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Medicine Sep 2023Hepatocellular carcinoma (HCC) poses a global health challenge. Effective biomarkers are required for early diagnosis to improve survival rates of patients with HCC....
Hepatocellular carcinoma (HCC) poses a global health challenge. Effective biomarkers are required for early diagnosis to improve survival rates of patients with HCC. Spindle and kinetochore-associated complex subunits 1 (SKA1) is essential for proper chromosome segregation in the mitotic cell cycle. Previous studies have shown that overexpression of SKA1 is associated with a poor prognosis in various cancers. The expression, prognostic value, and clinical functions of SKA1 in HCC were evaluated with several bioinformatics web portals. Additionally, we identified target long non-coding RNAs (lncRNAs) and microRNAs by analyzing messenger RNA (mRNA)-miRNA and miRNA-lncRNA interaction data and elucidated the potential competing endogenous RNA (ceRNA) mechanism associated with SKA1. High SKA1 expression was associated with poor prognosis in patients with HCC. Furthermore, multivariate Cox regression analysis revealed that SKA1 expression was an independent prognostic factor for HCC. GO and KEGG analyses showed that SKA1 is related to the cell cycle checkpoints, DNA replication and repair, Rho GTPases signaling, mitotic prometaphase, and kinesins. Gene set enrichment analysis revealed that high levels of SKA1 are associated with cancer-promoting pathways. DNA methylation of SKA1 in HCC tissues was lower than that in normal tissues. Ultimately, the following 9 potential ceRNA-based pathways targeting SKA1 were identified: lncRNA: AC026401.3, Small Nucleolar RNA Host Gene 3 (SNHG3), and AC124798.1-miR-139-5p-SKA1; lncRNA: AC26356.1, Small Nucleolar RNA Host Gene 16 (SNHG16), and FGD5 Antisense RNA 1-miR-22-3p-SKA1; lncRNA: Cytoskeleton Regulator RNA (CYTOR), MIR4435-2 Host Gene, and differentiation antagonizing non-protein coding RNA-miR-125b-5p-SKA1. SKA1 expression levels significantly correlated with immune cell infiltration and immune checkpoint genes in the HCC tissues. SKA1 is a potential prognostic biomarker for HCC. This study provides a meaningful direction for research on SKA1-related mechanisms, which will be beneficial for future research on HCC-related molecular biological therapies and targeted immunotherapy.
Topics: Humans; Carcinoma, Hepatocellular; RNA, Long Noncoding; RNA, Small Nucleolar; Liver Neoplasms; MicroRNAs; Computational Biology; Immunoassay; Chromosomal Proteins, Non-Histone
PubMed: 37746945
DOI: 10.1097/MD.0000000000034826 -
Cellular Signalling Sep 2023c-Src tyrosine kinase plays roles in a wide range of signaling events and its increased activity is frequently observed in a variety of epithelial and non-epithelial...
c-Src tyrosine kinase plays roles in a wide range of signaling events and its increased activity is frequently observed in a variety of epithelial and non-epithelial cancers. v-Src, an oncogene first identified in the Rous sarcoma virus, is an oncogenic version of c-Src and has constitutively active tyrosine kinase activity. We previously showed that v-Src induces Aurora B delocalization, resulting in cytokinesis failure and binucleated cell formation. In the present study, we explored the mechanism underlying v-Src-induced Aurora B delocalization. Treatment with the Eg5 inhibitor (+)-S-trityl-L-cysteine (STLC) arrested cells in a prometaphase-like state with a monopolar spindle; upon further inhibition of cyclin-dependent kinase (CDK1) by RO-3306, cells underwent monopolar cytokinesis with bleb-like protrusions. Aurora B was localized to the protruding furrow region or the polarized plasma membrane 30 min after RO-3306 addition, whereas inducible v-Src expression caused Aurora B delocalization in cells undergoing monopolar cytokinesis. Delocalization was similarly observed in monopolar cytokinesis induced by inhibiting Mps1, instead of CDK1, in the STLC-arrested mitotic cells. Importantly, western blotting analysis and in vitro kinase assay revealed that v-Src decreased the levels of Aurora B autophosphorylation and its kinase activity. Furthermore, like v-Src, treatment with the Aurora B inhibitor ZM447439 also caused Aurora B delocalization at concentrations that partially inhibited Aurora B autophosphorylation. Given that phosphorylation of Aurora B by v-Src was not observed, these results suggest that v-Src causes Aurora B delocalization by indirectly suppressing Aurora B kinase activity.
