽  
Katherine Rodriguez-Lukey ◽  
Susanta K. Behura ◽  
Ahmed Z. Balboula
Keyword(s):  
Polar Body ◽  
Spindle Pole ◽  
Meiotic Spindle ◽  
Cytoplasmic Microtubule ◽  
Spindle Positioning ◽  
Nuclear Envelope Breakdown ◽  
Oocyte Meiosis ◽  
Polar Bodies ◽  
Polar Body Extrusion

ABSTRACTDuring oocyte meiosis, migration of the spindle and its positioning must be tightly regulated to ensure elimination of the polar bodies and provide developmentally competent euploid eggs. Although the role of F-actin in regulating these critical processes has been studied extensively, little is known whether microtubules (MTs) participate in regulating these processes. Here, we characterize a pool of MTOCs in the oocyte that does not contribute to spindle assembly but instead remains free in the cytoplasm during metaphase I (metaphase cytoplasmic MTOCs; mcMTOCs). In contrast to spindle pole MTOCs, which primarily originate from the perinuclear region in prophase I, the mcMTOCs are found near the cortex of the oocyte. At nuclear envelope breakdown, they exhibit robust nucleation of MTs, which diminishes during polar body extrusion before returning robustly during metaphase II. The asymmetric positioning of the mcMTOCs provides the spindle with a MT-based anchor line to the cortex opposite the site of polar body extrusion. Depletion of mcMTOCs, by laser ablation, or manipulating their numbers, through autophagy inhibition, revealed that the mcMTOCs are required to regulate the timely migration and positioning of the spindle in meiosis. We discuss how forces exerted by F-actin in mediating movement of the spindle to the oocyte cortex are balanced by MT-mediated forces from the mcMTOCs to ensure spindle positioning and timely spindle migration.

scholarly journals Basic mechanism for biorientation of mitotic chromosomes is provided by the kinetochore geometry and indiscriminate turnover of kinetochore microtubules

2015 ◽  
Vol 26 (22) ◽  
pp. 3985-3998 ◽  
Author(s):  
Anatoly V. Zaytsev ◽  
Ekaterina L. Grishchuk
Keyword(s):  
Error Correction ◽  
Chromosome Segregation ◽  
Basic Mechanism ◽  
Spindle Pole ◽  
Mitotic Division ◽  
Human Cells ◽  
Chromosome Loss ◽  
Mitotic Chromosomes ◽  
Optimal Regime ◽  
Sister Kinetochore

Accuracy of chromosome segregation relies on the ill-understood ability of mitotic kinetochores to biorient, whereupon each sister kinetochore forms microtubule (MT) attachments to only one spindle pole. Because initial MT attachments result from chance encounters with the kinetochores, biorientation must rely on specific mechanisms to avoid and resolve improper attachments. Here we use mathematical modeling to critically analyze the error-correction potential of a simplified biorientation mechanism, which involves the back-to-back arrangement of sister kinetochores and the marked instability of kinetochore–MT attachments. We show that a typical mammalian kinetochore operates in a near-optimal regime, in which the back-to-back kinetochore geometry and the indiscriminate kinetochore–MT turnover provide strong error-correction activity. In human cells, this mechanism alone can potentially enable normal segregation of 45 out of 46 chromosomes during one mitotic division, corresponding to a mis-segregation rate in the range of 10−1–10−2 per chromosome. This theoretical upper limit for chromosome segregation accuracy predicted with the basic mechanism is close to the mis-segregation rate in some cancer cells; however, it cannot explain the relatively low chromosome loss in diploid human cells, consistent with their reliance on additional mechanisms.

scholarly journals Fission Yeast Bub1 Is a Mitotic Centromere Protein Essential for the Spindle Checkpoint and the Preservation of Correct Ploidy through Mitosis

