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Replication stress describes endogenous and exogenous challenges to DNA replication in the S-phase. Stress during this critical process causes helicase-polymerase decoupling at replication forks, triggering the S-phase checkpoint, which orchestrates global replication fork stalling and delayed entry into G2. The replication stressor most often used to induce the checkpoint response in yeast is hydroxyurea (HU), a clinically used chemotherapeutic. The primary mechanism of S-phase checkpoint activation by HU has thus far been considered to be a reduction of deoxynucleotide triphosphate synthesis by inhibition of ribonucleotide reductase (RNR), leading to helicase-polymerase decoupling and subsequent activation of the checkpoint, facilitated by the replisome-associated mediator Mrc1. In contrast, we observe that HU causes cell cycle arrest in budding yeast independent of both the Mrc1-mediated replication checkpoint response and the Psk1-Mrc1 oxidative signaling pathway. We demonstrate a direct relationship between HU incubation and reactive oxygen species (ROS) production in yeast and human cells and show that antioxidants restore growth of yeast in HU. We further observe that ROS strongly inhibits the in vitro polymerase activity of replicative polymerases (Pols), Pol α, Pol δ, and Pol ε, causing polymerase complex dissociation and subsequent loss of DNA substrate binding, likely through oxidation of their integral iron-sulfur (Fe-S) clusters. Finally, we present "RNR-deg," a genetically engineered alternative to HU in yeast with greatly increased specificity of RNR inhibition, allowing researchers to achieve fast, nontoxic, and more readily reversible checkpoint activation compared to HU, avoiding harmful ROS generation and associated downstream cellular effects that may confound interpretation of results., Competing Interests: Competing interests statement:The authors declare no competing interest.

Purpose: Loddo et al. (Br J Cancer 100:959-70, 2009) established the prognostic significance of cell cycle markers and "Cell-Cycle Phenotypes" in breast carcinoma. This study aims to 1) identify prognostic cell-cycle markers in sarcoma, and 2) assess the prognostic potential of specific cell-cycle phenotypes in sarcoma., Methods: Tissue samples from 128 soft tissue sarcomas were stained for four cell cycle-specific markers: Mcm2, Geminin, Plk1, and H3S10ph. Only primary soft tissue tumors (liposarcoma, leiomyosarcoma, synovial sarcoma, and undifferentiated pleomorphic sarcoma) were included in the analysis. Any tumor coming from a recurrent or metastatic lesion were excluded from the analysis. Three cell-cycle phenotypes (I, II, III) were derived from marker expression patterns. Prognostic significance was evaluated in a subset of primary soft tissue sarcomas using Cox regression for survival analysis., Results: Compared to phenotype I, the phenotype III tumors had a decreased 5-year overall survival (HR 6.81 [2.36-19.61]; p =  < 0.001), 5-year disease-free survival (HR 1.07 (1.02-1.18); p = 0.004), and 5-year metastasis-free survival (HR 4.34 [1.58-11.93]; p = 0.004). High expression of Plk1 was associated with decreased 5-year overall survival (HR: 4.04 CI [1.21-6.67; p = 0.02) and 5-year metastasis-free survival (HR: 2.91 CI [1.15-7.37]; p = 0.03). Geminin was also found to have a decreased 5-year overall survival (HR:2.84 CI [1.21-6.67]; p = 0.02). No statistical difference in prognostication were noted between phenotypes and the AJCC system., Conclusions: We identified three unique sarcoma cell cycle phenotypes that have prognostic significance. This performs similarly to the AJCC staging system., (© 2024. The Author(s).)

Fetal growth restriction (FGR) is a relatively common complication of pregnancy, and insufficient syncytialization in the placenta may play an important role in the pathogenesis of FGR. However, the mechanism of impaired formation of the syncytiotrophoblast layer in FGR patients requires further exploration. In the present study, we demonstrated that the level of syncytialization was decreased in FGR patient placentas, while the expression of connective tissue growth factor (CTGF) was significantly upregulated. CTGF was found to inhibit trophoblast fusion via regulating cell cycle progress of BeWo cells. Furthermore, we found that CTGF negatively regulates cell cycle arrest in a p21-dependent manner as overexpression of p21 could rescue the impaired syncytialization induced by CTGF-overexpression. Besides, we also identified that CTGF inhibits the expression of p21 through ITGB4/PI3K/AKT signaling pathway. Our study provided a new insight for elucidating the pathogenic mechanism of FGR and a novel idea for the clinical therapy of FGR., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Background: Centrosomal protein of 55 kDa (CEP55) overexpression has been linked to tumor stage, aggressiveness of the tumor, poor prognosis, and metastasis. This study aims to elucidate the action of CEP55 in ovarian cancer (OC) and the regulation by the alpha-ketoglutarate-dependent dioxygenase alkB homolog 5 (ALKBH5)/Forkhead box protein P2 (FOXP2) axis., Methods: Differentially expressed genes in OC were identified using in silico identification, followed by prognostic value assessment. Lentiviral vectors were constructed to downregulate CEP55 in OC cells, and colony formation, EdU, TUNEL, flow cytometry, Transwell assays, and Phalloidin staining were conducted. Transcription factors regulating CEP55 were predicted and verified, and rescue experiments were performed. The effect of ALKBH5-mediated demethylation on FOXP2 mRNA stability and OC cell cycle and EMT were analyzed., Results: High expression of CEP55 in OC was linked to unsatisfactory prognosis of patients. Knockdown of CEP55 repressed proliferation, invasiveness, and epithelial-mesenchymal transition (EMT) while inducing apoptosis and cell cycle arrest in OC cells. FOXP2 bound to the promoter of CEP55 to repress CEP55 transcription. FOXP2 regulated transcriptional repression of CEP55 to impede the malignant progression of OC and inhibit tumor metastasis. ALKBH5-mediated demethylation modification induced mRNA degradation of FOXP2. Knockdown of ALKBH5 induced cell cycle arrest and inhibited EMT in OC cells., Conclusions: ALKBH5 hinders FOXP2-mediated transcriptional repression of CEP55 to promote the malignant progression of OC via cell cycle and EMT., (© 2024. The Author(s).)

