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Depletion interactions are thought to significantly contribute to the organization of intracellular structures in the crowded cytosol. The strength of depletion interactions depends on physical parameters such as the depletant number density and the depletant size ratio. Cells are known to dynamically regulate these two parameters by varying the copy number of proteins of a wide distribution of sizes. However, mammalian cells are also known to keep the total protein mass density remarkably constant, to within 0.5% throughout the cell cycle. We thus ask how the strength of depletion interactions varies when the total depletant mass is held fixed, a.k.a. fixed-mass depletion. We answer this question via scaling arguments, as well as by studying depletion effects on networks of reconstituted semiflexible actin in silico and in vitro. We examine the maximum strength of the depletion interaction potential U∗ as a function of q, the size ratio between the depletant and the matter being depleted. We uncover a scaling relation U∗ ∼ qζ for two cases: fixed volume fraction φ and fixed mass density ρ. For fixed volume fraction, we report ζ < 0. For the fixed mass density case, we report ζ > 0, which suggests that the depletion interaction strength increases as the depletant size ratio is increased. To test this prediction, we prepared our filament networks at fixed mass concentrations with varying sizes of the depletant molecule poly(ethylene glycol) (PEG). We characterize the depletion interaction strength in our simulations via the mesh size. In experiments, we observe two distinct actin network morphologies, which we call weakly bundled and strongly bundled. We identify a mass concentration where different PEG depletant sizes lead to weakly bundled or strongly bundled morphologies. For these conditions, we find that the mesh size and intra-bundle spacing between filaments across the different morphologies do not show significant differences, while the dynamic light scattering relaxation time and storage modulus between the two states do show significant differences. Our results demonstrate the ability to tune actin network morphology and mechanics by controlling depletant size and give insights into depletion interaction mechanisms under the fixed-depletant-mass constraint relevant to living cells. [ABSTRACT FROM AUTHOR]

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.

Key HIV cure strategies involve reversing immune dysfunction and limiting the proliferation of infected T cells. We evaluate the safety of sirolimus, a mammalian target of rapamycin (mTOR) inhibitor, in people with HIV (PWH) and study the impact of sirolimus on HIV-1 reservoir size and HIV-1-specific immunity in a single-arm study of 20 weeks of treatment in PWH on antiretroviral therapy (ART). Sirolimus treatment does not impact HIV-1-specific CD8 T cell responses but leads to a significant decrease in CD4+ T cell-associated HIV-1 DNA levels at 20 weeks of therapy in the primary efficacy population (n = 16; 31% decline, p = 0.008). This decline persists for at least 12 weeks following cessation of the study drug. Sirolimus treatment also leads to a significant reduction in CD4+ T cell cycling and PD-1 expression on CD8+ lymphocytes. These data suggest that homeostatic proliferation of infected cells, an important mechanism for HIV persistence, is an intriguing therapeutic target.

Active loop extrusion—the process of formation of dynamically growing chromatin loops due to the motor activity of DNA–binding protein complexes—is a firmly established mechanism responsible for chromatin spatial organization at different stages of a cell cycle in eukaryotes and bacteria. The theoretical insight into the effect of loop extrusion on the experimentally measured statistics of chromatin conformation can be gained with an appropriately chosen polymer model. Here, we consider the simplest analytically solvable model of an interphase chromosome, which is treated as an ideal chain with disorder of sufficiently sparse random loops whose conformations are sampled from the equilibrium ensemble. This framework allows us to arrive at the closed-form analytical expression for the mean-squared distance between pairs of genomic loci, which is valid beyond the one-loop approximation in diagrammatic representation. In addition, we analyze the loop-induced deviation of chain conformations from the Gaussian statistics by calculating kurtosis of probability density of the pairwise separation vector. The presented results suggest the possible ways of estimating the characteristics of the loop extrusion process based on the experimental data on the scale-dependent statistics of intra-chromosomal pair-wise distances. [ABSTRACT FROM AUTHOR]

We studied the D3-type cyclin function during gynoecium development in Arabidopsis and how they are related to the hormone cytokinin and the transcription factor SPATULA. Growth throughout the life of plants is sustained by cell division and differentiation processes in meristematic tissues. In Arabidopsis, gynoecium development implies a multiphasic process where the tissues required for pollination, fertilization, and seed development form. The Carpel Margin Meristem (CMM) is a mass of undifferentiated cells that gives rise to the gynoecium internal tissues, such as septum, ovules, placenta, funiculus, transmitting tract, style, and stigma. Different genetic and hormonal factors, including cytokinin, control the CMM function. Cytokinin regulates the cell cycle transitions through the activation of cell cycle regulators as cyclin genes. D3-type cyclins are expressed in proliferative tissues, favoring the mitotic cell cycle over the endoreduplication. Though the role of cytokinin in CMM and gynoecium development is highly studied, its specific role in regulating the cell cycle in this tissue remains unclear. Additionally, despite extensive research on the relationship between CYCD3 genes and cytokinin, the regulatory mechanism that connects them remains elusive. Here, we found that D3-type cyclins are expressed in proliferative medial and lateral tissues. Conversely, the depletion of the three CYCD3 genes showed that they are not essential for gynoecium development. However, the addition of exogenous cytokinin showed that they could control the division/differentiation balance in gynoecium internal tissues and outgrowths. Finally, we found that SPATULA can be a mechanistic link between cytokinin and the D3-type cyclins. The data suggest that the role of D3-type cyclins in gynoecium development is related to the cytokinin response, and they might be activated by the transcription factor SPATULA.

Mutations in human nonsense-mediated mRNA decay (NMD) factors are enriched in neurodevelopmental disorders. We show that deletion of key NMD factor Upf2 in mouse embryonic neural progenitor cells causes perinatal microcephaly but deletion in immature neurons does not, indicating NMD's critical roles in progenitors. Upf2 knockout (KO) prolongs the cell cycle of radial glia progenitor cells, promotes their transition into intermediate progenitors, and leads to reduced upper-layer neurons. CRISPRi screening identified Trp53 knockdown rescuing Upf2KO progenitors without globally reversing NMD inhibition, implying marginal contributions of most NMD targets to the cell cycle defect. Integrated functional genomics shows that NMD degrades selective TRP53 downstream targets, including Cdkn1a, which, without NMD suppression, slow the cell cycle. Trp53KO restores the progenitor cell pool and rescues the microcephaly of Upf2KO mice. Therefore, one physiological role of NMD in the developing brain is to degrade selective TRP53 targets to control progenitor cell cycle and brain size.

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.

Entry into the cell cycle in late G1 phase occurs only when sufficient growth has occurred. In budding yeast, a cyclin called Cln3 is thought to link cell-cycle entry to cell growth. Cln3 accumulates during growth in early G1 phase and eventually helps trigger expression of late G1 phase cyclins that drive cell-cycle entry. All current models for cell-cycle entry assume that expression of late G1 phase cyclins is initiated at the transcriptional level. Current models also assume that the sole function of Cln3 in cell-cycle entry is to promote transcription of late G1 phase cyclins, and that Cln3 works solely in G1 phase. Here, we show that cell cycle-dependent expression of the late G1 phase cyclin Cln2 does not require any functions of the CLN2 promoter. Moreover, Cln3 can influence accumulation of Cln2 protein via posttranscriptional mechanisms. Finally, we show that Cln3 has functions in mitosis that strongly influence cell size. Together, these discoveries reveal the existence of surprising new mechanisms that challenge current models for control of cell-cycle entry and cell size.

BACKGROUND: A traditional view is that stem cells (SCs) divide slowly. Meanwhile, both embryonic and pluripotent SCs display a shorter cell cycle duration (CCD) in comparison to more committed progenitors (CPs). METHODS: We examined the in vitro proliferation and cycling behavior of somatic adult human cells using live cell imaging of passage zero keratinocytes and single-cell RNA sequencing. RESULTS: We found two populations of keratinocytes: those with short CCD and protracted near exponential growth, and those with long CCD and terminal differentiation. Applying the ergodic principle, the comparative numbers of cycling cells in S phase in an enriched population of SCs confirmed a shorter CCD than CPs. Further, analysis of single-cell RNA sequencing of cycling adult human keratinocyte SCs and CPs indicated a shortening of both G1 and G2M phases in the SC. CONCLUSIONS: Contrary to the pervasive paradigm, SCs progress through cell cycle more quickly than more differentiated dividing CPs. Thus, somatic human adult keratinocyte SCs may divide infrequently, but divide rapidly when they divide. Additionally, it was found that SC-like proliferation persisted in vitro.

