Polyploid Cells Research Articles

Therapeutic potential of PLK1 inhibition in triple-negative breast cancer

Triple negative breast cancer (TNBC) is responsible for significant number of breast cancer-associated deaths because of lacking of successful molecular-targeted therapy. To explore a therapeutic target for TNBC, we performed a siRNA-mediated knockdown screening and identified Polo-like kinase 1 (PLK1) as a potential therapeutic target for TNBC. Knockdown of PLK1 as well as a small compound inhibitor for PLK1, BI-2536, induced G2/M arrest and created polyploid cell population, shown by increased DNA content and nuclear size. Inhibition of PLK1 eventually triggered apoptosis in multiple TNBC cell lines. In addition, we confirmed that PLK1 was significantly overexpressed in the tissues from TNBC patients compared with the tissues of normal mammary glands and benign breast tumors. Our data indicated that PLK1 plays a pivotal role in the regulation of mitosis of TNBC cells. Although future in vivo studies are warranted, targeting PLK1 by a selective inhibitor such as BI-2536 can be an attractive molecular-targeted therapy for TNBC.

Structural and functional differentiation of a fat body-like tissue adhering to testis follicles facilitates spermatogenesis in locusts

The fat body is distributed throughout the body of insects, playing the essential role in intermediary metabolism and nutrient storage. However, the function of differentiation of fat bodies adhering to different tissues remains largely unknown. Here, we identified a fat body-like tissue (FLT) surrounding testis follicles and described its features at morphological, cellular and molecular levels. The FLT is morphologically distinguished with the abdominal fat body (FB) and dominated by diploid cells instead of polyploid cells. The transcriptomic analysis demonstrated that the FLT and FB have dramatically different gene expression profiles. Moreover, genes in the cell cycle pathway, which include both DNA replication- and cell division-related genes, were successively active during development of the FLT, suggesting that FLT cells possibly undergo a mitotic cycle rather than an endocycle. Deprivation of the FLT resulted in distortion of the testis follicles, disappearance of sperm bundles, reduction of total sperm number and increase of dead sperm, indicating a critical role of the FLT in the spermatogenesis in testis follicles. The special functional differentiation of the two similar tissues suggested that FLT-FB cells are able to establish a promising system to study mitotic-to-endocycle transition.

Heat Stress-Induced Multiple Multipolar Divisions of Human Cancer Cells.

Multipolar divisions of heated cells has long been thought to stem from centrosome aberrations of cells directly caused by heat stress. In this paper, through long-term live-cell imaging, we provide direct cellular evidences to demonstrate that heat stress can promote multiple multipolar divisions of MGC-803 and MCF-7 cells. Our results show that, besides facilitating centrosome aberration, polyploidy induced by heat stress is another mechanism that causes multipolar cell divisions, in which polyploid cancer cells engendered by mitotic slippage, cytokinesis failure, and cell fusion. Furthermore, we also find that the fates of theses polyploid cells depend on their origins, in the sense that the polyploid cells generated by mitotic slippage experience bipolar divisions with a higher rate than multipolar divisions, while those polyploid cells induced by both cytokinesis failure and cell fusion have a higher frequency of multipolar divisions compared with bipolar divisions. This work indicates that heat stress-induced multiple multipolar divisions of cancer cells usually produce aneuploid daughter cells, and might lead to genetically unstable cancer cells and facilitate tumor heterogeneity.

A CRISPR/Cas9 method to generate heterozygous alleles in Saccharomyces cerevisiae.

Saccharomyces cerevisiae is a genetically facile organism, yet multiple CRISPR/Cas9 techniques are widely used to edit its genome more efficiently and cost effectively than conventional methods. The absence of selective markers makes CRISPR/Cas9 editing particularly useful when making mutations within genes or regulatory sequences. Heterozygous mutations within genes frequently arise in the winners of evolution experiments. The genetic dissection of heterozygous alleles can be important to understanding gene structure and function. Unfortunately, the high efficiency of genome cutting and repair makes the introduction of heterozygous alleles by standard CRISPR/Cas9 technique impossible. To be able to quickly and reliably determine the individual phenotypes of the thousands of heterozygous mutations that can occur during directed evolutions is of particular interest to industrial strain improvement research. In this report, we describe a CRISPR/Cas9 method that introduces specific heterozygous mutations into the S.cerevisiae genome. This method relies upon creating silent point mutations in the protospacer adjacent motif site or removing the protospacer adjacent motif site entirely to stop the multiple rounds of genome editing that prevent heterozygous alleles from being generated. This technique should be able to create heterozygous alleles in other diploid yeasts and different allelic copy numbers in polyploid cells.