Multiple myeloma, which is a clonal plasma cell tumor, derives from a postmitotic lymphoid B-cell lineage and remains untreatable. Group XVI phospholipase A2 (PLA2G16) can either be a tumor suppressor or an oncogene in different types of cancer. This study was intended to explore the role of PLA2G16 in multiple myeloma and to reveal the reaction mechanism. The mRNA and protein expressions of PLA2G16 in human bone marrow stromal cell line HS-5 and multiple myeloma cells were assessed using reverse transcription-quantitative PCR and western blot. The transfection efficacy of sh-PLA2G16 and oe-YAP was examined using reverse transcription-quantitative PCR and western blot. Through cell counting kit-8 assay and 5-ethynyl-2'- deoxyuridine staining, multiple myeloma cell viability and proliferation were detected. Flow cytometry was used to measure cell apoptosis and cell cycle distribution. Oxygen consumption rate, the activities of mitochondrial respiratory chain complexes I-V, and the activity of caspase-3 were estimated with Seahorse XF24 analyzer, oxidative phosphorylation activity assay kit, and caspase-3 assay kit, respectively. Lactate production and glucose consumption were evaluated usingcorresponding assay kits. Western blot was employed to meaure proteins associated with cell cycle, glycolysis, pentose phosphate pathway as well as Hippo/YAP signaling pathway. In this study, PLA2G16 expression was greatly increased in multiple myeloma cells and PLA2G16 silence inhibited cell proliferation, promoted cell apoptosis, facilitated cell cycle arrest, and suppressed the reprogramming of glucose metabolism in multiple myeloma. It was also identified that PLA2G16 depletion inhibited the Hippo/YAP signaling pathway. Further experiments revealed that the overexpression of YAP partially reversed the inhibitory effects of PLA2G16 silence on multiple myeloma cell malignant development and the reprogramming of glucose metabolism. Collectively, PLA2G16 silence impeded multiple myeloma progression and inhibited glucose metabolism reprogramming by blocking the Hippo/YAP signaling pathway., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

NIPP1 is a ubiquitously expressed regulatory subunit of PP1. Its embryonic deletion in keratinocytes causes chronic sterile skin inflammation, epidermal hyperproliferation, and resistance to mutagens in adult mice. To explore the primary effects of NIPP1 deletion, we first examined hair cycle progression of NIPP1 skin knockouts (SKOs). The entry of the first hair cycle in the SKOs was delayed owing to prolonged quiescence of hair follicle stem cells. In contrast, the entry of the second hair cycle in the SKOs was advanced as a result of precocious activation of hair follicle stem cells. The epidermis of SKOs progressively accumulated senescent cells, and this cell-fate switch was accelerated by DNA damage. Primary keratinocytes from SKO neonates and human NIPP1-depleted HaCaT keratinocytes failed to proliferate and showed an increase in the expression of cell cycle inhibitors (p21, p16/Ink4a, and/or p19/Arf) and senescence-associated-secretory-phenotype factors as well as in DNA damage (γH2AX and 53BP1). Our data demonstrate that the primary effect of NIPP1 deletion in keratinocytes is a cell cycle arrest and premature senescence that gradually progresse to chronic senescence and likely contribute to the decreased sensitivity of SKOs to mutagens., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The Nile Tilapia (Oreochromis niloticus) evolved in warm, freshwater rivers, but possesses a broad physiological tolerance to a range of environmental conditions. Due to this hardiness and resilience, this species has been successfully introduced to regions widely outside of its native range. Here, we examine the impact of temperature and salinity variation on this species at the sub-lethal level. Specifically, Nile Tilapia were exposed to two temperatures (21 °C or 14 °C) and three salinities (0, 16, 34 ppt) for 1-h. Given their native habitat, the 21 °C / 0 ppt exposure was considered the control condition. Both cell cycle arrest and apoptosis represent sub-lethal but deleterious responses to environmental stress. Flow cytometry was used to assess the percentage of cells in a given stage of the cell cycle as a metric of cell cycle arrest in spleen and liver. Percentage of apoptotic cells were also quantified. Spleen was more sensitive to cold stress, demonstrating an increase in cells in the G2/M phase after experimental treatment. Liver, however, was more sensitive to salinity stress, with a significant increase in cells stalled in G2/M phase at higher salinities, which is in keeping with the freshwater evolutionary history of the species. A modest apoptotic signal was observed in liver but not in spleen. Together, these findings demonstrate that even short, acute exposures to cold temperatures and elevated salinity can cause sub-lethal damage in a species that is otherwise tolerant of environmental stress at the whole organism level., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: The author is an Associate Editor for Comparative Biochemistry and Physiology but was not involved in the decision to publish this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Pharmacological properties of flavonoids have been reported, with an anticancer role amongst them, however, its mechanisms are not fully elucidated. In this study, the activity of quercetin and chrysin towards MCF-7 and MDA-MB-231 breast cancer cells was investigated. Cellular viability was determined after treatment with the compounds in different concentrations for 24 h. Secondly, cells were treated with fixed concentration of chrysin and different concentrations of quercetin with preincubation for 1 h. Both compounds inhibited cellular proliferation in dose-dependent manner. The association showed improvement in their cytotoxicity, more expressively with preincubation of quercetin. Quercetin and chrysin association induced cell cycle arrest in sub-G0/G1 phase in MDA-MB-231 cells, modified the expression of caspases-3 and -8,-8, inducing late apoptosis cell death. Taken together, our results demonstrate that both flavonoids inhibited cells growth in a dose-dependent manner and the association of quercetin improved chrysin's toxic effect over the cell lines., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