1998 ◽  
Vol 143 (7) ◽  
pp. 1775-1787 ◽  
Author(s):  
Pascal Bernard ◽  
Kevin Hardwick ◽  
Jean-Paul Javerzat
Keyword(s):  
Fission Yeast ◽  
Genomic Instability ◽  
Chromosome Segregation ◽  
Mitotic Spindle ◽  
Spindle Checkpoint ◽  
Chromosome Loss ◽  
Checkpoint Response ◽  
Chromosome Missegregation ◽  
Centromere Function

The spindle checkpoint ensures proper chromosome segregation by delaying anaphase until all chromosomes are correctly attached to the mitotic spindle. We investigated the role of the fission yeast bub1 gene in spindle checkpoint function and in unperturbed mitoses. We find that bub1+ is essential for the fission yeast spindle checkpoint response to spindle damage and to defects in centromere function. Activation of the checkpoint results in the recruitment of Bub1 to centromeres and a delay in the completion of mitosis. We show that Bub1 also has a crucial role in normal, unperturbed mitoses. Loss of bub1 function causes chromosomes to lag on the anaphase spindle and an increased frequency of chromosome loss. Such genomic instability is even more dramatic in Δbub1 diploids, leading to massive chromosome missegregation events and loss of the diploid state, demonstrating that bub1+ function is essential to maintain correct ploidy through mitosis. As in larger eukaryotes, Bub1 is recruited to kinetochores during the early stages of mitosis. However, unlike its vertebrate counterpart, a pool of Bub1 remains centromere-associated at metaphase and even until telophase. We discuss the possibility of a role for the Bub1 kinase after the metaphase–anaphase transition.

scholarly journals Moonlighting of mitotic regulators in cilium disassembly

2021 ◽  
Author(s):  
Cenna Doornbos ◽  
Ronald Roepman
Keyword(s):  
Chromosome Segregation ◽  
Protein Interactions ◽  
Spindle Pole ◽  
Specific Protein ◽  
Protein Protein Interactions ◽  
Functional Protein ◽  
Cellular Processes ◽  
Microtubule Organizing Centres ◽  
Tumour Formation ◽  
Protein Modules

AbstractCorrect timing of cellular processes is essential during embryological development and to maintain the balance between healthy proliferation and tumour formation. Assembly and disassembly of the primary cilium, the cell’s sensory signalling organelle, are linked to cell cycle timing in the same manner as spindle pole assembly and chromosome segregation. Mitotic processes, ciliary assembly, and ciliary disassembly depend on the centrioles as microtubule-organizing centres (MTOC) to regulate polymerizing and depolymerizing microtubules. Subsequently, other functional protein modules are gathered to potentiate specific protein–protein interactions. In this review, we show that a significant subset of key mitotic regulator proteins is moonlighting at the cilium, among which PLK1, AURKA, CDC20, and their regulators. Although ciliary assembly defects are linked to a variety of ciliopathies, ciliary disassembly defects are more often linked to brain development and tumour formation. Acquiring a better understanding of the overlap in regulators of ciliary disassembly and mitosis is essential in finding therapeutic targets for the different diseases and types of tumours associated with these regulators.

scholarly journals Csi1 links centromeres to the nuclear envelope for centromere clustering

2012 ◽  
Vol 199 (5) ◽  
pp. 735-744 ◽  
Author(s):  
Haitong Hou ◽  
Zhou Zhou ◽  
Yu Wang ◽  
Jiyong Wang ◽  
Scott P. Kallgren ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Protein ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
High Rate ◽  
Chromosome Loss ◽  
Functional Importance ◽  
Pole Body ◽  
Domain Protein ◽  
Sun Domain

In the fission yeast Schizosaccharomyces pombe, the centromeres of each chromosome are clustered together and attached to the nuclear envelope near the site of the spindle pole body during interphase. The mechanism and functional importance of this arrangement of chromosomes are poorly understood. In this paper, we identified a novel nuclear protein, Csi1, that localized to the site of centromere attachment and interacted with both the inner nuclear envelope SUN domain protein Sad1 and centromeres. Both Csi1 and Sad1 mutants exhibited centromere clustering defects in a high percentage of cells. Csi1 mutants also displayed a high rate of chromosome loss during mitosis, significant mitotic delays, and sensitivity to perturbations in microtubule–kinetochore interactions and chromosome numbers. These studies thus define a molecular link between the centromere and nuclear envelope that is responsible for centromere clustering.