Activation of Meiotic Genes Mediates Ploidy Reduction and Generates Phenotypic Diversity During Cryptococcal Infection

Cryptococcus neoformans is a global human fungal pathogen that causes fatal meningoencephalitis in mostly immunocompromised individuals. During pulmonary infection, cryptococcal cells form big polyploid cells that exhibit increased resistance to host immune attack and are proposed to contribute to the latency of cryptococcal infection. These polyploidy titan cells can generate haploid and aneuploid progeny that may result in systematic infection. What triggers cryptococcal polyploidization and how ploidy reduction is achieved remain open questions. Here we discovered that Cryptococcus cells polyploidize in response to genotoxic stresses that cause DNA double strand breaks. Intriguingly, meiosis-specific genes are activated in C. neoformans and contribute to ploidy reduction, both in vitro and during infection in mice. Strikingly, cryptococcal cells that activated their meiotic genes in mice showed increased phenotypic diversity and were resistant to specific genotoxic stress comparing with sister cells recovered from the same host tissue but without activation of meiotic genes. Our findings support the idea that meiotic genes, in addition to the conventional roles during classic sexual reproduction, contribute to adaptation of eukaryotic cells undergoing dramatic genome changes in response to genotoxic stress. The discovery has additional implication for evolution of sexual reproduction and the paradox of the presence of meiotic machinery in asexual species. Finally, our findings in this eukaryotic microbe mirror the revolutionary discoveries of gametogenesis (polyploidization and meiosis-like process) in cancer cells, suggesting that the gametogenesis process itself could provide a general mechanism for rejuvenation in response to stress to promote individual survival.

Wound-induced polyploidization is driven by Myc and supports tissue repair in the presence of DNA damage.

ABSTRACTTissue repair usually requires either polyploid cell growth or cell division, but the molecular mechanism promoting polyploidy and limiting cell division remains poorly understood. Here, we find that injury to the adult Drosophila epithelium causes cells to enter the endocycle through the activation of Yorkie-dependent genes (Myc and E2f1). Myc is even sufficient to induce the endocycle in the uninjured post-mitotic epithelium. As result, epithelial cells enter S phase but mitosis is blocked by inhibition of mitotic gene expression. The mitotic cell cycle program can be activated by simultaneously expressing the Cdc25-like phosphatase String (stg), while genetically depleting APC/C E3 ligase fizzy-related (fzr). However, forcing cells to undergo mitosis is detrimental to wound repair as the adult fly epithelium accumulates DNA damage, and mitotic errors ensue when cells are forced to proliferate. In conclusion, we find that wound-induced polyploidization enables tissue repair when cell division is not a viable option.

A Novel Polyhalogenated Monoterpene Induces Cell Cycle Arrest and Apoptosis in Breast Cancer Cells.

Breast cancer is the most common cancer type and a primary cause of cancer mortality among females worldwide. Here, we analyzed the anticancer efficacy of a novel bromochlorinated monoterpene, PPM1, a synthetic analogue of polyhalogenated monoterpenes from Plocamium red algae and structurally similar non-brominated monoterpenes. PPM1, but not the non-brominated monoterpenes, decreased selectively the viability of several triple-negative as well as triple-positive breast cancer cells with different p53 status without significantly affecting normal breast epithelial cells. PPM1 induced accumulation of triple-negative MDA-MB-231 cells with 4N DNA content characterized by decreased histone H3-S10/T3 phosphorylation indicating cell cycle arrest in the G2 phase. Western immunoblot analysis revealed that PPM1 treatment triggered an initial rapid activation of Aurora kinases A/B/C and p21Waf1/Cip1 accumulation, which was followed by accumulation of polyploid >4N cells. Flow cytometric analysis showed mitochondrial potential disruption, caspase 3/7 activation, phosphatidylserine externalization, reduction of the amount polyploid cells, and DNA fragmentation consistent with induction of apoptosis. Cell viability was partially restored by the pan-caspase inhibitor Z-VAD-FMK indicating caspase contribution. In vivo, PPM1 inhibited growth, proliferation, and induced apoptosis in MDA-MB-231 xenografted onto the chick chorioallantoic membrane. Hence, Plocamium polyhalogenated monoterpenes and synthetic analogues deserve further exploration as promising anticancer lead compounds.

