Nuclear Organelles Research Articles

The coilin N-terminus mediates multivalent interactions between coilin and Nopp140 to form and maintain Cajal bodies

Cajal bodies (CBs) are ubiquitous nuclear membraneless organelles (MLOs) that concentrate and promote efficient biogenesis of snRNA-protein complexes involved in splicing (snRNPs). Depletion of the CB scaffolding protein coilin disperses snRNPs, making CBs a model system for studying the structure and function of MLOs. Although it is assumed that CBs form through condensation, the biomolecular interactions responsible remain elusive. Here, we discover the unexpected capacity of coilin’s N-terminal domain (NTD) to form extensive fibrils in the cytoplasm and discrete nuclear puncta in vivo. Single amino acid mutational analysis reveals distinct molecular interactions between coilin NTD proteins to form fibrils and additional NTD interactions with the nuclear Nopp140 protein to form puncta. We provide evidence that Nopp140 has condensation capacity and is required for CB assembly. From these observations, we propose a model in which coilin NTD–NTD mediated assemblies make multivalent contacts with Nopp140 to achieve biomolecular condensation in the nucleus.

Keeping up with the condensates: The retention, gain, and loss of nuclear membrane-less organelles

In recent decades, a growing number of biomolecular condensates have been identified in eukaryotic cells. These structures form through phase separation and have been linked to a diverse array of cellular processes. While a checklist of established membrane-bound organelles is present across the eukaryotic domain, less is known about the conservation of membrane-less subcellular structures. Many of these structures can be seen throughout eukaryotes, while others are only thought to be present in metazoans or a limited subset of species. In particular, the nucleus is a hub of biomolecular condensates. Some of these subnuclear domains have been found in a broad range of organisms, which is a characteristic often attributed to essential functionality. However, this does not always appear to be the case. For example, the nucleolus is critical for ribosomal biogenesis and is present throughout the eukaryotic domain, while the Cajal bodies are believed to be similarly conserved, yet these structures are dispensable for organismal survival. Likewise, depletion of the Drosophila melanogaster omega speckles reduces viability, despite the apparent absence of this domain in higher eukaryotes. By reviewing primary research that has analyzed the presence of specific condensates (nucleoli, Cajal bodies, amyloid bodies, nucleolar aggresomes, nuclear speckles, nuclear paraspeckles, nuclear stress bodies, PML bodies, omega speckles, NUN bodies, mei2 dots) in a cross-section of organisms (e.g., human, mouse, D. melanogaster, Caenorhabditis elegans, yeast), we adopt a human-centric view to explore the emergence, retention, and absence of a subset of nuclear biomolecular condensates. This overview is particularly important as numerous biomolecular condensates have been linked to human disease, and their presence in additional species could unlock new and well characterized model systems for health research.

Identification of Signal Pathways and Hub Genes of Pulmonary Arterial Hypertension by Bioinformatic Analysis.

Pulmonary arterial hypertension (PAH) is a progressive and complex pulmonary vascular disease with poor prognosis. The aim of this study was to provide a new understanding of the pathogenesis of disease and potential treatment targets for patients with PAH based on multiple-microarray analysis.Two microarray datasets (GSE53408 and GSE113439) downloaded from the Gene Expression Omnibus (GEO) database were analysed. All the raw data were processed by R, and differentially expressed genes (DEGs) were screened out by the “limma” package. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed and visualized by R and Cytoscape software. Protein-protein interactions (PPI) of DEGs were analysed based on the NetworkAnalyst online tool. A total of 442 upregulated DEGs and 84 downregulated DEGs were identified. GO enrichment analysis showed that these DEGs were mainly enriched in mitotic nuclear division, organelle fission, chromosome segregation, nuclear division, and sister chromatid segregation. Significant KEGG pathway enrichment included ribosome biogenesis in eukaryotes, RNA transport, proteoglycans in cancer, dilated cardiomyopathy, rheumatoid arthritis, vascular smooth muscle contraction, focal adhesion, regulation of the actin cytoskeleton, and hypertrophic cardiomyopathy. The PPI network identified 10 hub genes including HSP90AA1, CDC5L, MDM2, LRRK2, CFTR, IQGAP1, CAND1, TOP2A, DDX21, and HIF1A. We elucidated potential biomarkers and therapeutic targets for PAH by bioinformatic analysis, which provides a theoretical basis for future study.