Point mutations that separate the role of Saccharomyces cerevisiae centromere binding factor 1 in chromosome segregation from its role in transcriptional activation.

Genetics ◽  
1993 ◽  
Vol 135 (2) ◽  
pp. 287-296
Author(s):  
P K Foreman ◽  
R W Davis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Chromosome Segregation ◽  
Transcriptional Activation ◽  
Point Mutations ◽  
Random Mutagenesis ◽  
Chromosome Loss ◽  
Gene Encoding ◽  
Helix Loop Helix ◽  
Binding Factor

Abstract Centromere binding factor 1 (Cbf1p or CP1) binds to the CDEI region of Saccharomyces cerevisiae centromeres and is a member of the basic helix-loop-helix (bHLH) class of proteins. Deletion of the gene encoding Cbf1p results in an increased frequency of chromosome loss, hypersensitivity to low levels of microtubule disrupting drugs (such as thiabendazole and benomyl) and methionine auxotrophy. By polymerase chain reaction-based random mutagenesis of the CBF1 gene we have obtained a number of mutant alleles that make full-length protein with impaired function. The mutations in these alleles are clustered in or just downstream from the bHLH domain. Among the alleles obtained was a class that was more compromised for transcriptional activation and a class that was more compromised for chromosome loss and thiabendazole hypersensitivity. These results indicate that at least some aspects of the role of Cbf1p in chromosome segregation and transcriptional activation are distinct. In contrast, increased chromosome loss and thiabendazole hypersensitivity were not separated in any of the alleles, suggesting that these phenotypes reflect the same mechanistic defect. These observations are consistent with a model that suggests that one role of Cbf1p in chromosome segregation may be to improve the efficiency with which contact between the kinetochore and spindle microtubules is established or maintained.

Role of small nuclear RNAs in eukaryotic gene expression

2013 ◽  
Vol 54 ◽  
pp. 79-90 ◽  
Author(s):  
Saba Valadkhan ◽  
Lalith S. Gunawardane
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Rna Stability ◽  
Splice Sites ◽  
Branch Site ◽  
Small Nuclear Rnas ◽  
Eukaryotic Gene Expression ◽  
Eukaryotic Gene ◽  
Rna Biogenesis

Eukaryotic cells contain small, highly abundant, nuclear-localized non-coding RNAs [snRNAs (small nuclear RNAs)] which play important roles in splicing of introns from primary genomic transcripts. Through a combination of RNA–RNA and RNA–protein interactions, two of the snRNPs, U1 and U2, recognize the splice sites and the branch site of introns. A complex remodelling of RNA–RNA and protein-based interactions follows, resulting in the assembly of catalytically competent spliceosomes, in which the snRNAs and their bound proteins play central roles. This process involves formation of extensive base-pairing interactions between U2 and U6, U6 and the 5′ splice site, and U5 and the exonic sequences immediately adjacent to the 5′ and 3′ splice sites. Thus RNA–RNA interactions involving U2, U5 and U6 help position the reacting groups of the first and second steps of splicing. In addition, U6 is also thought to participate in formation of the spliceosomal active site. Furthermore, emerging evidence suggests additional roles for snRNAs in regulation of various aspects of RNA biogenesis, from transcription to polyadenylation and RNA stability. These snRNP-mediated regulatory roles probably serve to ensure the co-ordination of the different processes involved in biogenesis of RNAs and point to the central importance of snRNAs in eukaryotic gene expression.

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