When Three Isn't a Crowd: A Digyny Concept for Treatment-Resistant, Near-Triploid Human Cancers.

Near-triploid human tumors are frequently resistant to radio/chemotherapy through mechanisms that are unclear. We recently reported a tight association of male tumor triploidy with XXY karyotypes based on a meta-analysis of 15 tumor cohorts extracted from the Mitelman database. Here we provide a conceptual framework of the digyny-like origin of this karyotype based on the germline features of malignant tumors and adaptive capacity of digyny, which supports survival in adverse conditions. Studying how the recombinatorial reproduction via diploidy can be executed in primary cancer samples and HeLa cells after DNA damage, we report the first evidence that diploid and triploid cell sub-populations constitutively coexist and inter-change genomes via endoreduplicated polyploid cells generated through genotoxic challenge. We show that irradiated triploid HeLa cells can enter tripolar mitosis producing three diploid sub-subnuclei by segregation and pairwise fusions of whole genomes. Considering the upregulation of meiotic genes in tumors, we propose that the reconstructed diploid sub-cells can initiate pseudo-meiosis producing two “gametes” (diploid “maternal” and haploid “paternal”) followed by digynic-like reconstitution of a triploid stemline that returns to mitotic cycling. This process ensures tumor survival and growth by (1) DNA repair and genetic variation, (2) protection against recessive lethal mutations using the third genome.

Building bridges between chromosomes: novel insights into the abscission checkpoint.

In the presence of chromatin bridges, mammalian cells delay completion of cytokinesis (abscission) to prevent chromatin breakage or tetraploidization by regression of the cleavage furrow. This abscission delay is called "the abscission checkpoint" and is dependent on Aurora B kinase. Furthermore, cells stabilize the narrow cytoplasmic canal between the two daughter cells until the DNA bridges are resolved. Impaired abscission checkpoint signaling or unstable intercellular canals can lead to accumulation of DNA damage, aneuploidy, or generation of polyploid cells which are associated with tumourigenesis. However, the molecular mechanisms involved have only recently started to emerge. In this review, we focus on the molecular pathways of the abscission checkpoint and describe newly identified triggers, Aurora B-regulators and effector proteins in abscission checkpoint signaling. We also describe mechanisms that control intercellular bridge stabilization, DNA bridge resolution, or abscission checkpoint silencing upon satisfaction, and discuss how abscission checkpoint proteins can be targeted to potentially improve cancer therapy.

PROBING BREAST CANCER THERAPEUTIC RESPONSES BY DNA CONTENT PROFILING

Background. Discrepancies in the interpretation of breast cancer therapeutic responses still exist mainly because of lack of standardized assessment criteria and methods. Objective. DNA content profiling of cells in the affected (cancerous) tissue before and after neoadjuvant chemotherapy (NAC) was applied to facilitate interpretation of therapeutic responses. Methods. Both diagnostic biopsy and operation materials representing the tissue of primary tumors surgically removed after NAC were subjected to DNA image cytometry. Polyploidy and aneuploidy in DNA histograms were evaluated with a prognostic Auer typing. Stemline DNA index (DI) values and percentages of cells that polyploidize (>4.5C) were also determined. Immunofluorescence staining was applied to evaluate proliferation (Ki-67), invasiveness (CD44), and self-renewal factors characteristic for stem cells (SOX2 and NANOG). Results. DNA content profiles of 12 breast cancer cases, of which 7 were triple-negative, revealed the features of tumor non-responsiveness to NAC in 7 cases, of which 5 were triple-negative. Among non-responsive cases there were 3 cases that showed enhanced polyploidization, suggesting the negative NAC effect. Near-triploid (DI=1.26-1.74) triple-negative cases were determined as most resistant to NAC. Cycling near-triploid cells may contribute to the excessive numbers of >4.5C cells. Polyploid cells were positive for Ki-67, CD44, SOX2, and NANOG. Conclusions. DNA content profiling data provide additional helpful information for interpreting therapeutic responses in NAC-treated breast cancers. Polyploid tumor cells possessing stem cell features can be induced by NAC. Because NAC effects in some cases may be unfavorable, the use of the further treatment strategy should be carefully considered.