The road from transcription to protein synthesis is paved with many obstacles, allowing for several modes of post-transcriptional regulation of gene expression. A fundamental player in mRNA biology is DDX3X, an RNA binding protein that canonically regulates mRNA translation. By monitoring dynamics of mRNA abundance and translation following DDX3X depletion, we observe stabilization of translationally suppressed mRNAs. We use interpretable statistical learning models to uncover GC content in the coding sequence as the major feature underlying RNA stabilization. This result corroborates GC content-related mRNA regulation detectable in other studies, including hundreds of ENCODE datasets and recent work focusing on mRNA dynamics in the cell cycle. We provide further evidence for mRNA stabilization by detailed analysis of RNA-seq profiles in hundreds of samples, including a Ddx3x conditional knockout mouse model exhibiting cell cycle and neurogenesis defects. Our study identifies a ubiquitous feature underlying mRNA regulation and highlights the importance of quantifying multiple steps of the gene expression cascade, where RNA abundance and protein production are often uncoupled.

UNLABELLED: 11q13 amplification is a frequent event in human cancer and in particular in squamous cell carcinomas (SCC). Despite almost invariably spanning 10 genes, it is unclear which genetic components of the amplicon are the key driver events in SCC. A combination of computational, in vitro, ex vivo, and in vivo models leveraging efficient primary human keratinocyte genome editing by Cas9-RNP electroporation, identified ORAOV1, CCND1, and MIR548K as the critical drivers of the amplicon in head and neck SCC. CCND1 amplification drives the cell cycle in a CDK4/6/RB1-independent fashion and may confer a novel dependency on RRM2. MIR548K contributes to epithelial-mesenchymal transition. Finally, we identify ORAOV1 as an oncogene that acts likely via its ability to modulate reactive oxygen species. Thus, the 11q13 amplicon drives SCC through at least three independent genetic elements and suggests therapeutic targets for this morbid and lethal disease. IMPLICATIONS: This work demonstrates novel mechanisms and ways to target these mechanisms underlying the most common amplification in squamous cell carcinoma, one of the most prevalent and deadly forms of human cancer.

Histone deacetylases (HDACs) play a crucial role in transcriptional regulation and are implicated in various diseases, including cancer. They are involved in histone tail deacetylation and canonically linked to transcriptional repression. Previous studies suggested that HDAC recruitment to cell-cycle gene promoters via the retinoblastoma (RB) protein or the DREAM complex through SIN3B is essential for G1/S and G2/M gene repression during cell-cycle arrest and exit. Here we investigate the interplay among DREAM, RB, SIN3 proteins, and HDACs in the context of cell-cycle gene repression. Knockout of SIN3B does not globally derepress cell-cycle genes in non-proliferating HCT116 and C2C12 cells. Loss of SIN3A/B moderately upregulates several cell-cycle genes in HCT116 cells but does so independently of DREAM/RB. HDAC inhibition does not induce general upregulation of RB/DREAM target genes in arrested transformed or non-transformed cells. Our findings suggest that E2F:RB and DREAM complexes can repress cell-cycle genes without relying on HDAC activity.

The preparation of cellular metabolomics samples and how to achieve comprehensive coverage of different polar metabolites in cell samples in the analysis pose a challenge for cellular metabolomics. In this study, we optimized a metabolomics protocol based on ultra-high-performance liquid chromatography high-resolution mass spectrometry (UPLC/HRMS) for the extraction and detection of metabolites in A549 cells and exploration of the intervention effect of Qi-Yu-San-Long decoction (QYSLD) on A549 cells. The results indicate that the lowest level of ATP leakage was observed when A549 cells were quenched under liquid nitrogen. MeOH/chloroform/H2O (1:2:1) extraction yielded more chromatographic peaks and excellent reproducibility, and the relative extraction efficiency of most target metabolites was also high. And we optimized the chromatographic separation conditions in both HILIC and RPLC modes, enabling comprehensive detection and analysis of metabolites with varying polarities. Then, we applied the optimized method to UPLC-Q-TOF/MS-based metabolomics of A549 cells to study the mechanism of QYSLD intervention in non-small cell lung cancer (NSCLC). The CCK-8, EdU staining, and cell cycle assay showed that QYSLD inhibited the proliferation of A549 cells by interfering with the cell cycle and blocking them in the G1 phase. A total of 36 differential metabolites associated with the antitumor effects of QYSLD on NSCLC were identified, mainly involving nicotinate and nicotinamide metabolism, sphingolipid metabolism, and glycerophospholipid metabolism. And western blotting confirmed that the change in 1-methylnicotinamide levels after QYSLD intervention was associated with the inhibition of nicotinamide N-methyltransferase expression in A549 cells. [ABSTRACT FROM AUTHOR]

Chinese hamster ovary (CHO) cells represent the most preferential host cell system for therapeutic monoclonal antibody (mAb) production. Enhancing mAb production in CHO cells can be achieved by adding chemical compounds that regulate the cell cycle and cell survival pathways. This study investigated the impact of ectoine supplementation on mAb production in CHO cells. The results showed that adding ectoine at a concentration of 100 mM on the 3rd day of cultivation improved mAb production by improving cell viability and extending the culture duration. RNA sequencing analysis revealed differentially expressed genes associated with cell cycle regulation, cell proliferation, and cellular homeostasis, in particular promotion of cell cycle arrest, which was then confirmed by flow cytometry analysis. Ectoine-treated CHO cells exhibited an increase in the number of cells in the G0/G1 phase. In addition, the cell diameter was also increased. These findings support the hypothesis that ectoine enhances mAb production in CHO cells through mechanisms involving cell cycle arrest and cellular homeostasis. Overall, this study highlights the potential of ectoine as a promising supplementation strategy to enhance mAb production not only in CHO cells but also in other cell lines. [Display omitted] • Ectoine improved mAb production in CHO cells. • Ectoine improved cell viability and extended the culture duration of CHO cells. • Ectoine increased the number of CHO cells in the G0/G1 phase. • Enhanced mAb production involved cell cycle arrest and cellular homeostasis. • Ectoine is a promising supplement to enhance mAb production in cell lines. [ABSTRACT FROM AUTHOR]

Three novel complexes of deprotonated diflunisal (dif) with neocuproine (neo) were synthesized and characterized via elemental, spectral (UV-vis, FTIR, fluorescence, and mass spectrometry), and single-crystal X-ray diffraction analyses. Although the compounds shared a similar composition of [MCl(dif)(neo)], where M represents Zn(II) (1), Co(II) (2) and Cu(II) (3), only 1 and 2 were isostructural, while 3 differed in both the molecular and supramolecular structures. In all three complex molecules, the central atom is coordinated by two nitrogen atoms of neo in a bidentate chelate mode, and one chlorido ligand and dif is bonded in either a monodentate mode via one oxygen atom of the carboxylate in 1 and 2 or in a bidentate chelate mode via both carboxylate oxygen atoms in 3. All three compounds demonstrated remarkable antiproliferative activity against human prostate (PC-3), colon (HCT116) and breast (MDA-MB-468) cancer cell lines with IC50 values in the nanomolar range, with the lowest values observed in the case of PC-3 and MDA-MB-468 with 2 (20.0 nM) and 3 (31.1 nM), respectively. Moreover, complex 2, as the most active, was further investigated for its potential to induce perturbations in the cell cycle of PC-3 cells. The results indicated an induction of caspase-independent apoptosis. The interaction of the complexes with genomic DNA isolated from the respective cancer cell lines was evaluated for the intercalative mode, with binding strength correlated with the antiproliferative activity against PC-3 and MDA-MB-468 cancer cell lines. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer death worldwide, and classifying the developmental stages of HCC can help with early prognosis and treatment. This study aimed to investigate diagnostic and prognostic molecular signatures underlying the progression of HCC, including tumor initiation and growth, and to classify its developmental stages based on gene expression levels. We integrated data from two cancer systems, including 78 patients with Edmondson-Steiner (ES) grade and 417 patients with TNM stage cancer. Functional profiling was performed using identified signatures. Using a multinomial logistic regression model (MLR), we classified controls, early-stage HCC, and advanced-stage HCC. The model was validated in three independent cohorts comprising 45 patients (neoplastic stage), 394 patients (ES grade), and 466 patients (TNM stage). Multivariate Cox regression was employed for HCC prognosis prediction. We identified 35 genes with gradual upregulation or downregulation in both ES grade and TNM stage patients during HCC progression. These genes are involved in cell division, chromosome segregation, and mitotic cytokinesis, promoting tumor cell proliferation through the mitotic cell cycle. The MLR model accurately differentiated controls, early-stage HCC, and advanced-stage HCC across multiple cancer systems, which was further validated in various independent cohorts. Survival analysis revealed a subset of five genes from TNM stage (HR: 3.27, p < 0.0001) and three genes from ES grade (HR: 7.56, p < 0.0001) that showed significant association with HCC prognosis. The identified molecular signature not only initiates tumorigenesis but also promotes HCC development. It has the potential to improve clinical diagnosis, prognosis, and therapeutic interventions for HCC. This study enhances our understanding of HCC progression and provides valuable insights for precision medicine approaches. • 35 genes gradually regulate in HCC progression in both cancer stage and grade. • These molecular signatures enhance cell proliferation via impact on the mitotic cycle. • Using MLR model, 10 genes accurately classify HCC stages across cancer systems. • Statistically significant subsets of genes emerge as prognostic markers for HCC. [ABSTRACT FROM AUTHOR]