A-MYB (MYBL1) is a transcription factor with a role in meiosis in spermatocytes. The related B-MYB protein is a key oncogene and a master regulator activating late cell cycle genes. To activate genes, B-MYB forms a complex with MuvB and is recruited indirectly to cell cycle genes homology region (CHR) promoter sites of target genes. Activation through the B-MYB-MuvB (MMB) complex is essential for successful mitosis. Here, we discover that A-MYB has a function in transcriptional regulation of the mitotic cell cycle and can substitute for B-MYB. Knockdown experiments in cells not related to spermatogenesis show that B-MYB loss alone merely delays cell cycle progression. Only dual knockdown of B-MYB and A-MYB causes G2/M cell cycle arrest, endoreduplication, and apoptosis. A-MYB can substitute for B-MYB in binding to MuvB. The resulting A-MYB-MuvB complex activates genes through CHR sites. We find that A-MYB activates the same target genes as B-MYB. Many of the corresponding proteins are central regulators of the cell division cycle. In summary, we demonstrate that A-MYB is an activator of the mitotic cell cycle by activating late cell cycle genes., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The measurement of dynamic changes in protein level and localization throughout the cell cycle is of major relevance to studies of cellular processes tightly coordinated with the cycle, such as replication, transcription, DNA repair, and checkpoint control. Currently available methods include biochemical assays of cells in bulk following synchronization, which determine protein levels with poor temporal and no spatial resolution. Taking advantage of genetic engineering and live-cell microscopy, we performed time-lapse imaging of cells expressing fluorescently tagged proteins under the control of their endogenous regulatory elements in order to follow their levels throughout the cell cycle. We effectively discern between cell cycle phases and S subphases based on fluorescence intensity and distribution of co-expressed proliferating cell nuclear antigen (PCNA)-mCherry. This allowed us to precisely determine and compare the levels and distribution of multiple replication-associated factors, including Rap1-interacting factor 1 (RIF1), minichromosome maintenance complex component 6 (MCM6), origin recognition complex subunit 1 (ORC1, and Claspin, with high spatiotemporal resolution in HeLa Kyoto cells. Combining these data with available mass spectrometry-based measurements of protein concentrations reveals the changes in the concentration of these proteins throughout the cell cycle. Our approach provides a practical basis for a detailed interrogation of protein dynamics in the context of the cell cycle.

The present study aimed to assess the effects of simulated microgravity (SMG) on 3T3 cell proliferation and the expression of cell cycle regulators. 3T3 cells were induced to SMG by Gravite® for 8 days, while the control group was treated with 1G condition. The result showed that the SMG condition causes a decrease in proliferative activity in 3T3 cells. In the SMG group, the expression of cell cycle-related proteins was lower than the control on day 3. However, these proteins were upregulated in 3T3 cells of the SMG group on day 5, suggesting that these cells were rescued from the arrest and retrieved a higher proliferation. A down-regulation of cell cycle-related proteins was observed in 3T3 cells of both SMG and control groups on day 7. In conclusion, SMG results in the attenuation of cell proliferation during the initial exposure to SMG, but the cells will adapt to this condition and retrieve normal proliferation by increasing the expression of cell cycle regulators., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: CDK4 is highly expressed and associated with poor prognosis and decreased survival in advanced neuroblastoma (NB). Targeting CDK4 degradation presents a potentially promising therapeutic strategy compared to conventional CDK4 inhibitors. However, the autophagic degradation of the CDK4 protein and its anti-proliferation effect in NB cells has not been mentioned., Results: We identified autophagy as a new pathway for the degradation of CDK4. Firstly, autophagic degradation of CDK4 is critical for NVP-BEZ235-induced G0/G1 arrest, as demonstrated by the overexpression of CDK4, autophagy inhibition, and blockade of autophagy-related genes. Secondly, we present the first evidence that p62 binds to CDK4 and then enters the autophagy-lysosome to degrade CDK4 in a CTSB-dependent manner in NVP-BEZ235 treated NB cells. Similar results regarding the interaction between p62 and CDK4 were observed in the NVP-BEZ235 treated NB xenograft mouse model., Conclusions: Autophagic degradation of CDK4 plays a pivotal role in G0/G1 cell cycle arrest in NB cells treated with NVP-BEZ235.

Recent studies showed an interphase chromosome architecture-a specific coiled nucleosome structure-derived from cryopreserved EM tomograms, and dispersed throughout the nucleus. The images were computationally processed to fill in the missing wedges of data caused by incomplete tomographic tilts. The resulting structures increased z-resolution enabling an extension of the proposed architecture to that of mitotic chromosomes. Here, we provide additional insights into the chromosome architecture that was recently published [M. Elbaum et al., Proc. Natl. Acad. Sci. U.S.A. 119, e2119101119 (2022)]. We build on the defined chromosomes time-dependent structures in an effort to probe their dynamics. Variants of the coiled chromosome structures, possibly further defining specific regions, are discussed. We propose, based on generalized specific uncoiling of mitotic chromosomes in telophase, large-scale reorganization of interphase chromosomes. Chromosome territories, organized as micron-sized small patches, are constructed, satisfying complex volume considerations. Finally, we unveiled the structures of replicated coiled chromosomes, still attached to centromeres, as part of chromosome architecture.