A Cyclin A-Myb-MuvB-Aurora B network regulates the choice between mitotic cycles and polyploid endoreplication cycles.

Endoreplication is a cell cycle variant that entails cell growth and periodic genome duplication without cell division, and results in large, polyploid cells. Cells switch from mitotic cycles to endoreplication cycles during development, and also in response to conditional stimuli during wound healing, regeneration, aging, and cancer. In this study, we use integrated approaches in Drosophila to determine how mitotic cycles are remodeled into endoreplication cycles, and how similar this remodeling is between induced and developmental endoreplicating cells (iECs and devECs). Our evidence suggests that Cyclin A / CDK directly activates the Myb-MuvB (MMB) complex to induce transcription of a battery of genes required for mitosis, and that repression of CDK activity dampens this MMB mitotic transcriptome to promote endoreplication in both iECs and devECs. iECs and devECs differed, however, in that devECs had reduced expression of E2F1-dependent genes that function in S phase, whereas repression of the MMB transcriptome in iECs was sufficient to induce endoreplication without a reduction in S phase gene expression. Among the MMB regulated genes, knockdown of AurB protein and other subunits of the chromosomal passenger complex (CPC) induced endoreplication, as did knockdown of CPC-regulated cytokinetic, but not kinetochore, proteins. Together, our results indicate that the status of a CycA—Myb-MuvB—AurB network determines the decision to commit to mitosis or switch to endoreplication in both iECs and devECs, and suggest that regulation of different steps of this network may explain the known diversity of polyploid cycle types in development and disease.

Abstract 2902: Non-genetic TPX2/AURKA signaling facilitates tumor evolution in EGFR-TKI resistance in NSCLC

Abstract Background: EGFR tyrosine kinase inhibitors are effective for treatment of non-small cell lung cancer. Benefit from these therapies is often transient due to residual disease leading to acquired resistance. Osimertinib now used in first-line setting, but mechanisms of resistance are ill-defined and no effective second-line therapies exist. Disease progression occurs through tumor evolution that involves the emergence of distinct genetic alterations and non-genetic changes in cell state. In this study, we determine the molecular drivers of drug sensitivity and tumor evolution which forms the basis of clinical genomic heterogeneity. We show that the increase in genomic instability correlates with increased chromosomal alterations or rate of mutation which creates evolutionary dependencies. Method: Using our previously established EGFR-TKI acquired resistance models, we observed heightened TPX2/AURKA activity. We measured the temporal activation of TPX2/AURKA in sensitive phase, a drug-tolerant phase (residual phase) and acquired resistant phase. Chromosomal segregation errors, mitotic deformation and aneuploidy were used as a functional read-out to the measure genomic instability upon EGFR-TKI treatment. Using clinical specimens, the contribution of non-genetic TPX2/AURKA axis in disease progression was evaluated. Results: In-vitro models of residual disease exhibited increased AURKA activation via upregulation of its co-activator TPX2. Pharmacological inhibition of AURKA activation by MLN8237 together with osimertinib enhanced the magnitude of response and forestalled the emergence of resistance. Therefore, tumor cells reply on AURKA to transition from a sensitive to drug-tolerant phase and is maintained in acquired resistance. Since high levels of TPX2 and AURKA are known to cause mitotic errors and polyploidy, we surveyed for mitotic defects induced by EGFR-TKI treatment. EGFR-TKI resulted in a marked accumulation of errors in centrosome biogenesis, spindle assembly, and chromosome segregation resulting in polyploid cells with abnormal DNA content indicating persistent mitotic stress is a feature of EGFR inhibition. We conclude that the abnormal activation of TPX2/AURKA leaves a signature of defects associated with mitotic stress, a prominent feature of genomic instability. Examination of clinical samples with T790M mutation or MET amplification upon disease progression had high levels of TPX2. This nominates TPX2 as a novel biomarker therapeutic resistance in a significant fraction of EGFR-mutant lung cancers. Conclusion: Our results indicate that heightened TPX2/AURKA activation is required for tumor cells to transition out of sensitive cell state and persist upon drug pressure. This deregulation creates a highly genomically unstable environment and give rise to genomic heterogeneity providing a fertile ground for the subsequent survival of fittest subclones. Citation Format: Khyati N. Shah, Roma Bhatt, Julia Rotow, Julia Rohrberg, Victor Olivas, Victoria E. Wang, Jonathan Kuhn, Sophie Dumont, Frank Mccormick, Andrei Goga, Collin M. Blakely, Trever G. Bivona, Sourav Bandyopadhyay. Non-genetic TPX2/AURKA signaling facilitates tumor evolution in EGFR-TKI resistance in NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2902.