X-irradiation of blood products is an alternative for gamma-ray to prevent post-transfusion GvHD. However, commercial X-irradiators are not widely available while little is known about their safety and efficacy for platelet products. This study introduces an efficient, accessible and cost-effective "X irradiation system" for platelet concentrates (PCs). By constructing a suitable radiation box (phantom) for a clinically available linear accelerator, an "X irradiation system" was designed specifically for PCs. PCs were divided into three equal bags either exposed to X- and gamma-irradiation or kept unirradiated (control). Irradiation-induced inhibition of T cells proliferation was examined by MTT and cell cycle assays on mononuclear cells (MNCs) obtained from PCs. The inhibitory effect of irradiation on allorecognition ability of MNCs was assessed by mixed lymphocyte reaction where MTT evaluated lymphocyte proliferation responses and flowcytometry examined CD8+T lymphocytes activity. Platelet activation was also examined with P-selectin expression and PAC-1 binding by flowcytometry. X- and gamma-irradiation reduced T cell proliferation while disturbing the cell-cycle with reduced entry of T-cells into the S phase and their G2 arrest. Both types of irradiations also effectively reduced "lymphocyte allorecognition responses" while inactivating CD8+T lymphocytes in platelet products but with no significant effect on platelet activity. This is the first study that showed "X irradiation system" effectively suppresses T cell proliferation and CD8+T lymphocyte activity in platelet products, with no effect to platelet quality and activation markers. This may suggest the LINAC-based "X irradiation system" with a dose of 30Gy as efficient and safe as gamma-irradiation for platelet products. [ABSTRACT FROM AUTHOR]

To ensure an even segregation of chromosomes during somatic cell division, eukaryotes rely on mitotic spindles. Here, we measured prime characteristics of the Arabidopsis mitotic spindle and built a three-dimensional dynamic model using Cytosim. We identified the cell-cycle regulator CYCLIN-DEPENDENT KINASE B1 (CDKB1) together with its cyclin partner CYCB3;1 as key regulators of spindle morphology in Arabidopsis. We found that the augmin component ENDOSPERM DEFECTIVE1 (EDE1) is a substrate of the CDKB1;1-CYCB3;1 complex. A non-phosphorylatable mutant rescue of ede1 resembled the spindle phenotypes of cycb3;1 and cdkb1 mutants and the protein associated less efficiently with spindle microtubules. Accordingly, reducing the level of augmin in simulations recapitulated the phenotypes observed in the mutants. Our findings emphasize the importance of cell-cycle-dependent phospho-control of the mitotic spindle in plant cells and support the validity of our model as a framework for the exploration of mechanisms controlling the organization of the eukaryotic spindle. [Display omitted] • Generation of a 3D simulation of the Arabidopsis spindle • B1-type CDKs together with a B3-type cyclin control spindle morphology via augmin • The requirement of augmin for spindle morphology is recapitulated by 3D simulations Romeiro Motta et al. show that spindle morphology is controlled by the phosphorylation of augmin performed by a cell-cycle regulator complex in Arabidopsis. By constructing a tridimensional simulation of the Arabidopsis mitotic spindle, combined with several experimental approaches, they show that changes in augmin activity largely impact spindle organization. [ABSTRACT FROM AUTHOR]

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 (IC50, 3.51 μM vs. 35.45 μM) and DU145 (IC50, 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. [ABSTRACT FROM AUTHOR]

Background and Purpose: Gastrointestinal tumours overexpress voltage‐gated calcium (CaV3) channels (CaV3.1, 3.2 and 3.3). CaV3 channels regulate cell growth and apoptosis colorectal cancer. Gossypol, a polyphenolic aldehyde found in the cotton plant, has anti‐tumour properties and inhibits CaV3 currents. A systematic study was performed on gossypol blocking mechanism on CaV3 channels and its potential anticancer effects in colon cancer cells, which express CaV3 isoforms. Experimental Approach: Transcripts for CaV3 proteins were analysed in gastrointestinal cancers using public repositories and in human colorectal cancer cell lines HCT116, SW480 and SW620. The gossypol blocking mechanism on CaV3 channels was investigated by combining heterologous expression systems and patch‐clamp experiments. The anti‐tumoural properties of gossypol were estimated by cell proliferation, viability and cell cycle assays. Ca2+ dynamics were evaluated with cytosolic and endoplasmic reticulum (ER) Ca2+ indicators. Key Results: High levels of CaV3 transcripts correlate with poor prognosis in gastrointestinal cancers. Gossypol blockade of CaV3 isoforms is concentration‐ and use‐dependent interacting with the closed, activated and inactivated conformations of CaV3 channels. Gossypol and CaV3 channels down‐regulation inhibit colorectal cancer cell proliferation by arresting cell cycles at the G0/G1 and G2/M phases, respectively. CaV3 channels underlie the vectorial Ca2+ uptake by endoplasmic reticulum in colorectal cancer cells. Conclusion and Implications: Gossypol differentially blocked CaV3 channel and its anticancer activity was correlated with high levels of CaV3.1 and CaV3.2 in colorectal cancer cells. The CaV3 regulates cell proliferation and Ca2+ dynamics in colorectal cancer cells. Understanding this blocking mechanism maybe improve cancer therapies. [ABSTRACT FROM AUTHOR]

The transcriptional coactivator Yes-associated protein (YAP)/transcriptional co-activator with PDZ binding motif (TAZ) induces cell proliferation through nuclear localization at low cell density. Conversely, at extremely high cell density, the Hippo pathway, which regulates YAP/TAZ, is activated. This activation leads to the translocation of YAP/TAZ into the cytoplasm, resulting in cell cycle arrest. Various cancer cells have several times more YAP/TAZ than normal cells. However, it is not entirely clear whether this several-fold increase in YAP/TAZ alone is sufficient to overcome proliferation inhibition (contact inhibition) under high-density conditions, thereby allowing continuous proliferation. In this study, we construct a three-dimensional (3D) mathematical model of cell proliferation incorporating the Hippo-YAP/TAZ pathway. Herein, a significant innovation in our approach is the introduction of a novel modeling component that inputs cell density, which reflects cell dynamics, into the Hippo pathway and enables the simulation of cell proliferation as the output response. We assume such 3D model with cell-cell interactions by solving reaction and molecular dynamics (MD) equations by applying adhesion and repulsive forces that act between cells and frictional forces acting on each cell. We assume Lennard-Jones (12-6) potential with a softcore character so that each cell secures its exclusive domain. We set cell cycles composed of mitotic and cell growth phases in which cells divide and grow under the influence of cell kinetics. We perform mathematical simulations at various YAP/TAZ levels to investigate the extent of YAP/TAZ increase required for sustained proliferation at high density. The results show that a twofold increase in YAP/TAZ levels of cancer cells was sufficient to evade cell cycle arrest compared to normal cells, enabling cells to continue proliferating even under high-density conditions. Finally, this mathematical model, which incorporates cell-cell interactions and the Hippo-YAP/TAZ pathway, may be applicable for evaluating cancer malignancy based on YAP/TAZ levels, developing drugs to suppress the abnormal proliferation of cancer cells, and determining appropriate drug dosages. The source codes are freely available. Author summary: This study numerically clarifies multicellular spatiotemporal properties by solving MD and reaction equations for application to any reaction network. We applied the Hippo-YAP/TAZ signaling pathway as a reaction network model for normal and cancer tissues. Moreover, we quantitatively discuss the cell proliferation outcomes in normal and cancerous tissues by calculating time-dependent multicellular structures on the 3D model using the MD method. This approach paves the way for the mathematical exploration of patterns in physical therapies, such as drug administration. [ABSTRACT FROM AUTHOR]

Breast cancer is the most common cancer diagnosed in women worldwide. However, the effective treatment for breast cancer progression is still being sought. The activation of cannabinoid receptor (CB) has been shown to negatively affect breast cancer cell survival. Our previous study also reported that breast cancer cells responded to various combinations of CB1 and CB2 agonists differently. Nonetheless, the mechanism underlying this effect and whether this phenomenon can be seen in other cancer characteristics remain unknown. Therefore, this study aims to further elucidate the effects of highly selective CB agonists and their combination on triple-negative breast cancer proliferation, cell cycle progression, invasion, lamellipodia formation as well as proteomic profile of MDA-MB-231 breast cancer cells. The presence of CB agonists, specifically a 2:1 (ACEA: GW405833) combination, prominently inhibited colony formation and induced the S-phase cell cycle arrest in MDA-MB-231 cells. Furthermore, cell invasion ability and lamellipodia formation of MDA-MB-231 were also attenuated by the exposure of CB agonists and their 2:1 combination ratio. Our proteomic analysis revealed proteomic profile alteration in MDA-MB-231 upon CB exposure that potentially led to breast cancer suppression, such as ZPR1/SHC1/MAPK-mediated cell proliferation and AXL/VAV2/RAC1-mediated cell motility pathways. Our findings showed that selective CB agonists and their combination suppressed breast cancer characteristics in MDA-MB-231 cells. The exposure of CB agonists also altered the proteomic profile of MDA-MB-231, which could lead to cell proliferation and motility suppression. [ABSTRACT FROM AUTHOR]