Topics: Humans; Cytokinesis; Aurora Kinase B; Quinolines; Phosphorylation; Oncogenes; Mitosis; HeLa Cells
PubMed: 37315749
DOI: 10.1016/j.cellsig.2023.110764 -
ELife Jun 2024Cohesin is a multi-subunit protein that plays a pivotal role in holding sister chromatids together during cell division. Sister chromatid cohesion 3 (SCC3), constituents...
Cohesin is a multi-subunit protein that plays a pivotal role in holding sister chromatids together during cell division. Sister chromatid cohesion 3 (SCC3), constituents of cohesin complex, is highly conserved from yeast to mammals. Since the deletion of individual cohesin subunit always causes lethality, it is difficult to dissect its biological function in both mitosis and meiosis. Here, we obtained weak mutants using CRISPR-Cas9 system to explore its function during rice mitosis and meiosis. The weak mutants displayed obvious vegetative defects and complete sterility, underscoring the essential roles of SCC3 in both mitosis and meiosis. SCC3 is localized on chromatin from interphase to prometaphase in mitosis. However, in meiosis, SCC3 acts as an axial element during early prophase I and subsequently situates onto centromeric regions following the disassembly of the synaptonemal complex. The loading of SCC3 onto meiotic chromosomes depends on REC8. shows severe defects in homologous pairing and synapsis. Consequently, SCC3 functions as an axial element that is essential for maintaining homologous chromosome pairing and synapsis during meiosis.
Topics: Chromosome Pairing; Meiosis; Cell Cycle Proteins; Oryza; Chromosomal Proteins, Non-Histone; Cohesins; Mitosis; Synaptonemal Complex; CRISPR-Cas Systems
PubMed: 38864853
DOI: 10.7554/eLife.94180 -
BMC Cancer Aug 2023HeberFERON is a co-formulation of α2b and γ interferons, based on their synergism, which has shown its clinical superiority over individual interferons in basal cell...
BACKGROUND
HeberFERON is a co-formulation of α2b and γ interferons, based on their synergism, which has shown its clinical superiority over individual interferons in basal cell carcinomas. In glioblastoma (GBM), HeberFERON has displayed promising preclinical and clinical results. This led us to design a microarray experiment aimed at identifying the molecular mechanisms involved in the distinctive effect of HeberFERON compared to the individual interferons in U-87MG model.
METHODS
Transcriptional expression profiling including a control (untreated) and three groups receiving α2b-interferon, γ-interferon and HeberFERON was performed using an Illumina HT-12 microarray platform. Unsupervised methods for gene and sample grouping, identification of differentially expressed genes, functional enrichment and network analysis computational biology methods were applied to identify distinctive transcription patterns of HeberFERON. Validation of most representative genes was performed by qPCR. For the cell cycle analysis of cells treated with HeberFERON for 24 h, 48 and 72 h we used flow cytometry.
RESULTS
The three treatments show different behavior based on the gene expression profiles. The enrichment analysis identified several mitotic cell cycle related events, in particular from prometaphase to anaphase, which are exclusively targeted by HeberFERON. The FOXM1 transcription factor network that is involved in several cell cycle phases and is highly expressed in GBMs, is significantly down regulated. Flow cytometry experiments corroborated the action of HeberFERON on the cell cycle in a dose and time dependent manner with a clear cellular arrest as of 24 h post-treatment. Despite the fact that p53 was not down-regulated, several genes involved in its regulatory activity were functionally enriched. Network analysis also revealed a strong relationship of p53 with genes targeted by HeberFERON. We propose a mechanistic model to explain this distinctive action, based on the simultaneous activation of PKR and ATF3, p53 phosphorylation changes, as well as its reduced MDM2 mediated ubiquitination and export from the nucleus to the cytoplasm. PLK1, AURKB, BIRC5 and CCNB1 genes, all regulated by FOXM1, also play central roles in this model. These and other interactions could explain a G2/M arrest and the effect of HeberFERON on the proliferation of U-87MG.
CONCLUSIONS
We proposed molecular mechanisms underlying the distinctive behavior of HeberFERON compared to the treatments with the individual interferons in U-87MG model, where cell cycle related events were highly relevant.
Topics: Humans; Glioblastoma; Apoptosis; Cell Line, Tumor; G2 Phase Cell Cycle Checkpoints; Interferon-alpha; Anaphase; Interferon-gamma; Skin Neoplasms
PubMed: 37644431
DOI: 10.1186/s12885-023-11330-2 -
International Journal of Surgical... Apr 2024The identification of mitotic figures is essential for the diagnosis, grading, and classification of various different tumors. Despite its importance, there is a...