1,3-Dinitrobenze-Induced Genotoxicity Through Altering Nuclear Integrity of Diploid and Polyploidy Germ Cells.

1,3-Dinitrobenzene (mDNB) is a widely used intermediate in commercial products and causes testicular injury. However, genotoxic effects upon low-level exposure are poorly understood. The present study evaluated the effects of very low-chronic doses of mDNB on sperm nuclear integrity. Male hamsters were treated with 1.5 mg/kg/d/4 wks (group A), 1.5 mg/kg/mDNB/d/week/4 weeks (group B), 1.0 mg/kg/mDNB/3 d/wk/4 wks (group C), or polyethylene glycol 600 (control). Nuclear integrity of distal cauda epididymal sperm was determined using the sperm chromatin structure assay and acridine orange staining (AOS). The germ cell nuclear integrity was assessed by the comet assay. Testicular histopathology was conducted to evaluate the sensitive stages. The comet assay revealed denatured nuclear DNA in group A (in diploid and polyploid cells from weeks 2-5); respectively at week 4 and weeks 3 to 4 in groups B and C. According to AOS, only group A animals exhibited denatured sperm DNA (weeks 1 and 3). The effective sperm count declined from weeks 1 to 6. Mean sperm DNA denaturation extent, percentage cells outside the main population, and standard deviation indicated altered sperm nuclear integrity in group A. Same animals exhibited progressive disruption of the Sertoli cells, while groups B and C exhibited damages on germ cells. The results suggest that mDNB affects sperm nuclear integrity at very low chronic doses targeting cell-specific testicular damage.

Abstract 3774: Elevated cancer evolution dynamics: Emergence of polyploid cancer cells in response to multimodal therapy as an adaptive response on both individual and collective levels