Aurora kinase A (AURKA) is a major regulator of the cell cycle. A prominent association exists between high expression of AURKA and cancer, and impairment of AURKA levels can trigger its oncogenic activity. In order to explore the contribution of post-transcriptional regulation to AURKA expression in different cancers, we carried out a meta-analysis of -omics data of 18 cancer types from The Cancer Genome Atlas (TCGA). Our study confirmed a general trend for increased AURKA mRNA in cancer compared to normal tissues and revealed that AURKA expression is highly dependent on post-transcriptional control in several cancers. Correlation and clustering analyses of AURKA mRNA and protein expression, and expression of AURKA-targeting hsa-let-7a miRNA, unveiled that hsa-let-7a is likely involved to varying extents in controlling AURKA expression in cancers. We then measured differences in the short/long ratio (SLR) of the two alternative cleavage and polyadenylation (APA) isoforms of AURKA mRNA across cancers compared to the respective healthy counterparts. We suggest that the interplay between APA and hsa-let-7a targeting of AURKA mRNA may influence AURKA expression in some cancers. hsa-let-7a and APA may also independently contribute to altered AURKA levels. Therefore, we argue that AURKA mRNA and protein expression are often discordant in cancer as a result of dynamic post-transcriptional regulation. [ABSTRACT FROM AUTHOR]

Inhibition of CDK4/6 kinases has led to improved outcomes in breast cancer. Nevertheless, only a minority of patients experience long-term disease control. Using a large, clinically annotated cohort of patients with metastatic hormone receptor-positive (HR+) breast cancer, we identify TP53 loss (27.6%) and MDM2 amplification (6.4%) to be associated with lack of long-term disease control. Human breast cancer models reveal that p53 loss does not alter CDK4/6 activity or G1 blockade but instead promotes drug-insensitive p130 phosphorylation by CDK2. The persistence of phospho-p130 prevents DREAM complex assembly, enabling cell-cycle re-entry and tumor progression. Inhibitors of CDK2 can overcome p53 loss, leading to geroconversion and manifestation of senescence phenotypes. Complete inhibition of both CDK4/6 and CDK2 kinases appears to be necessary to facilitate long-term response across genomically diverse HR+ breast cancers. [Display omitted] • TP53 mutation is associated with lack of long-term disease control on CDK4/6i • p53 deficiency suppresses DREAM complex in BC cells, enabling cell-cycle re-entry • Disruption of DREAM complex is sufficient to promote long-term CDK4/6i resistance • Inhibition of CDK2 overcomes p53 loss, promoting geroconversion and disease control Kudo et al. demonstrate that TP53 loss in HR+ breast cancer is clinically associated with long-term failure of CDK4/6i due to cellular escape from quiescence via impaired DREAM complex assembly. Selective CDK2 inhibition is found to reinforce p130 dephosphorylation in p53 mutant models and facilitate durable tumor growth suppression. [ABSTRACT FROM AUTHOR]

PTBP1KD-BMSCs can reduce the cerebral infarction volume, neurological function score, and improve neuronal damage in rats with ischemia–reperfusion, which is attributed to the down-regulation of PTBP1 , which inhibits the proliferation of BMSCs, slows down cell aging, and enhances cell viability. PTBP1KD-BMSCs may play a protective role in regulating ferroptosis by inhibiting the JNK/P38 pathway to increase the antioxidant activity and inhibit lipid peroxidation in neurons with ischemia–reperfusion injury. [Display omitted] • PTBP1KD-BMSCs treat ischemia–reperfusion injury in neurons. • PTBP1 regulates the BMSC cell cycle and increases cell viability. • PTBP1KD-BMSCs conditioned medium protects PC12 cells from OGD/R. • It inhibits ferroptosis and regulates the P38/JNK pathway in PC12 cells. Over the years, the neuroprotective potential of bone marrow mesenchymal stem cells (BMSCs) in acute ischemic stroke has attracted significant attention. However, BMSCs face challenges like short metabolic cycles and low survival rates post-transplant. Polypyrimidine tract-binding protein 1 (PTBP1) is an immunomodulatory RNA-binding protein that regulates the cell cycle and increases cell viability. This study investigated the protective effects and underlying mechanism of PTBP1 knockdown in BMSCs (PTBP1KD-BMSCs) following ischemia–reperfusion injury (IRI) in neurons. BMSCs were isolated from Sprague-Dawley rat femurs and characterized through flow cytometry and differentiation induction. PTBP1 knockdown inhibited BMSCs proliferation. Co-culture with PTBP1KD-BMSCs decreased reactive oxygen species (ROS) and malondialdehyde (MDA) levels, while increasing glutathione (GSH) production in oxygen and glucose deprivation/reperfusion-induced PC12 cells. Transcriptome sequencing analysis of PC12 cells suggested that the protective effect of PTBP1KD-BMSCs against injury may involve ferroptosis. Furthermore, western blotting showed upregulation of glutathione synthetase (GSS), glutathione peroxidase 4 (GPX4), and solute carrier family 7 member 11 (SLC7A11) in PTBP1KD-BMSCs, known negative regulators of ferroptosis. Moreover, PTBP1KD-BMSCs inhibited p38MAPK and JNK activation. In addition, PTBP1KD-BMSCs transplantation into middle cerebral artery occlusion/reperfusion (MCAO/R) rats reduced cerebral infarction volume and improved neurological function. Immunofluorescence analysis confirmed the upregulation of GSS expression in neurons of the ischemic cortex, while immunohistochemistry indicated a downregulation of p-P38. These result suggest that PTBP1KD-BMSCs can alleviate neuronal IRI by reducing oxidative stress, inhibiting ferroptosis, and modulating the MAPK pathway, providing a theoretical basis for potential treatment strategies for cerebral IRI. [ABSTRACT FROM AUTHOR]

The reversibility and stability of zinc (Zn) metal anode are closely related to inner Helmholtz plane (IHP) chemistry. The H2O‐rich IHP raises severe parasitic reactions and irregular Zn deposition, impeding the practical utility of Zn anode in aqueous Zn‐ion batteries (AZIBs). In this study, tetrahydropyran (THP), a five‐carbon heterocyclic ether with permanent dipole moment and hydrophobic characteristic, is introduced as a self‐adsorptive additive to reshape the IHP. It squeezes out partial H2O molecules and forms a H2O‐lean IHP, benefitting for alleviating active H2O decomposition and improving the stability of Zn anode. Moreover, the adsorbed THP induces the preferential nucleation of Zn (002) plane, facilitating dendrite‐free growth and improving the reversibility of Zn anode. Consequently, the Zn||Zn symmetric cell enables to cycle over 3600 h at 5 mA cm−2@ 1 mAh cm−2. The Zn||Cu half‐cell can stably cycle over 400 cycles with 99.9% coulombic efficiency even under harsh test conditions (10 mA cm−2@5 mAh cm−2) with 30 µm Zn foil. The Zn||NH4V4O10 full cell maintains 92.6% capacity retention after 800 cycles at 1 A g−1 and the Zn||I2 full cell enables to perform steadily over 10000 cycles with a capacity decay rate of merely 0.003% per cycle at 5 C. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are short, nonprotein‐coding RNAs, and their expression is dysregulated in malignant germ cell tumors (GCTs). Here, we investigated the causes and consequences of downregulated miR‐99a‐5p/miR‐100‐5p (functionally identical) and miR‐125b‐5p levels in malignant GCTs regardless of age, site, or subtype. Quantitative RT‐PCR was used to assess miR‐99a‐5p/miR‐100‐5p, miR‐125b‐5p, and associated gene expression in malignant GCT tissues/cell lines [seminoma (Sem), yolk sac tumor (YST), embryonal carcinoma (EC)]. Cells were treated with demethylating 5‐azacytidine and pyrosequencing was performed. Combination miR‐100‐5p/miR‐125b‐5p mimic replenishment was used to treat malignant GCT cells. Global messenger RNA (mRNA) targets of the replenished miRNAs were identified and Metascape used to study pathway effects. We found that expression levels of miR‐99a‐5p/miR‐100‐5p and miR‐125b‐5p, their respective pri‐miRNAs, and associated genes from chromosomes 11 and 21 (chr11/chr21) were downregulated and highly correlated in malignant GCT cells. Treatment with 5‐azacytidine caused upregulation of these miRNAs, with pyrosequencing revealing hypermethylation of their chr11/chr21 loci, likely contributing to miR‐100‐5p/miR‐125b‐5p downregulation. Combination miR‐100‐5p/miR‐125b‐5p mimic replenishment resulted in growth inhibition in Sem/YST cells, with miR‐100‐5p/miR‐125b‐5p mRNA targets enriched in downregulated genes, which were involved in cell cycle (confirmed by flow cytometry) and signaling pathways. Knockdown of the miR‐100‐5p/miR‐125b‐5p target tripartite motif containing 71 (TRIM71kd) recapitulated miR‐100‐5p/miR‐125b‐5p replenishment, with growth inhibition and cell cycle disruption of Sem/YST/EC cells. Further, replenishment led to reduced lin‐28 homolog A (LIN28A) levels and concomitant increases in let‐7 (MIRLET7B) tumor suppressor miRNAs, creating a sustained reversion of cell phenotype. In summary, combination miR‐100‐5p/miR‐125b‐5p mimic replenishment or TRIM71kd caused growth inhibition in malignant GCT cells via cell cycle disruption. Further studies are now warranted, including mimic treatment alongside conventional platinum‐based chemotherapy. [ABSTRACT FROM AUTHOR]