The identification of mitotic figures is essential for the diagnosis, grading, and classification of various different tumors. Despite its importance, there is a paucity of literature reporting the consistency in interpreting mitotic figures among pathologists. This study leverages publicly accessible datasets and social media to recruit an international group of pathologists to score an image database of more than 1000 mitotic figures collectively. Pathologists were instructed to randomly select a digital slide from The Cancer Genome Atlas (TCGA) datasets and annotate 10-20 mitotic figures within a 2 mm area. The first 1010 submitted mitotic figures were used to create an image dataset, with each figure transformed into an individual tile at 40x magnification. The dataset was redistributed to all pathologists to review and determine whether each tile constituted a mitotic figure. Overall pathologists had a median agreement rate of 80.2% (range 42.0%-95.7%). Individual mitotic figure tiles had a median agreement rate of 87.1% and a fair inter-rater agreement across all tiles (kappa = 0.284). Mitotic figures in prometaphase had lower percentage agreement rates compared to other phases of mitosis. This dataset stands as the largest international consensus study for mitotic figures to date and can be utilized as a training set for future studies. The agreement range reflects a spectrum of criteria that pathologists use to decide what constitutes a mitotic figure, which may have potential implications in tumor diagnostics and clinical management.
PubMed: 38627896
DOI: 10.1177/10668969241234321 -
Physical Review. E Sep 2023Variation in the chromosome numbers can arise from the erroneous mitosis or fusion and fission of chromosomes. While the mitotic errors lead to an increase or decrease...
Variation in the chromosome numbers can arise from the erroneous mitosis or fusion and fission of chromosomes. While the mitotic errors lead to an increase or decrease in the overall chromosomal substance in the daughter cells, fission and fusion keep this conserved. Variations in chromosome numbers are assumed to be a crucial driver of speciation. For example, the members of the muntjac species are known to have very different karyotypes with the chromosome numbers varying from 2n=70+3B in the brown brocket deer to 2n=46 in the Chinese muntjac and 2n=6/7 in the Indian muntjac. The chromosomal content in the nucleus of these closely related mammals is roughly the same and various chromosome fusion and fission pathways have been suggested as the evolution process of these karyotypes. Similar trends can also be found in lepidoptera and yeast species which show a wide variation of chromosome numbers. The effect of chromosome number variation on the spindle assembly time and accuracy is still not properly addressed. We computationally investigate the effect of conservation of the total chromosomal substance on the spindle assembly during prometaphase. Our results suggest that chromosomal fusion pathways aid the microtubule-driven search and capture of the kinetochore in cells with monocentric chromosomes. We further report a comparative analysis of the site and percentage of amphitelic captures, dependence on cell shape, and position of the kinetochore in respect to chromosomal volume partitioning.
Topics: Animals; Muntjacs; Deer; Mitosis; Microtubules; Kinetochores
PubMed: 37849183
DOI: 10.1103/PhysRevE.108.034401 -
Research Square Feb 2024The Survivin protein has roles in repairing incorrect microtubule-kinetochore attachments at prometaphase, and the faithful execution of cytokinesis, both as part of the...
The Survivin protein has roles in repairing incorrect microtubule-kinetochore attachments at prometaphase, and the faithful execution of cytokinesis, both as part of the (CPC) (1). In this context, errors frequently lead to aneuploidy, polyploidy and cancer (1). Adding to these well-known roles of this protein, this paper now shows for the first time that Survivin is required for cancer cells to enter mitosis, and that, in its absence, HeLa cells accumulate at early prophase, or prior to reported before (2, 3). This early prophase blockage is demonstrated by the presence of an intact nuclear lamina and low Cdk1 activity (4). Importantly, escaping the arrest induced by Survivin abrogation leads to multiple mitotic defects, or , and eventually cell death. Mechanistically, Cdk1 does not localize at the centrosome in the absence of Survivin pointing at an impairment in signaling through the Cdc25B-Cdk1 axis. In agreement, even though Survivin directly interacts with Cdc25B, both and , in its absence, an inactive cytosolic Cdc25B-Cdk1-Cyclin B1 complex accumulates. This flaw in Cdc25B activation can however be reversed in Survivin-depleted HeLa cell extracts to which the recombinant Survivin protein is added back. Finally, a role for Survivin in the Cdc25B-mediated activation of Cdk1 is confirmed by overriding the early prophase blockage induced in cells lacking Survivin through the expression of a gain-of-function Cdc25B mutant.
PubMed: 38464014
DOI: 10.21203/rs.3.rs-3949429/v1 -
Development (Cambridge, England) Jun 2024The RNA-binding protein cytoplasmic polyadenylation element binding 1 (CPEB1) plays a fundamental role in regulating mRNA translation in oocytes. However, the specifics...