Abstract Polyploidy has been often associated with poor prognosis and has been described to some extent in histological samples for most types of solid tumors. In cancer biology, it has been proposed to induce the gain of a stem-like phenotype and drive tumor progression by increasing the potential for cellular transformation. Experiments with chemotherapy gradients in our engineered microfluidic device with prostate cancer cell lines exhibited a stiff polyploidy distribution pattern of heterogeneous cell populations across the gradient. This demonstrates the generation of polyploid giant cancer cells (PGCCs) is an emergent response to high doses across the microfluidic drug stress landscape. Furthermore, in these experiments we observed polyploidization events occurring concurrently with the development of drug resistance and delayed relapse by non-polypoidal cell budding off from PGCCs leading to a complete repopulation, even in the high-dose areas of the array. Recent publications suggest polyploidy might play a crucial role in resistance to chemo-, hormone- and radio-therapy across most solid tumors investigated, thus independent of the therapy’s mechanism of action. To confirm these findings, we conducted experiments in conventional cell culture using different chemotherapeutic agents and ionizing radiation in several cell lines and confirmed the development of polyploidy was a common feature among them. Although these data supporting the association of polyploidy with resistance are extensive, the effects of these therapies on tumor heterogeneity and the cancer stem niche remains unknown. Different molecular mechanisms, such as mitotic slippage, aborted cytokinesis, endo-replication or cell fusion can lead to polyploidy. We hypothesize a subset of polyploid cells with stem-like features may be the reservoir of therapeutic resistance in cancer. In this work we focused on the characterization of this subset of cells within the tumor leading to the polyploid-stem phenotype, particularly the subpopulation with potential to reverse polyploidy and repopulate tumor heterogeneity after therapy. In order to establish causality and chronology of these phenomena, as well as quantify the complex population dynamics involved in this process, we engineered several cell lines from various common types of solid tumors with two fluorescence biosensors enabling simultaneous monitoring of SOX2/OCT4 activity and cell cycle status at the single-cell level using high-throughput confocal videomicroscopy. The emergence of polyploid cancer cells in response to multimodal therapy as an adaptive response on both individual and collective levels may be a hallmark of elevated cancer evolution dynamics and constitutes a promising potential target to prevent relapse. Citation Format: Gonzalo Torga, Ke-Chih Lin, Kayla Myers, Kimberly Smith, Robert H. Austin, Kenneth J. Pienta. Elevated cancer evolution dynamics: Emergence of polyploid cancer cells in response to multimodal therapy as an adaptive response on both individual and collective levels [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3774.

Hepatic polyploidy: Dr Jekyll or Mr Hyde

Polyploidy (alias whole genome amplification) refers to organisms containing more than two basic sets of chromosomes. Polyploidy was first observed in plants more than a century ago, and it is known that such processes occur in many eukaryotes under a variety of circumstances. In mammals, the development of polyploid cells can contribute to tissue differentiation and therefore possibly a gain of function. Alternately, it can be associated with development of disease such as cancer. Polyploidy can occur because of cell fusion or abnormal cell division. Polyploidy is a common characteristic of the mammalian liver. Polyploidization occurs notably during liver development, but also in adults because of cellular stress. Recent progresses have unraveled the mechanisms and functional consequences of hepatocytes polyploidization during normal and pathological liver growth.

Modulation of megakaryopoiesis and platelet production during inflammation

Megakaryocytes (MKs) are widely known as the progenitor cells of platelets. These large, polyploid cells are a derivative of the hematopoietic stem cell (HSC), and reside in the bone marrow, lining blood vessel walls where they release their platelet progeny into circulation. Although little is known about how MKs differ under various environmental stressors, both chronic and acute inflammation alter the differentiation and molecular content of MKs. Furthermore, evidence suggests that the release of inflammatory cytokines may induce MK rupture and rapid release of platelets as a mechanism to quickly replenish diminished platelet counts in response to inflammation. Similarities between MKs and their close relatives, white blood cells, have introduced the notion that MKs may play a role in combating infection by engulfing and presenting antigens, and passing this information to circulating platelets. In addition, MKs exposed to varying bone marrow environments produce different platelets which enter circulation primed to respond to and combat inflammation, infection, or injury. This review focuses on how inflammation alters MK production, maturation, and platelet production. In addition, it introduces the idea that inflammation reprograms MKs to create different, more pathogenic platelets and leads them to take on different roles as responders to deleterious conditions. In the future, studies determining how platelets are altered in disease states may lead to novel MK- and platelet-based therapeutic targets to mitigate inflammation-related morbidity and mortality.

Karyological Study of Siberian Larch Species Larix sibirica and Larix gmelinii in Taimyr

A comparative study of karyotypes of two Siberian species of larch—L. sibirica and L. gmelinii—growing in the zone of forest-tundra on the Taimyr Peninsula was carried out. The number of chromosomes has been analyzed and the increase in chromosomal instability in larches in forest-tundra conditions has been shown. The size and morphology of chromosomes and the localization and frequency of secondary constrictions in chromosomes of L. sibirica and L. gmelinii were estimated. The parameters of chromosomes in polyploid cells of L. gmelinii were studied. The asymmetry of karyotypes of L. sibirica and L. gmelinii was analyzed. The spectrum and frequency of chromosome rearrangements in L. sibirica and L. gmelinii were studied. Using the data of the karyological analysis, the issues of differentiation and adaptation of the investigated larch species in Taimyr are discussed.