MCPH1 has been identified as the causal gene for primary microcephaly type 1, a neurodevelopmental disorder characterized by reduced brain size and delayed growth. As a multifunction protein, MCPH1 has been reported to repress the expression of TERT and interact with transcriptional regulator E2F1. However, it remains unclear whether MCPH1 regulates brain development through its transcriptional regulation function. This study showed that the knockout of Mcph1 in mice leads to delayed growth as early as the embryo stage E11.5. Transcriptome analysis (RNA-seq) revealed that the deletion of Mcph1 resulted in changes in the expression levels of a limited number of genes. Although the expression of some of E2F1 targets, such as Satb2 and Cdkn1c , was affected, the differentially expressed genes (DEGs) were not significantly enriched as E2F1 target genes. Further investigations showed that primary and immortalized Mcph1 knockout mouse embryonic fibroblasts (MEFs) exhibited cell cycle arrest and cellular senescence phenotype. Interestingly, the upregulation of p19ARF was detected in Mcph1 knockout MEFs, and silencing p19Arf restored the cell cycle and growth arrest to wild-type levels. Our findings suggested it is unlikely that MCPH1 regulates neurodevelopment through E2F1-mediated transcriptional regulation, and p19ARF-dependent cell cycle arrest and cellular senescence may contribute to the developmental abnormalities observed in primary microcephaly.

Alzheimer's disease (AD) is the most common type of dementia. Studies have shown that neuronal cell cycle re-entry is an abnormal change in the early stage of AD, and it is also one of the key reasons for mediating neuronal death. The mechanism of acupuncture and moxibustion in treating AD may be related to the regulation of abnormal cell cycle. In this article, we reviewed the occurrence of AD neuronal cell cycle re-entry and its mediated cell death mechanism, listed the factors affecting neuronal cell cycle re-entry, explored the possible mechanism of acupuncture and moxibustion in preventing and treating AD by regulating AD cell cycle protein, AD neuronal cell cycle re-entry influencing factors (β-amyloid protein and Tau protein phosphorylation level, oxidative stress, brain-derived neurotrophic factor), the multi-target regulation of neuronal cell cycle re-entry, so as to provide reference for the application and research of acupuncture and moxibustion in preventing and treating AD.

Main Conclusion: In the angiosperm root apical meristem, the holoploid DNA content is not directly related to cell cycle time. Instead, ploidy, chromosome number, and taxa emerge as key factors that influence this interaction. It is commonly considered that cell cycle time in the angiosperm root apical meristem is directly related to the holoploid DNA content, and this is one of the manifestations of the nucleotypic effect. In this paper, we significantly expanded the previously reported data on cell cycle time using the thymidine method and the rate-of-cell-production method and investigated the nucleotypic effect in different taxonomic groups, in diploids with varying numbers of chromosomes, and in species of different ploidy levels. The nucleotypic effect was most pronounced in diploids with a chromosome number of less than 17, especially in the orders Asparagales and Liliales. In diploids with 17 and a greater number of chromosomes, and in polyploids, the nucleotypic effect was only evident in these two orders and was practically absent in the rest of angiosperms, which have generally lower the holoploid DNA content and average DNA content per chromosome. Overall, our work demonstrates that in angiosperms the relationship between cell cycle time in the root apical meristem and the holoploid DNA content is complex and not homogeneous and it is not directly related to cell cycle time. Instead, ploidy, chromosome number, and taxa emerge as key factors that influence this interaction., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

From a biochemical viewpoint, the cell cycle is controlled by the phosphorylation of cyclin-dependent kinase (CDK) substrates, and the phosphorylation level is determined by the enzymatic balance between CDK and protein phosphatase 2A (PP2A). However, the conventional techniques for analyzing protein phosphorylation using radioisotopes and antibodies involve many operational steps and take days before obtaining results, making them difficult to apply to high-throughput screening and real-time observations. In this study, we developed luminescent probes with a light intensity that changes depending on its phosphorylation state. We modified the Nano-lantern probe (Renilla luciferase-based Ca 2+ probe) by introducing a CDK-substrate peptide and a phosphopeptide-binding domain into the luciferase. Our initial trial resulted in new probes that could report the CDK/PP2A balance in a purified system. Further modifications of these probes (replacing the phospho-Ser with phospho-Thr and randomly replacing its surrounding amino acids) improved the dynamic range by up to four-fold, making them practical for use in the Xenopus egg extracts system, where many physiological events can be reproduced. Taken together, our new probes enabled the monitoring of the CDK/PP2A balance in real time, and are applicable to high-throughput systems; the new probes thus appear promising for use in substrate and drug screening., (© 2024 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.)

Background and Purpose: Prostate cancer remains a major public health burden worldwide. Polo like kinase 4 (PLK4) has emerged as a promising therapeutic target in prostate cancer due to its key roles in cell cycle regulation and tumour progression. This study aims to develop and characterize the novel curcumin analogue NL13 as a potential therapeutic agent and PLK4 inhibitor against prostate cancer., Experimental Approach: NL13 was synthesized and its effects were evaluated in prostate cancer cells and mouse xenograft models. Kinome screening and molecular modelling identified PLK4 as the primary target. Antiproliferative and proapoptotic mechanisms were explored via cell cycle, apoptosis, gene and protein analyses., Key Results: Compared with curcumin, NL13 exhibited much greater potency in inhibiting PC3 (IC 50 , 3.51 μM vs. 35.45 μM) and DU145 (IC 50 , 2.53 μM vs. 29.35 μM) prostate cancer cells viability and PLK4 kinase activity (2.32 μM vs. 246.88 μM). NL13 induced G2/M cell cycle arrest through CCNB1/CDK1 down-regulation and triggered apoptosis via caspase-9/caspase-3 cleavage. These effects were mediated by PLK4 inhibition, which led to the inactivation of the AKT signalling pathway. In mice, NL13 significantly inhibited tumour growth and modulated molecular markers consistent with in vitro findings, including decreased p-AKT and increased cleaved caspase-9/3., Conclusion and Implications: NL13, a novel PLK4-targeted curcumin analogue, exerts promising anticancer properties against prostate cancer by disrupting the PLK4-AKT-CCNB1/CDK1 and apoptosis pathways. NL13 represents a promising new agent for prostate cancer therapy., (© 2024 British Pharmacological Society.)