Esophageal cancer (EC) poses a substantial threat to human health. The development of radioresistance in esophageal cancer cells is a critical factor contributing to local treatment failure and an unfavorable prognosis in affected patients. A comprehensive analysis was performed using bulk RNA sequencing (RNA-seq) and single-cell RNA sequencing (scRNA-seq) data from esophageal squamous cell carcinoma (ESCC) samples. Radioresistant ESCC cell lines were generated to explore the functional role of CCNG1. Various techniques, including gene knockdown, flow cytometry, and apoptosis assays, were utilized to evaluate alterations in radiosensitivity, cell cycle progression, and cell survival in response to CCNG1 modulation. Elevated CCNG1 expression was associated with poor clinical outcomes in ESCC patients and contributed to various malignant phenotypes in ESCC cells. In radioresistant ESCC cell lines, CCNG1 knockdown markedly increased radiosensitivity, as demonstrated by enhanced G2/M phase arrest and apoptosis following radiation exposure. CellChat analysis indicated a correlation between CCNG1 and the Wnt/β-catenin signaling pathway, while western blot (WB) analysis confirmed that CCNG1 functions as a downstream effector of Wnt/β-catenin. Our study has identified CCNG1 as a key regulator of radiosensitivity in ESCC, mediated through its interaction with the Wnt/β-catenin signaling pathway. Targeting the Wnt/β-catenin/CCNG1 axis presents a promising therapeutic strategy to enhance the efficacy of radiotherapy in ESCC, potentially overcoming radioresistance and improving patient outcomes. [ABSTRACT FROM AUTHOR]

Background: Sepsis survivors exhibit immune dysregulation that contributes to poor long-term outcomes. Phenotypic and functional alterations within the myeloid compartment are believed to be a contributing factor. Here we dissect the cellular and transcriptional heterogeneity of splenic CD11b+Ly6Chigh myeloid cells that are expanded in mice that survive the cecal ligation and puncture (CLP) murine model of polymicrobial sepsis to better understand the basis of immune dysregulation in sepsis survivors. Methods: Sham or CLP surgeries were performed on C57BL/6J and BALB/c mice. Four weeks later splenic CD11b+Ly6Chigh cells from both groups were isolated for phenotypic (flow cytometry) and functional (phagocytosis and glycolysis) characterization and RNA was obtained for single-cell RNA-seq (scRNA-seq) and subsequent analysis. Results: CD11b+Ly6Chigh cells from sham and CLP surviving mice exhibit phenotypic and functional differences that relate to immune function, some of which are observed in both C57BL/6J and BALB/c strains and others that are not. To dissect disease-specific and strain-specific distinctions within the myeloid compartment, scRNA-seq analysis was performed on CD11b+Ly6Chigh cells from C57BL/6J and BALB/c sham and CLP mice. Uniform Manifold Approximation and Projection from both strains identified 13 distinct clusters of sorted CD11b+Ly6Chigh cells demonstrating significant transcriptional heterogeneity and expressing gene signatures corresponding to classical-monocytes, non-classical monocytes, M1- or M2-like macrophages, dendritic-like cells, monocyte-derived dendritic-like cells, and proliferating monocytic myeloid-derived suppressor cells (M-MDSCs). Frequency plots showed that the percentages of proliferating M-MDSCs (clusters 8, 11 and 12) were increased in CLP mice compared to sham mice in both strains. Pathway and UCell score analysis in CLP mice revealed that cell cycle and glycolytic pathways were upregulated in proliferating M-MDSCs in both strains. Notably, granule protease genes were upregulated in M-MDSCs from CLP mice. ScRNA-seq analyses also showed that phagocytic pathways were upregulated in multiple clusters including the classical monocyte cluster, confirming the increased phagocytic capacity in CD11b+Ly6Chigh cells from CLP mice observed in ex vivo functional assays in C57BL/6J mice. Conclusion: The splenic CD11b+Ly6Chigh myeloid populations expanded in survivors of CLP sepsis correspond to proliferating cells that have an increased metabolic demand and gene signatures consistent with M-MDSCs, a population known to have immunosuppressive capacity. [ABSTRACT FROM AUTHOR]

The abnormal structure of tumor vascular seriously hinders the delivery and deep penetration of drug in tumor therapy. Herein, an integrated and tumor microenvironment (TME)-responsive nanocarrier is designed, which can dilate vessle and improve the drug penetration by in situ releasing nitric oxide (NO). Briefly, S-nitroso-glutathione (GSNO) and curcumin (Cur) were encapsulatd into the Cu-doped zeolite imidazole framework-8 (Cu-ZIF-8) and modified with hyaluronic acid. The nanocarrier degradation in the weakly acidic of TME releases Cu2+, then deplete overexpressed intratumourally glutathione and transformed into Cu+, thus disrupting the balance between nicotinamide adenine dinucleotide phosphate and flavin adenine dinucleotide (NADPH/FAD) during the metabolism homeostasis of tumor. The Cu+ can generate highly toxic hydroxyl radical through the Fenton-like reaction, enhancing the chemodynamic therapeutic effect. In addition, Cu+ also decomposes GSNO to release NO by ionic reduction, leading to vasodilation and increased vascular permeability, significantly promoting the deep penetration of Cur in tumor. Afterwards, the orderly operation of cell cycle is disrupted and arrested in the S-phase to induce tumor cell apoptosis. Deep-hypothermia potentiated 2D/3D fluorescence imaging demonstrated nanocarrier regulated endogenous metabolism homeostasis of tumor. The cascade-catalysed multifunctional nanocarrier provides an approach to treat orthotopic tumor. [ABSTRACT FROM AUTHOR]

Background: Staphylococcus aureus (S. aureus) mastitis results in economic losses during dairy production. Understanding the biological progression of bovine S. aureus mastitis is vital for its prevention. Lipoteichoic acid is a key virulence factor of S. aureus (aLTA), but the main biological pathways involved in its effect on bovine mammary epithetionallial cells (Mac-T) apoptosis and necrosis have not been fully explored. Folic acid (FA) has anti-inflammatory and anti-apoptotic effects. However, the role of FA in mediating the effects of aLTA on apoptosis and necrosis remains unknown. Results: We found that low concentration of aLTA inhibited apoptosis and necrosis and that high concentration promoted the apoptosis and necrosis of Mac-T. FA pretreatment alleviated high concentration of aLTA induced apoptosis. Through transcriptomic analysis, we found that nuclear receptor subfamily 4 group A (NR4A), which alters the expression of downstream genes involved in apoptosis, proliferation, and inflammation, decreased under stimulation with a low concentration of aLTA and increased under stimulation with a high concentration of aLTA. Under stimulation with a high concentration of aLTA, the expression of the NR4A subfamily could be inhibited by FA. The results showed that aLTA may affect apoptosis and necrosis through the NR4A subfamily by targeting genes involved in bacterial invasion of epithelial cells, the IL-17 signaling pathway, DNA replication, longevity regulation, the cell cycle, and tight junction pathways. We further found that the expression trends of NR4A1 and the target genes of the NR4A subfamily (PTGS2, ESPL1, MCM5, and BUB1B) in the blood of healthy cows (Healthy), subclinical mastitis cows (SCM), and SCM supplemented with FA (SCM_FA) were consistent with those observed at the cellular level in this study. Conclusions: Our study revealed that low and high concentrations of aLTA have opposite effects on apoptosis and necrosis of Mac-T and that FA can alleviate the apoptosis induced by high concentration of aLTA. Transcriptome analysis revealed that the NR4A subfamily play a role in the ability of FA to alleviate the apoptosis and necrosis induced by high concentration of aLTA. [ABSTRACT FROM AUTHOR]