The DNA damage checkpoint and the spindle position checkpoint: guardians of meiotic commitment.

Exogenous signals induce cells to enter the specialized cell division process of meiosis, which produces haploid gametes from diploid progenitor cells. Once cells initiate the meiotic divisions, it is imperative that they complete meiosis. Inappropriate exit from meiosis and entrance into mitosis can create polyploid cells and can lead to germline tumors. Saccharomyces cerevisiae cells enter meiosis when starved of nutrients but can return to mitosis if provided nutrient-rich medium before a defined commitment point. Once past the meiotic commitment point in prometaphase I, cells stay committed to meiosis even in the presence of a mitosis-inducing signal. Recent research investigated the maintenance of meiotic commitment in budding yeast and found that two checkpoints that do not normally function in meiosis I, the DNA damage checkpoint and the spindle position checkpoint, have crucial functions in maintaining meiotic commitment. Here, we review these findings and discuss how the mitosis-inducing signal of nutrient-rich medium could activate these two checkpoints in meiosis to prevent inappropriate meiotic exit.

Spectraplakin Shot Maintains Perinuclear Microtubule Organization in Drosophila Polyploid Cells

Polyploid cells endoreplicate their DNA through a modified cell cycle that skips mitosis as part of their differentiation programs. Upon cell-cycle exit and differentiation, non-centrosomal sites govern microtubule distribution in most cells. Little is known on how polyploid cells, differentiated but cycling, organize their microtubules. We show that microtubules in Drosophila adipocytes and other polyploid tissues form a dense perinuclear cortex responsible for nuclearsize and position. Confirming a relation between this perinuclear cortex and the polyploid endocycle,polyploidization of normally diploid cells was sufficient for cortex formation. A critical component of the perinuclear microtubule organizer (pnMTOC) is Shot, absence of which caused collapse of the perinuclear network into a condensed organizer through kinesin-dependent microtubule sliding. Furthermore, this ectopic organizer was capable of directing partial assembly of a deeply disruptive cytokinesis furrow. In all, our study revealed the importance of perinuclear microtubule organization for stability of endocycling Drosophila cells.

Chromosomal anomalies in dairy cattle as reasons of impaired fertility

Aim. The evaluation of animals for the presence of chromosomal anomalies is one of the main tasks of prac- tical selection, aimed at detecting undesired chromosomal anomalies in early age, which may have negative impact on the reproductive and productive capability of cows and lead to considerable economic losses. The aim of the work is a cytogenetic analysis of the chromosome set in cows of Ukrainian Red-and-Motley dairy cattle breed, which will allow assuming a decrease in reproductive functions with chromosomal aberrations. Methods. We examined 53 cows of the Ukrainian Red-and-Motley dairy cattle breed in SE Research Farm Khrystynivske, IABG named after M.V. Zubets, NAAS. The investigation of chromosomal anomalies involved 72-h cultivation of lymphocytes from the peripheral blood of animals using the common methods. During a routine analysis the preparations were stained with 2 % Giemsa staining solution. The induction of G-bands for differential staining of chromosomes was conducted using 0.25 % solution of trypsin. The processing of study results was performed with Microsoft Excel software package. Results. The investigations in the aberration spectrum detected aneuploid and polyploid cells, breaks and fragments of chromosomes, premature chromo- some disjunction in mitosis and translocation. The total number of aberrant cells in cows with decreased fertil- ity was 14.69 ± 0.56 %, the number of aberrations per one investigated cell was 0.144, which was almost twice reliably (Р < 0.999) exceeding the values of similar features for cows which did not have problems with repro- duction. GTG-banding method was used to detect a new RT 13/23 Robertsonian translocation. Conclusions. The cytogenetic analysis of chromosome set of Ukrainian Red-and-Motley dairy breed cows allows assuming the connection between a decrease in the fertility of cows and chromosomal instability. A routine screening of dairy cows allows both evaluating the karyotype saturation with undesired chromosomal aberrations and using the obtained results to forecast the reproductive ability of an animal in the early age.