The RNA-binding protein cytoplasmic polyadenylation element binding 1 (CPEB1) plays a fundamental role in regulating mRNA translation in oocytes. However, the specifics of how and which protein kinase cascades modulate CPEB1 activity are still controversial. Using genetic and pharmacological tools, and detailed time courses, we have re-evaluated the relationship between CPEB1 phosphorylation and translation activation during mouse oocyte maturation. We show that both the CDK1/MAPK and AURKA/PLK1 pathways converge on CPEB1 phosphorylation during prometaphase of meiosis I. Only inactivation of the CDK1/MAPK pathway disrupts translation, whereas inactivation of either pathway alone leads to CPEB1 stabilization. However, CPEB1 stabilization induced by inactivation of the AURKA/PLK1 pathway does not affect translation, indicating that destabilization and/or degradation is not linked to translational activation. The accumulation of endogenous CCNB1 protein closely recapitulates the translation data that use an exogenous template. These findings support the overarching hypothesis that the activation of translation during prometaphase in mouse oocytes relies on a CDK1/MAPK-dependent CPEB1 phosphorylation, and that translational activation precedes CPEB1 destabilization.
Topics: Animals; Oocytes; Meiosis; mRNA Cleavage and Polyadenylation Factors; Phosphorylation; Mice; Protein Biosynthesis; Female; CDC2 Protein Kinase; Transcription Factors; Aurora Kinase A; Cyclin B1; Cell Cycle Proteins; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Signal Transduction
PubMed: 38785133
DOI: 10.1242/dev.202712 -
BioRxiv : the Preprint Server For... Jan 2024Genetic screens for recessive alleles induce mutations, make the mutated chromosomes homozygous, and then assay those homozygotes for the phenotype of interest. When...
Genetic screens for recessive alleles induce mutations, make the mutated chromosomes homozygous, and then assay those homozygotes for the phenotype of interest. When screening for genes required for female meiosis, the phenotype of interest has typically been nondisjunction from chromosome segregation errors. As this requires that mutant females be viable and fertile, any mutants that are lethal or sterile when homozygous cannot be recovered by this approach. To overcome these limitations, our lab has screened the VALIUM22 collection produced by the Harvard TRiP Project, which contains RNAi constructs targeting genes known to be expressed in the germline in a vector optimized for germline expression. By driving RNAi with GAL4 under control of a germline-specific promoter ( or , we can test genes that would be lethal if knocked down in all cells, and by examining unfertilized metaphase-arrested mature oocytes, we can identify defects associated with genes whose knockdown results in sterility or causes other errors besides nondisjunction. We screened this collection to identify genes that disrupt either of two phenotypes when knocked down: the ability of meiotic chromosomes to congress to a single mass at the end of prometaphase, and the sequestration of Mps1-GFP to ooplasmic filaments in response to hypoxia. After screening >1450 lines of the collection, we obtained multiple hits for both phenotypes, identified novel meiotic phenotypes for genes that had been previously characterized in other processes, and identified the first phenotypes to be associated with several previously uncharacterized genes.
PubMed: 38293152
DOI: 10.1101/2024.01.12.575435 -
BioRxiv : the Preprint Server For... Jan 2024The RNA-binding protein cytoplasmic polyadenylation element binding 1 (CPEB1) plays a fundamental role in the regulation of mRNA translation in oocytes. However, the...
The RNA-binding protein cytoplasmic polyadenylation element binding 1 (CPEB1) plays a fundamental role in the regulation of mRNA translation in oocytes. However, the nature of protein kinase cascades modulating the activity of CPEB1 is still a matter of controversy. Using genetic and pharmacological tools and detailed time courses, here we have reevaluated the relationship between CPEB1 phosphorylation and the activation of translation during mouse oocyte maturation. We show that both the CDK1/MAPK and AURKA/PLK1 pathways converge on the phosphorylation of CPEB1 during prometaphase. Only inactivation of the CDK1/MAPK pathway disrupts translation, while inactivation of either pathway leads to CPEB1 stabilization. However, stabilization of CPEB1 induced by inactivation of the AURKA/PLK1 does not affect translation, indicating that destabilization/degradation can be dissociated from translational activation. The accumulation of the endogenous CCNB1 protein closely recapitulates the translation data. These findings support the overarching hypothesis that the activation of translation in prometaphase in mouse oocytes relies on a CDK1-dependent CPEB1 phosphorylation, and this translational activation precedes CPEB1 destabilization.
PubMed: 38293116
DOI: 10.1101/2024.01.17.575938