Background: Transcript variants and protein isoforms are central to unique tissue functions and maintenance of homeostasis, in addition to being associated with aberrant states such as cancer, where their crosstalk with the mutated tumor suppressor p53 may contribute to genomic instability and chromosomal rearrangements. We previously identified several novel splice variants in ovarian cancer RNA-sequencing datasets; herein, we aimed to elucidate the biological effects of the Integrin Subunit Beta 8 variant (termed pITGB8-205). Methods: Resolution of the full-length sequence of pITGB8-205 through rapid amplification of cDNA ends (RACE-PCR). Cell cycle analysis and karyotype studies were performed to further explore genomic instability. RNA-seq and proteomics analyses were used to identify the differential expression of the genes. Results: This full-length study revealed a unique 5' sequence in pITGB8-205 that differed from the reported ITGB8-205 sequence, suggesting differential regulation of this novel transcript. Under a p53 mutant background, overexpression of pITGB8-205 triggered genetic instability reminiscent of oncogene-induced replicative stress with extensive abnormal mitoses and chromosomal and nuclear aberrations indicative of chromosomal instability, leading to near whole-genome duplication that imposes energy stress on cellular resources. Micronuclei and aneuploidy are striking features of pITGB8-205-overexpressing p53-mutant cells but are not enhanced in p53 wild-type (WT) cells. RNA-seq and proteomics analyses further suggested that p53 inactivation in ovarian cancer provides a permissive intracellular molecular niche for pITGB8-205 to mediate its effects on genomic instability. This observation is pivotal considering that most high-grade serous ovarian carcinoma (HGSC) tumors express mutant p53. The resulting aneuploid clones with enhanced self-renewal and survival capabilities disrupt clonal dominance under stress yet maintain a balance between replicative stress and prosurvival advantages. Conclusion: pITGB8-205-overexpressing clones sustain ovarian tumor cell survival, achieve homeostasis and are formidable opponents of therapy. [ABSTRACT FROM AUTHOR]

Compounds that can induce oxidative stress in cancer cells while remaining nontoxic to healthy cells are extremely promising for potential anticancer drugs. 2,2′:6′,2′′-terpyridine-metal complexes possess these properties. The high level of activity (IC50 = 0.605 µM) of 2,2′:6′,2′′-terpyridine-metal complexes on lung, breast, pancreatic, and glioblastoma multiforme cancer lines and their selectivity (SI > 41.32) on human normal fibroblasts were confirmed and presented in this paper. The mechanism of action of these compounds is associated with the generation of reactive oxygen species, which affects several cellular pathways and signals. The results demonstrate that 2,2′:6′,2′′-terpyridine-metal complexes affect cell cycle inhibition in the G0/G1 phase as well as the activation of apoptosis and autophagy cell death. These results were confirmed in several independent studies, including experiments measuring the fluorescence levels of reactive oxygen species, flow cytometry, and gene and protein analysis. [ABSTRACT FROM AUTHOR]

Microtubule-targeting agents are an important class of anti-cancer drugs; their full potential is however not realized because of significant myelotoxicity and neurotoxicity. We here report 3-nitropyridine analogues as a novel group of microtubule-targeting agents with potent anti-cancer effects against a broad range of cancer types. We show that these 3-nitropyridines induce cell cycle arrest in the G2-M phase and inhibit tubulin polymerization by interacting with tubulin. Determination of the tubulin–4AZA2996 structure by X-ray crystallography demonstrated that this class of compounds binds to the colchicine-site of tubulin. Furthermore, the anti-cancer effect was demonstrated both in vitro and in vivo in a murine heterotopic xenograft model of colon cancer. When administered intravenously, 4AZA2891 effectively inhibited cancer growth. Whereas 3-nitropyridine compounds do not induce myelotoxicity at pharmacological doses, the neurotoxicity associated with microtubule-targeting agents is still present. [ABSTRACT FROM AUTHOR]

ABSTRACT Understanding the molecular signaling pathways of colorectal cancer (CRC) can be accepted as the first step in treatment strategy. Permanent mTOR signaling activation stimulates the CRC process via various biological processes. It supplies the survival of CRC stem cells, tumorigenesis, morbidity, and decreased response to drugs in CRC pathogenesis. Therefore, inhibition of the mTOR signaling by numerous bioactive components may be effective against CRC. The study aims to discuss the therapeutic capacity of various polyphenols, terpenoids, and alkaloids on CRC via the PI3K/Akt/mTOR pathway. The potential molecular effects of bioactive compounds on the mTOR pathway's upstream and downstream targets are examined. Each bioactive component causes various physiological processes, such as triggering free radical production, disruption of mitochondrial membrane potential, cell cycle arrest, inhibition of CRC stem cell migration, and suppression of glycolysis through mTOR signaling inhibition. As a result, carcinogenesis is inhibited by inducing apoptosis and autophagy. However, it should be noted that studies are primarily in vitro dose‐dependent treatment researchers. This study raises awareness about the role of phenolic compounds in treating CRC, contributing to their future use as anticancer agents. These bioactive compounds have the potential to be developed into food supplementation to prevent and treat various cancer types including CRC. This review has the potential to lead to further development of clinical studies. In the future, mTOR inhibition by applying several bioactive agents using advanced drug delivery systems may contribute to CRC treatment with 3D cell culture and in vivo clinical studies. [ABSTRACT FROM AUTHOR]

Understanding the dynamics of RNA targeting to membraneless organelles is essential to disentangle their functions. Here, we investigate how P-bodies (PBs) evolve during cell-cycle progression in HEK293 cells. PB purification across the cell cycle uncovers widespread changes in their RNA content, partly uncoupled from cell-cycle-dependent changes in RNA expression. Single-molecule fluorescence in situ hybridization (FISH) shows various mRNA localization patterns in PBs peaking in G1, S, or G2, with examples illustrating the timely capture of mRNAs in PBs when their encoded protein becomes dispensable. Rather than directly reflecting absence of translation, cyclic mRNA localization in PBs can be controlled by RBPs, such as HuR in G2, and by RNA features. Indeed, while PB mRNAs are AU rich at all cell-cycle phases, they are specifically longer in G1, possibly related to post-mitotic PB reassembly. Altogether, our study supports a model where PBs are more than a default location for excess untranslated mRNAs. [Display omitted] • Cyclic mRNA accumulation in PBs is partly uncoupled from cytoplasmic abundance • smFISH highlights various patterns of cyclic PB mRNA accumulation, notably in G1 • HuR-mRNA interactions reduce mRNA PB localization in G2 • PB mRNAs are particularly long in G1, while AU rich all along the cell cycle Safieddine et al. purify P-bodies across the cell cycle, revealing that their RNA content is dynamic and partly uncoupled from that of the surrounding cytoplasm. RNA imaging highlights various patterns of cyclic mRNA localization. Extrinsic and intrinsic factors such as HuR and RNA length contribute to cyclic mRNA localization. [ABSTRACT FROM AUTHOR]

Background: Prostate cancer (PCa) is one of the most common malignancies of the urinary system. Cuproptosis, a newly discovered form of cell death. The relationship between cuproptosis-related long non-coding RNAs (ClncRNAs) related to PCa and prognosis remains unclear. This study aimed to explore the clinical significance of novel ClncRNAs in the prognostic assessment of PCa. Methods: ClncRNAs and differentially expressed mRNAs linked to these ClncRNAs were identified using Pearson's correlation and differential expression analyses. A prognostic signature (risk score) comprising three ClncRNAs was established based on multivariable Cox regression analysis. The predictive performance of this ClncRNAs signature was validated using receiver operating characteristic curves and nomograms. Finally, further in vitro cell experiments were conducted for validation, including quantitative polymerase chain reaction (qPCR), western blot (WB), cell proliferation assays, cell migration assays, cell invasion assays, apoptosis, and cell cycle analysis. Results: We constructed a prognostic signature of ClncRNAs for PCa comprising three key differentially expressed ClncRNAs(AC010896-1, AC016394-2, and SNHG9). Multivariable Cox regression analysis indicated that clinical staging and risk scores of the ClncRNAs signature were independent prognostic factors for PCa. Compared to other clinical features, the ClncRNAs signature exhibited higher diagnostic efficiency and performed well in predicting the 1-, 3-, and 5-year progression-free intervals (PFIs) for PCa. Notably, in terms of immune activity, PCa patients with high-risk scores exhibited higher tumor mutational burden (TMB) levels, while their Tumor Immune Dysfunction and Exclusion (TIDE) scores were lower than those of PCa patients with low-risk scores. Additionally, in vitro cellular functional experiments, we knocked down SNHG9 that is the most significantly differentially expressed ClncRNA among the three key ClncRNAs. SNHG9 knockdown resulted in a significant increase in G1 phase cells and a decrease in S and G2 phases, indicating inhibition of DNA synthesis and cell cycle progression. Colony formation assays showed reduced clonogenic ability, with fewer and smaller colonies. Western blot analysis revealed the upregulation of the key cuproptosis-related mRNAs FDX1 and DLST. These findings suggested that SNHG9 promotes PCa cell proliferation, migration, and invasion. Conclusion: Building on the three ClncRNAs, we identified a novel prognostic signature of PCa. The ClncRNA SNHG9 can promote PCa cell proliferation, migration, and invasion. [ABSTRACT FROM AUTHOR]