Cardiomyocytes in the adult human heart are quiescent and those lost following heart injury are not replaced by proliferating survivors. Considerable effort has been made to understand the mechanisms underlying cardiomyocyte cell cycle exit and re-entry, with view to discovering therapeutics that could stimulate cardiomyocyte proliferation and heart regeneration. The advent of large compound libraries and robotic liquid handling platforms has enabled the screening of thousands of conditions in a single experiment but success of these screens depends on the appropriateness and quality of the model used. Quantification of (human) cardiomyocyte proliferation in high throughput has remained problematic because conventional antibody-based staining is costly, technically challenging and does not discriminate between cardiomyocyte division and failure in karyokinesis or cytokinesis. Live cell imaging has provided alternatives that facilitate high-throughput screening but these have other limitations. Here, we (i) review the cell cycle features of cardiomyocytes, (ii) discuss various cell cycle fluorescent reporter systems, and (iii) speculate on what could improve their predictive value in the context of cardiomyocyte proliferation. Finally, we consider how these new methods can be used in combination with state-of-the-art three-dimensional human cardiac organoid platforms to identify pro-proliferative signalling pathways that could stimulate regeneration of the human heart.

Malassezia is a lipophilic commensal yeast that resides mainly on the mammalian skin and is also found to associate with the internal organs. Dysbiosis of Malassezia is related to several diseases and often escapes detection as it is difficult to culture and maintain. Malassezia cell wall differs from other budding yeasts like S. cerevisiae due to the difference in the lipid content and is difficult to transform. In this study, we present a methodology to stain Malassezia's nucleus and perform cell cycle studies. However, staining presents a challenge due to its exceptionally thick cell wall with high lipid content, hindering conventional methods. Our novel methodology addresses this challenge and enables the staining of the Malassezia nucleus with a low background. This would allow researchers to visualize the overall nuclear health specifically nuclear morphology and analyze DNA content, crucial for cell cycle progression. By employing DNA-specific dyes like DAPI or Hoechst, we can observe the nuclear structure, and using PI we can differentiate cells in distinct cell cycle phases using techniques like flow cytometry. This novel staining methodology unlocks the door for in-depth cell cycle analysis in Malassezia which has challenged us through ages being refractory to genetic manipulations, paving the way for a deeper understanding of this commensal fungus and its potential role in human health., (© 2024. The Author(s).)

Background: Glioblastoma multiforme (GBM) is the most malignant brain tumor, with a poor prognosis and life expectancy of 14-16 months after diagnosis. The standard treatment for GBM consists of surgery, radiotherapy, and chemotherapy with temozolomide. Most patients become resistant to treatment after some time, and the tumor recurs. Therefore, there is a need for new drugs to manage GBM. Eslicarbazepine (ESL) is a well-known antiepileptic drug belonging to the dibenzazepine group with anticancer potentials. In this study, for the first time, we evaluated the potential effects of ESL on C6 cell growth, both in vitro and in vivo, and examined its molecular effects., Methods: To determine the effect of ESL on the c6 cell line, cell viability, proliferation, and migration were evaluated by MTT assay, colony formation, and wound healing assay. Also, apoptosis and cell cycle were examined by flow cytometry, qRT-PCR, and western blotting. In addition, an intracranial model in Wistar rats was used to investigate the effect of ESL in vivo, and the tumor size was measured using both Caliper and MRI., Results: The obtained results are extremely consistent and highly encouraging. C6 cell viability, proliferation, and migration were significantly suppressed in ESL-treated C6 cells (p < 0.001), as determined by cell-based assays. ESL treatment led to significant enhancement of apoptosis (p < 0.01), as determined by flow cytometry, and upregulation of genes involved in cell apoptosis, such as the Bax/Bcl2 ratio at RNA (p < 0.05) and protein levels (5.37-fold). Flow cytometric analysis of ESL-treated cells revealed G2/M phase cell cycle arrest. ESL-treated cells demonstrated 2.49-fold upregulation of p21 alongside, 0.22-fold downregulation of cyclin B1, and 0.34-fold downregulation of cyclin-dependent kinase-1 at the protein level. Administration of ESL (30 mg/kg) to male rats bearing C6 intracranial tumors also suppressed the tumor volume and weight (p < 0.01)., Conclusions: Based on these novel findings, ESL has the potential for further experimental and clinical studies in glioblastoma., (© 2024. The Author(s).)

Cytoskeleton-associated protein 2-like (CKAP2L) is a paralogue of cytoskeleton-associated protein 2 (CKAP2). We characterized the expression pattern, subcellular localization, and microtubule-stabilizing properties of human CKAP2L. The levels of both CKAP2L transcript and protein were cell cycle phase-dependent, peaking during the G 2 /M phase and relatively high in certain human tissues, including testis, intestine, and spleen. CKAP2L protein was detectable in all human cancer cell lines we tested. CKAP2L localized to the mitotic spindle apparatus during mitosis, as reported previously. During interphase, however, CKAP2L localized mainly to the nucleus. Ectopic overexpression of CKAP2L resulted in 'microtubule bundling', and, consequently, an elevated CKAP2L level led to prolonged mitosis. These findings support the mitotic role of CKAP2L during the human cell cycle., (© 2024 The Author(s). FEBS Open Bio published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Breast cancer stands out as one of the most prevalent malignancies worldwide, necessitating a nuanced understanding of its molecular underpinnings for effective treatment. Hormone receptors in breast cancer cells substantially influence treatment strategies, dictating therapeutic approaches in clinical settings, serving as a guide for drug development, and aiming to enhance treatment specificity and efficacy. Natural compounds, such as curcumin, offer a diverse array of chemical structures with promising therapeutic potential. Despite curcumin's benefits, challenges like poor solubility and rapid metabolism have spurred the exploration of analogs. Here, we evaluated the efficacy of the curcumin analog NC2603 to induce cell cycle arrest in MCF-7 breast cancer cells and explored its molecular mechanisms. Our findings reveal potent inhibition of cell viability (IC50 = 5.6 μM) and greater specificity than doxorubicin toward MCF-7 vs. non-cancer HaCaT cells. Transcriptome analysis identified 12,055 modulated genes, most notably upregulation of GADD45A and downregulation of ESR1 , implicating CDKN1A -mediated regulation of proliferation and cell cycle genes. We hypothesize that the curcumin analog by inducing GADD45A expression and repressing ESR1 , triggers the expression of CDKN1A , which in turn downregulates the expression of many important genes of proliferation and the cell cycle. These insights advance our understanding of curcumin analogs' therapeutic potential, highlighting not just their role in treatment, but also the molecular pathways involved in their activity toward breast cancer cells.

In human cells and yeast, an intact "hydrophobic patch" substrate docking site is needed for mitotic cyclin centrosomal localization. A hydrophobic patch mutant (HPM) of the fission yeast mitotic cyclin Cdc13 cannot enter mitosis, but whether this is due to defective centrosomal localization or defective cyclin-substrate docking more widely is unknown. Here, we show that artificially restoring Cdc13-HPM centrosomal localization promotes mitotic entry and increases CDK (cyclin-dependent kinase) substrate phosphorylation at the centrosome and in the cytoplasm. We also show that the S-phase B-cyclin hydrophobic patch is required for centrosomal localization but not for S phase. We propose that the hydrophobic patch is essential for mitosis due to its requirement for the local concentration of cyclin-CDK with CDK substrates and regulators at the centrosome. Our findings emphasize the central importance of the centrosome as a hub coordinating cell-cycle control and explain why the cyclin hydrophobic patch is essential for mitosis., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: This study aims to identify the essential cell cycle-related genes associated with prognosis in breast cancer (BRCA), and to verify the relationship between the central gene and immune infiltration, so as to provide detailed and comprehensive information for the treatment of BRCA., Materials and Methods: Gene expression profiles (GSE10780, GSE21422, GSE61304) and the Cancer Genome Atlas (TCGA) BRCA data were used to identify differentially expressed genes (DEGs) and further functional enrichment analysis. STRING and Cytoscape were employed for the protein-protein interaction (PPI) network construction. TPX2 was viewed as the crucial prognostic gene by the Survival and Cox analysis. Furthermore, the connection between TPX2 expression and immune infiltrating cells and immune checkpoints in BRCA was also performed by the TIMER online database and R software., Results: A total of 18 cell cycle-related DEGs were identified in this study. Subsequently, an intersection analysis based on TCGA-BRCA prognostic genes and the above DEGs identified three genes (TPX2, UBE2C, CCNE2) as crucial prognostic candidate biomarkers. Moreover, we also demonstrated that TPX2 is closely associated with immune infiltration in BRCA and a positive relation between TPX2 and PD-L1 expression was firstly detected., Conclusions: These results revealed that TPX2 is a potential prognostic biomarker and closely correlated with immune infiltration in BRCA, which could provide powerful and efficient strategies for breast cancer immunotherapy.

The cell cycle is a fundamental process essential for cell proliferation, differentiation and development. It consists of four major phases: G1, S, G2 and M. These phases collectively drive the reproductive cycle and are meticulously regulated by various proteins that play crucial roles in both the prevention and progression of cancer. Traditional methods for studying these functions, such as flow cytometry, require a substantial number of cells to ensure accuracy. In this study, we have developed a user-friendly immunofluorescence-based method for identifying cell cycle stages, providing single-cell resolution and precise identification of G1, early/mid S, late S, early/mid G2, late G2, and each sub-stage of the M phase using fluorescence microscopy called ImmunoCellCycle-ID. This method provides high-precision cell cycle identification and can serve as an alternative to, or in combination with, traditional flow cytometry to dissect detailed sub-stages of the cell cycle in a variety of cell lines., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Ataxia telangiectasia and Rad3-related protein (ATR) is a key DNA damage response protein that facilitates DNA damage repair and regulates cell cycle progression. As such, ATR is an important component of the cellular response to radiation, particularly in cancer cells, which show altered DNA damage response and aberrant cell cycle checkpoints. Therefore, ATR's pharmacological inhibition could be an effective radiosensitization strategy to improve radiotherapy. We assessed the ability of an ATR inhibitor, AZD6738, to sensitize cancer cell lines of various histologic types to photon and proton radiotherapy. We found that radiosensitization took place through persistent DNA damage and abrogated G2 cell cycle arrest. We also found that AZD6738 increased the number of micronuclei after exposure to radiotherapy. We found that combining radiation with AZD6738 led to tumor growth delay and prolonged survival relative to radiation alone in a breast cancer model. Combining AZD6738 with photons or protons also led to increased macrophage infiltration at the tumor microenvironment. These results provide a rationale for further investigation of ATR inhibition in combination with radiotherapy and with other agents such as immune checkpoint blockade.