Objective: Long non-coding RNAs (lncRNAs) LINC02466 is an lncRNA newly linked to the adverse outcomes in primary liver cancer patients, and its crucial involvement in the disease's escalation. Decoding the specific role of LINC02466 in HCC progression is of great significance to provide a potential therapeutic target for HCC. Methods: RT-qPCR and Western Blot techniques was used to analyze the expression levels of LINC02466 in both malignant and surrounding healthy liver tissues. CCK8 assays and colony formation experiments indicates the LINC02466's effect on the proliferation rates of liver cancer cells. Flow cytometry was pivotal in revealing its significant influence on the cell cycle of these cells. Kaplan–Meier survival analysis with log-rank tests were employed. Results: The suppression of LINC02466 markedly reduces the stemness attributes of liver cancer cells, indicating a potential therapeutic target. LINC02466 overexpression significantly increased tumor growth rates and final volumes. Further research indicated that LINC02466 significantly influences liver cancer progression through regulating the mTOR signaling pathway. Conclusion: LINC02466 regulating cell proliferation, the cell cycle, and stemness characteristics via the mTOR pathway, suggesting LINC02466 as a potential therapeutic target for primary liver cancer. [ABSTRACT FROM AUTHOR]

Kinetochores form the interface between chromosomes and spindle microtubules and are thus under tight control by a complex regulatory circuitry. The Aurora B kinase plays a central role within this circuitry by destabilizing improper kinetochore-microtubule attachments and relaying the attachment status to the spindle assembly checkpoint. Intriguingly, Aurora B is conserved even in kinetoplastids, a group of early-branching eukaryotes which possess a unique set of kinetochore proteins. It remains unclear how their kinetochores are regulated to ensure faithful chromosome segregation. Here, we show in Trypanosoma brucei that Aurora B activity controls the metaphase-to-anaphase transition through phosphorylation of the divergent Bub1-like protein KKT14. Depletion of KKT14 overrides the metaphase arrest resulting from Aurora B inhibition, while expression of non-phosphorylatable KKT14 delays anaphase onset. Finally, we demonstrate that re-targeting Aurora B to the outer kinetochore suffices to promote mitotic exit but causes extensive chromosome missegregation in anaphase. Our results indicate that Aurora B and KKT14 are involved in an unconventional circuitry controlling cell cycle progression in trypanosomes. [ABSTRACT FROM AUTHOR]

Adult oligodendrocyte progenitors (aOPCs) generate myelinating oligodendrocytes like neonatal progenitors (nOPCs), and they also display unique functional features. Here, using unbiased histone proteomics analysis and ChIP sequencing analysis of PDGFRa+ OPCs sorted from neonatal and adult Pdgfra-H2B-EGFP reporter mice, we identify the activating H4K8ac histone mark as enriched in the aOPCs. We detect increased occupancy of the H4K8ac activating mark at chromatin locations corresponding to genes related to the progenitor state (e.g., Hes5, Gpr17), metabolic processes (e.g., Txnip, Ptdgs), and myelin components (e.g., Cnp, Mog). aOPCs showed higher levels of transcripts related to lipid metabolism and myelin, and lower levels of transcripts related to cell cycle and proliferation compared with nOPCs. In addition, pharmacological inhibition of histone acetylation decreased the expression of the H4K8ac target genes in aOPCs and decreased their proliferation. Overall, this study identifies acetylation of the histone H4K8 as a regulator of the proliferative capacity of aOPCs. [ABSTRACT FROM AUTHOR]

The intracellular position of genes may impact their expression, but it has not been possible to accurately measure the 3D position of chromosomal loci. In 2D, loci can be tracked using arrays of DNA-binding sites for transcription factors (TFs) fused with fluorescent proteins. However, the same 2D data can result from different 3D trajectories. Here, we have developed a deep learning method for super-resolved astigmatism-based 3D localization of chromosomal loci in live E. coli cells which enables a precision better than 61 nm at a signal-to-background ratio of ~4 on a heterogeneous cell background. Determining the spatial localization of chromosomal loci, we find that some loci are at the periphery of the nucleoid for large parts of the cell cycle. Analyses of individual trajectories reveal that these loci are subdiffusive both longitudinally (x) and radially (r), but that individual loci explore the full radial width on a minute time scale. The intracellular position of genes in the chromosome may impact their expression. To measure the 3D positions of chromosomal loci over the bacterial cell cycle, we have developed a combination of microfluidics, optics, and deep learning. [ABSTRACT FROM AUTHOR]

To maintain protein homeostasis in changing nutrient environments, cells must precisely control the amount of their proteins, despite the accompanying changes in cell growth and biosynthetic capacity. As nutrients are major regulators of cell cycle length and progression, a particular challenge arises for the nutrient-dependent regulation of 'cell cycle genes', which are periodically expressed during the cell cycle. One important example are histones, which are needed at a constant histone-to-DNA stoichiometry. Here we show that budding yeast achieves histone homeostasis in different nutrients through a decoupling of transcript and protein abundance. We find that cells downregulate histone transcripts in poor nutrients to avoid toxic histone overexpression, but produce constant amounts of histone proteins through nutrient-specific regulation of translation efficiency. Our findings suggest that this allows cells to balance the need for rapid histone production under fast growth conditions with the tight regulation required to avoid toxic overexpression in poor nutrients. Synopsis: Changing nutrient environments challenge cells as they affect both protein biosynthesis and cell cycle length. This work shows that budding yeast maintains histone-to-DNA stoichiometry across nutrient conditions through promoter-mediated regulation of transcription and translation. Histone-to-DNA stoichiometry is maintained across nutrient conditions. Nutrient-dependent histone homeostasis can be mediated by histone promoters. Nutrient-dependent histone homeostasis involves separate regulation of transcription and translation. Histone mRNA concentration is lower, and histone overexpression more toxic, in poor growth conditions. Budding yeast maintains histone-to-DNA stoichiometry across nutrient conditions through promoter-mediated regulation of transcription and translation, potentially to avoid toxic overexpression. [ABSTRACT FROM AUTHOR]

A novel series of six [1,2,4]triazolo[3,4-a]isoquinolin-3-yl)-3-(1,3-diphenyl-1H-pyrazol-4-yl)prop-2-en-1-ones (3a–3f) was designed and synthesized. They were characterized based on spectral and elemental analyses. In silico studies were also committed to provide insights and a better understanding of their structural features. The six compounds were screened for their antiproliferative activity using the MTT assay against five human cancer cell lines, namely, A549, HCT116, PC3, HT29, and MCF-7 in parallel with the non-cancerous human lung cell line WI-38. The results showed that 3e and 3f have potential cytotoxic activities, especially on A549 cells with IC50 = 2.3 µM and 1.15 µM, respectively. Meanwhile, they recorded a minimal cytotoxic effect on WI-38 cells. Concerning the molecular mechanism of action, the present study showed the inhibitory effect of the six compounds against total EGFR. The most potent EGFR inhibitors were 3e and 3f with IC50 = 0.031 µM and 0.023 µM, respectively. The selectivity index of 3f for EGFRT790M was 1.81 times more selective than that of lapatinib. In addition, 3e and 3f initiated cell cycle arrest at the G2/M and pre-G1 phases along with the downregulation of anti-apoptotic protein Bcl2 and the upregulation of pro-apoptotic proteins: p53, Bax, and caspases 3, 8, and 9. Further studies are recommended to evaluate animal models' promising anticancer activity and molecular mechanism of triazolo[3,4-a]isoquinoline derivatives 3e and 3f. [ABSTRACT FROM AUTHOR]

Background: Prevention and treatment of metastatic breast cancer (BC) is an unmet clinical need. The retinoic acid derivative fenretinide (FeR) was previously evaluated in Phase I-III clinical trials but, despite its excellent tolerability and antitumor activity in preclinical models, showed limited therapeutic efficacy due to poor bioavailability. We recently generated a new micellar formulation of FeR, Bionanofenretinide (Bio-nFeR) showing enhanced bioavailability, low toxicity, and strong antitumor efficacy on human lung cancer, colorectal cancer, and melanoma xenografts. In the present study, we tested the effect of Bio-nFeR on a preclinical model of metastatic BC. Methods: We used BC cell lines for in vitro analyses of cell viability, cell cycle and migratory capacity. For in vivo studies, we used HER2/neu transgenic mice (neuT) as a model of spontaneously metastatic BC. Mice were treated orally with Bio-nFeR and at sacrifice primary and metastatic breast tumors were analyzed by histology and immunohistochemistry. Molecular pathways activated in primary tumors were analyzed by immunoblotting. Stem cell content was assessed by flow cytometry, immunoblotting and functional assays such as colony formation ex vivo and second transplantation assay in immunocompromised mice. Results: Bio-nFeR inhibited the proliferation and migration of neuT BC cells in vitro and showed significant efficacy against BC onset in neuT mice. Importantly, Bio-nFeR showed the highest effectiveness against metastatic progression, counteracting both metastasis initiation and expansion. The main mechanism of Bio-nFeR action consists of promoting tumor dormancy through a combined induction of antiproliferative signals and inhibition of the mTOR pathway. Conclusion: The high effectiveness of Bio-nFeR in the neuT model of mammary carcinogenesis, coupled with its low toxicity, indicates this formulation as a potential candidate for the treatment of metastatic BC and for the adjuvant therapy of BC patients at high risk of developing metastasis. [ABSTRACT FROM AUTHOR]