Non-excitable cells express sodium voltage-gated channel alpha subunit 1 gene and protein (known as SCN1A and NaV1.1, respectively); however, the functions of NaV1.1 are unclear. In this study, we investigated the role of SCN1A and NaV1.1 in human mesenchymal stem cells (MSCs). We found that SCN1A was expressed in MSCs, and abundant expression of NaV1.1 was observed in the endoplasmic reticulum; however, this expression was not found to be related to Na+ currents. SCN1A-silencing reduced MSC proliferation and delayed the cell cycle in the S phase. SCN1A silencing also suppressed the protein levels of CDK2 and AKT (herein referring to total AKT), despite similar mRNA expression, and inhibited AKT phosphorylation in MSCs. A cycloheximide-chase assay showed that SCN1A-silencing induced CDK2 but not AKT protein degradation in MSCs. A proteolysis inhibition assay using epoxomicin, bafilomycin A1 and NH4Cl revealed that both the ubiquitin-proteasome system and the autophagy and endo-lysosome system were irrelevant to CDK2 and AKT protein reduction in SCN1A-silenced MSCs. The AKT inhibitor LY294002 did not affect the degradation and nuclear localization of CDK2 in MSCs. Likewise, the AKT activator SC79 did not attenuate the SCN1A-silencing effects on CDK2 in MSCs. These results suggest that NaV1.1 contributes to the cell cycle of MSCs by regulating the post-translational control of AKT and CDK2., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Methods that predict fate potential or degree of differentiation from transcriptomic data have identified rare progenitor populations and uncovered developmental regulatory mechanisms. However, some state-of-the-art methods are too computationally burdensome for emerging large-scale data and all methods make inaccurate predictions in certain biological systems. We developed a method in R (stemFinder) that predicts single cell differentiation time based on heterogeneity in cell cycle gene expression. Our method is computationally tractable and is as good as or superior to competitors. As part of our benchmarking, we implemented four different performance metrics to assist potential users in selecting the tool that is most apt for their application. Finally, we explore the relationship between differentiation time and cell fate potential by analyzing a lineage tracing dataset with clonally labelled hematopoietic cells, revealing that metrics of differentiation time are correlated with the number of downstream lineages., (© The Author(s) 2024. Published by Oxford University Press.)

Kaposi's sarcoma-associated herpesvirus (KSHV) is a double-stranded DNA virus that encodes numerous cellular homologs, including cyclin D, G protein-coupled protein, interleukin-6, and macrophage inflammatory proteins 1 and 2. KSHV vCyclin encoded by ORF72, is the homolog of cellular cyclinD2. KSHV vCyclin can regulate virus replication and cell proliferation by constitutively activating cellular cyclin-dependent kinase 6 (CDK6). However, the regulatory mechanism of KSHV vCyclin has not been fully elucidated. In the present study, we identified a host protein named protein arginine methyltransferase 5 (PRMT5) that interacts with KSHV vCyclin. We further demonstrated that PRMT5 is upregulated by latency-associated nuclear antigen (LANA) through transcriptional activation. Remarkably, knockdown or pharmaceutical inhibition (using EPZ015666) of PRMT5 inhibited the cell cycle progression and cell proliferation of KSHV latently infected tumor cells. Mechanistically, PRMT5 methylates vCyclin symmetrically at arginine 128 and stabilizes vCyclin in a methyltransferase activity-dependent manner. We also show that the methylation of vCyclin by PRMT5 positively regulates the phosphorylate retinoblastoma protein (pRB) pathway. Taken together, our findings reveal an important regulatory effect of PRMT5 on vCyclin that facilitates cell cycle progression and proliferation, which provides a potential therapeutic target for KSHV-associated malignancies., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Niu et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Breast cancer (BRCA) remains to be among the main causes of cancer-associated mortality in women globally. HGH1 homolog (HGH1) has been reported to be associated with tumor immunity. However, the function of HGH1 in BRCA remains unclear. Therefore, the present study examined the potential role of HGH1 in BRCA., Methods: The Cancer Genome Atlas (TCGA) databases and Gene Expression Omnibus (GEO) were used to obtain RNA-seq data for BRCA. A protein localization of HGH1 was determined by using the Human Protein Atlas (HPA), and immunohistochemistry (IHC) staining revealed an upregulation in the expression of HGH1 in clinical BRCA tissues. Xenograft mice were used to test tumor growth and HGH1 expression in breast cancer cells. The protein interaction information of HGH1 was analyzed using the GeneMANIA website. Based on univariate Cox regression and Kaplan-Meier methods, we evaluated the role of HGH1 in BRCA prognosis. HGH1-related differentially expressed genes were analyzed using GO, KEGG, and GSEA. We also examined the relationship between HGH1 expression, immune checkpoints, and immune infiltration. CCK-8, EdU, and colony formation assays were used to measure cell proliferation, and western blot analysis was used to evaluate HGH1's role in BRCA., Results: IHC results showed that the expression of HGH1 was significantly upregulated in BRCA tissues compared to normal tissues. High levels of HGH1 expression was associated with worse clinical features and a worse prognosis. HGH1 expression was an independent predictor of BRCA outcomes in both univariate and multivariate analyses. Functionally, western blot analysis showed that HGH1 is implicated in cell cycle. As well, knocking down HGH1 significantly reduced BRCA cells' proliferative abilities. Crucially, HGH1 expression levels were positively correlated with Th2 cell infiltration and negatively correlated with Tcm cell infiltration., Conclusion: Biomarkers such as HGH1 can reliably predict prognosis in BRCA patients., (© 2024. The Author(s).)