The AgNPs‐Rsvl complex was investigated for its anti‐cancer effects on human breast cancer cell lines. The UV–vis spectrophotometer was used to perform synthesized AgNPs‐Rsvl binding analyses. The size and morphology of the AgNPs were determined by field emission gun‐transmission electron microscopy (FEG‐TEM). The surface chemical signatures of AgNPs nanocomposites were assessed using FT‐IR spectroscopy, and the zeta potential (ζ potential) of various AgNPs colloids was measured using the Zetasizer Nano ZS analyzer. Cell proliferation was analyzed using the XTT method and cell apoptosis was analyzed using the AnnexinV‐fluorescein isothiocyanate (FITC)/PI method. The effects of resveratrol were evaluated on human breast cancer cell lines (MDA‐MB‐231 and MCF‐7). As a result of flow cytometry, a decrease in viability was observed in the MDA‐MB‐231 cells, and an increase in early apoptotic, late apoptotic, and necrotic values was observed. Resveratrol arrests the cell cycle in the G0/G1 phase but when used with AgNPs, the cell cycle arrests at the S phase. The combination of resveratrol and AgNPs exhibited a cytotoxic effect on cancer cells, with MDA‐MB‐231 breast cancer cells being the most sensitive to AgNPs‐Rsvl. Resveratrol and AgNPs arrested the cell cycle and increased apoptosis, indicating their potential use in breast cancer treatment. [ABSTRACT FROM AUTHOR]

The zinc metal anodes are liable to experience detrimental dendrite growth and side reactions, thereby limiting the lifespan of aqueous Zn‐ion batteries. Here, a readily available supramolecule, dimethoxypillar[5]arene (DP[5]), is utilized as a shielding layer to stabilize the Zn anode by exploiting its zincophilicity derived from the ─OCH3 functional groups and its hydrophobicity from the hydrophobic backbone. The DP[5] shielding layer regulates the solvation sheath of Zn2+ and facilitates uniform zinc deposition. Swelling and dendrite formation are obviously suppressed in both the symmetric and full cells. The DP[5]‐Zn symmetrical cell cycles stably for 5500 h, and achieves a coulombic efficiency of 99.76% in a DP[5]‐Zn||Cu half‐cell after 2200 cycles. The DP[5]‐Zn||V2O5 full cell maintains 92.03% of initial capacity after 6000 cycles. Given the cost‐effective fabrication and environmental friendliness of DP[5] films, this material may pave the way for practical applications of zinc anodes. [ABSTRACT FROM AUTHOR]

Background: Extracellular vesicles (EVs) regulate cell metabolism and various biological processes by delivering specific proteins and nucleic acids to surrounding cells. We aimed to investigate the effects of the cargo contained in EVs derived from adipose-derived stem cells (ASCs) on the porcine embryonic development. Methods: ASCs were isolated from porcine adipose tissue and characterized using ASC-specific markers via flow cytometry. EVs were subsequently extracted from the conditioned media of the established ASCs. These EVs were added to the in vitro culture environment of porcine embryos to observe qualitative improvements in embryonic development. Furthermore, the proteins within the EVs were analyzed to investigate the underlying mechanisms. Results: We observed a higher blastocyst development rate and increased mitochondrial activity in early stage embryos in the ASC-EVs-supplemented group than in the controls (24.8% ± 0.8% vs. 28.6% ± 1.1%, respectively). The terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay of blastocysts also revealed significantly reduced apoptotic cells in the ASC-EVs-supplemented group. Furthermore, through proteomics, we detected the proteins in ASC-EVs and blastocysts from each treatment group. This analysis revealed a higher fraction of proteins in the ASC-EVs-supplemented group than in the controls (1,547 vs. 1,495, respectively). Gene analysis confirmed that ASC-EVs showed a high expression of tyrosine-protein kinase (SRC), whereas ASC-EVs supplemented blastocysts showed a higher expression of Cyclin-dependent kinase 1 (CDK1). SRC is postulated to activate protein kinase B (AKT), which inhibits the forkhead box O signaling pathway and activates CDK1. Subsequently, CDK1 activation influences the cell cycle, thereby affecting in vitro embryonic development. Conclusion: ASC-EVs promote mitochondrial activity, which is crucial for the early development of blastocysts and vital in the downregulation of apoptosis. Additionally, ASC-EVs supply SRC to porcine blastocysts, thereby elongating the cell cycle. [ABSTRACT FROM AUTHOR]

Background: Anlotinib, a highly selective inhibitor of VEGFR2, has demonstrated significant anti-tumor effects in various cancers. However, its potential synergistic effects with DDP (cisplatin) in breast cancer (BRCA) remain to be fully elucidated. This study aims to discover the therapeutic efficacy of anlotinib on BRCA, specifically the synergistic effects with DDP, and to elucidate the underlying molecular mechanisms. Methods: BRCA cells were treated with anlotinib and/or DDP. The proliferation, migration and invasion capabilities of BRCA cells were evaluated using CCK-8 assays, cell cycle distribution, clone formation assays, wound healing assays and transwell assays. Cell apoptosis was detected by flow cytometry technique and Hoechst33342 fluorescence staining. The potential mechanism of anlotinib in the development of BRCA was predicted through bioinformatics analysis, and the mRNA or protein levels were subsequently quantified using qPCR, immunofuorescence and western blot. The anti-breast cancer efficacy of anlotinib was evaluated in vivo using a xenograft tumor model. Results: Our findings reveal that increased VEGFA expression in BRCA patients is associated with poorer prognosis, underscoring the need for targeted therapeutic strategies. We also demonstrate that both anlotinib and DDP independently inhibit BRCA cell growth, migration, and invasion, while their combination exhibits a synergistic effect, significantly enhancing the inhibition of these oncogenic processes. This synergy is further evident through the induction of apoptosis and autophagy in BRCA cells. Mechanistically, anlotinib's effectiveness is linked to its inhibition of the JAK2/STAT3 pathway, a critical axis in BRCA progression. In vivo study further support these results, showing that anlotinib markedly inhibits tumor growth in xenografted mice. Conclusion: This study confirms the efficacy of anlotinib or in combination with DDP and elucidates the mechanism behind anlotinib's effectiveness, highlighting its role in inhibiting the JAK2/STAT3 pathway. [ABSTRACT FROM AUTHOR]

Colorectal cancer (CRC) is one of the primary contributors to cancer-related fatalities, with up to 80% of advanced CRC cases exhibiting mutations in the p53 gene. Unfortunately, the development of new compounds targeting mutant p53 is quite limited. The anticancer effects of Dehydroepiandrosterone (DHEA) on various cancers have been reported. However, the suppressive effect of DHEA on CRC cells harboring wild-type or mutant p53 gene remains controversial. This study emphasized revealing the suppressive mechanism and the effect of DHEA on CRC cell tumorigenesis in the presence of wild-type or mutant p53 gene. We demonstrate that DHEA causes CRC cell death and cell cycle arrest in a dose and time-dependent manner. Notably, DHEA exhibits similar inhibitory effects on CRC cells regardless of the p53 gene status. Further study reveals that DHEA induces endoplasmic reticulum (ER) stress and triggers PERK/eIF2/ATF4/CHOP UPR signaling pathway to activate autophagy followed by apoptosis, which was confirmed by suppression of 4-phenylbutyric acid (an ER stress inhibitor) or knockdown either ATF4 or CHOP. DHEA-induced apoptosis was attenuated by silencing ATG5 gene in either p53+/+ or p53−/− CRC cells, indicating autophagy regulation of apoptosis. Furthermore, DHEA treatment accompanied by bafilomycin A1 (a blocker of autophagosome degradation) leads to the accumulation of ATF4, CHOP, DR5, and p21 levels in CRC cells, implying that the degradative autophagy machinery regulates these four molecules. Consistently, DHEA demonstrates its inhibitory effect by suppressing CRC tumor formation in vivo. Altogether, we provide compelling evidence that DHEA is a potential therapeutic candidate for CRC patient treatment regardless of the p53 status through ER stress-PERK-autophagy-apoptosis axis. [ABSTRACT FROM AUTHOR]

Isoalantolactone (ISA) is a sesquiterpene lactone that could be isolated from Inula helenium as well as many other herbal plants belonging to Asteraceae. Over the past 2 decades, lots of researches have been made on ISA, which owns multiple pharmacological effects, such as antimicrobial, anticancer, anti-inflammatory, neuroprotective, antidepressant-like activity, as well as others. The anticancer effects of ISA involve proliferation inhibition, ROS overproduction, apoptosis induction and cell cycle arrest. Through inhibiting NF-κB signaling, ISA exerts its anti-inflammatory effects which are involved in the neuroprotection of ISA. This review hackled the reported pharmacological effects of ISA and associated mechanisms, providing an update on understanding its potential in drug development. [ABSTRACT FROM AUTHOR]