The cell division cycle-associated protein (CDCA) family is important in regulating cell division. High CDCA expression is significantly linked to tumor development. This review summarizes clinical and basic studies on CDCAs conducted in recent decades. Furthermore, it systematically introduces the molecular expression and function, key mechanisms, cell cycle regulation, and roles of CDCAs in tumor development, cell proliferation, drug resistance, invasion, and metastasis. Additionally, it presents the latest research on tumor diagnosis, prognosis, and treatment targeting CDCAs. These findings are pivotal for further in-depth studies on the role of CDCAs in promoting tumor development and provide theoretical support for their application as new anti-tumor targets., Competing Interests: Declaration of competing interest The authors do not have any possible conflicts of interest., (Copyright © 2024. Published by Elsevier B.V.)

Induced oncoproteins degradation provides an attractive anti-cancer modality. Activation of anaphase-promoting complex (APC/C CDH1 ) prevents cell-cycle entry by targeting crucial mitotic proteins for degradation. Phosphorylation of its co-activator CDH1 modulates the E3 ligase activity, but little is known about its regulation after phosphorylation and how to effectively harness APC/C CDH1 activity to treat cancer. Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (PIN1)-catalyzed phosphorylation-dependent cis-trans prolyl isomerization drives tumor malignancy. However, the mechanisms controlling its protein turnover remain elusive. Through proteomic screens and structural characterizations, we identify a reciprocal antagonism of PIN1-APC/C CDH1 mediated by domain-oriented phosphorylation-dependent dual interactions as a fundamental mechanism governing mitotic protein stability and cell-cycle entry. Remarkably, combined PIN1 and cyclin-dependent protein kinases (CDKs) inhibition creates a positive feedback loop of PIN1 inhibition and APC/C CDH1 activation to irreversibly degrade PIN1 and other crucial mitotic proteins, which force permanent cell-cycle exit and trigger anti-tumor immunity, translating into synergistic efficacy against triple-negative breast cancer., (© 2024. The Author(s).)

Inherently identical cells exhibit significant phenotypic variation. It can be essential for many biological processes and is known to arise from stochastic, 'noisy', gene expression that is determined by intrinsic and extrinsic components. It is now obvious that the noise varies as a function of inducer concentration. However, its fluctuation over the cell cycle is limited. Applying dual colour fluorescence protein reporter system, Cyan Fluorescent Protein (CFP) and Yellow fluorescent protein (YFP) tagged multi-copy plasmids, we determine variation of the noise components over the phases in lac promoter induced by Isopropyl β-D-1-thiogalactopyranoside (IPTG) and in presence of additional Magnesium, Mg 2+ ion. We, also, estimate the how such system deviates from observations of single-copy plasmid. Found 25 % difference between multi-copy system and single-copy system clarifies that observed noise is considerable and estimates population behaviour during the cell cycle. We show that total variation in cells induced with IPTG is determined by higher extrinsic than intrinsic noise. It increases from Lag to Exponential phase and decreases from Retardation to Stationary phase. By observing slow and fast dividing cells, we show that 5 mM Mg 2+ increases population homogeneity compared to 2.5 mM Mg 2+ in the environment. The experimental data obtained using dual colour fluorescence protein reporter system demonstrates that protein expression noise, depending on intra cellular ionic concentration, is tightly controlled by phase of the cell., Competing Interests: Declaration of competing interest The authors declare no competing financial interests., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

The endoplasmic reticulum (ER) is a single-copy organelle that cannot be generated de novo, suggesting coordination between the mechanisms overseeing ER integrity and those controlling the cell cycle to maintain organelle inheritance. The Unfolded Protein Response (UPR) is a conserved signaling network that regulates ER homeostasis. Here, we show that pharmacological and genetic inhibition of the UPR sensors IRE1, ATF6, and PERK in unstressed cells delays the cell cycle, with PERK inhibition showing the most penetrant effect, which was associated with a slowdown of the G1-to-S/G2 transition. Treatment with the small molecule ISRIB to bypass the effects of PERK-dependent phosphorylation of the translation initiation factor eIF2α had no such effect, suggesting that cell cycle timing depends on PERKs kinase activity but is independent of eIF2α phosphorylation. Using complementary light and electron microscopy and flow cytometry-based analyses, we also demonstrate that the ER enlarges before mitosis. Together, our results suggest coordination between UPR signaling and the cell cycle to maintain ER physiology during cell division.

Prostate cancer is the most frequent solid tumor in terms of incidence and ranks second only to lung cancer in terms of cancer mortality among men. It has a considerably high mortality rate; around 375,000 deaths occurred worldwide in 2020. In 2024, the American Cancer Society estimated that the number of new prostate cancer cases will be around 299,010 cases, and the estimated deaths will be around 32,250 deaths only in the USA. Cell cycle dysregulation is inevitable in cancer etiology and is targeted by various therapies in cancer treatment. MicroRNAs (miRNAs) are small, endogenous, non-coding regulatory molecules involved in both normal and abnormal cellular events. One of the cellular processes regulated by miRNAs is the cell cycle. Although there are some exceptions, tumor suppressor miRNAs could potentially arrest the cell cycle by downregulating several molecular machineries involved in catalyzing the cell cycle progression. In contrast, oncogenic miRNAs (oncomirs) help the cell cycle to progress by targeting various regulatory proteins such as retinoblastoma (Rb) or cell cycle inhibitors such as p21 or p27, and hence may contribute to prostate cancer progression; however, this is not always the case. In this review, we emphasize how a dysregulated miRNA expression profile is linked to an abnormal cell cycle progression in prostate cancer, which subsequently paves the way to a new therapeutic option for prostate cancer., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)