КЛИНИКО-ДИАГНОСТИЧЕСКОЕ ЗНАЧЕНИЕ АНТИНУКЛЕАРНЫХ АНТИТЕЛ В ПРАКТИКЕ ПЕДИАТРА (ЧАСТЬ I)

In recent decades, there has been an increase in the incidence of various autoimmune, including rheumatic diseases, while their debut has been shifted to an earlier age. The clinical picture of rheumatic diseases is distinguished by severe polymorphism; therefore, laboratory diagnostic methods are important for diagnostic search. One of the most widely used tests in the rheumatological practice is the detection and identification of antinuclear antibodies (ANA). ANA are a large group of autoantibodies (auto-Abs) directed against various cell structures, including not only the nucleus but also the nuclear membrane, mitotic apparatus, cytoplasm components and cell organelles, as well as cell membranes. Different methods are used for testing, but the method of ANA detection through indirect immunofluorescence antinuclear antibody test (IF-ANA) is recommended as a “gold standard” with human laryngeal carcinoma HEp-2 cells being recognized as a standard substrate. IF-ANA with HEp-2 has the highest diagnostic sensitivity compared with other methods, it includes the phase of serial dilutions of positive serums and a visual analysis of the type of glow, the list of which currently has 30 patterns. ANA glow types are caused by various auto-Abs that respond with antigens located in the nucleus and cytoplasm of the HEp-2 cells line. Assessment of the type of glow in conjunction with clinical and laboratory data allows planning of further examination with confirming tests to determine the specificity of ANA, some of which, in particular, are auto-Abs to DFS70 antigen, which in the absence of other types of autoantibodies, are more likely to doubt the presence of a systemic rheumatic disease. The diagnostic specificity of various screening techniques for ANA detection is relatively low, they are detected with a fairly high frequency in patients with other diseases and in healthy individuals of different gender and age, including children.

SUMO Regulates Histone mRNA Processing and Polyadenylation

Replication‐dependent histone genes encode the only non‐polyadenylated mRNAs in mammals. Histone pre‐mRNAs, which do not contain introns, therefore require only one processing step that involves 3’ end recognition and cleavage. Both histone gene expression and pre‐mRNA processing take place in specialized nuclear organelles known as histone locus bodies. Recent studies have revealed that polyadenylation of histone mRNAs does occur, however, under specific physiological conditions. These include during normal terminal cellular differentiation, tumorigenesis, and in disease patients with Aicardi–Goutières syndrome. Despite these links between polyadenylation of histone mRNAs, human physiology, and disease, the triggers, the mechanisms, and the consequences of histone mRNA polyadenylation remain poorly defined. In recent studies of small ubiquitin‐related modifier (SUMO) knockout cell lines, we have obtained evidence that SUMO1 and SUMO2 play roles in regulating histone pre‐mRNA 3’ end processing and polyadenylation. This evidence includes alterations in histone locus body size and structure in SUMO1 and SUMO2 knockout cells, as well as mislocalization of key pre‐mRNA processing factors required for normal processing. Collectively, our findings provide new insights into control of histone pre‐mRNA processing that involves posttranslational protein modifications by SUMO1 and SUMO2.

Maternal, placental, and fetal distribution of titanium after repeated titanium dioxide nanoparticle inhalation through pregnancy

Epidemiological studies have associated ambient engineered nanomaterials or ultrafine particulate matter (PM0.1), collectively referred to as nanoparticles (NPs), with adverse pregnancy outcomes including miscarriage, preterm labor, and fetal growth restriction. Evidence from non-pregnant models demonstrate that NPs can cross the lung air-blood barrier and circulate systemically. Therefore, inhalation of NPs during pregnancy leading to fetoplacental exposure has garnered attention. The purpose of this study was to evaluate the distribution of inhaled titanium dioxide nanoparticles (nano-TiO2) from the maternal lung to maternal and fetal systemic tissues. Pregnant Sprague Dawley rats were administered whole-body exposure to filtered air or of nano-TiO2 aerosols (9.96 ± 0.06 mg/m3) between gestational day (GD) 4 and 19. On GD 20 maternal, placental, and fetal tissues were harvested then digested for ICP-MS analysis to measure concentrations of titanium (Ti). TEM was used to visualize particle internalization by the placental syncytium. The results demonstrate the extrapulmonary distribution of Ti to various maternal organs during pregnancy. Our study found Ti accumulation in the decidua/junctional and labyrinth zones of placentas embedded in all sections of uterine horns. Further, NPs deposited in the placenta, identified by TEM, were found intracellularly within nuclear, endoplasmic reticulum, and vesicle organelles. This study identified the systemic distribution and placental accumulation of Ti after nano-TiO2 aerosol inhalation in a pregnancy model. These findings arouse concerns for poor air quality for pregnant women and possible contributions to adverse pregnancy outcomes.

Identification of key genes and carcinogenic pathways in hepatitis B virus-associated hepatocellular carcinoma through bioinformatics analysis.

Backgrounds/AimsMechanisms for the development of hepatocellular carcinoma (HCC) in hepatitis B virus (HBV)-infected patients remain unclear. The aim of the present study was to identify genes and pathways involved in the development of HBV-associated HCC.MethodsThe GSE121248 gene dataset, which included 70 HCCs and 37 adjacent liver tissues, was downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) in HCCs and adjacent liver tissues were identified. Gene ontology and Kyoto Encyclopedia of Genes and Genome pathway enrichment analyses were then performed.ResultsOf 134 DEGs identified, 34 were up-regulated and 100 were down-regulated in HCCs. The 34 up-regulated DEGs were mainly involved in nuclear division, organelle fission, spindle and midbody formation, histone kinase activity, and p53 signaling pathway, whereas the 100 down-regulated DEGs were involved in steroid and hormone metabolism, collagen-coated extracellular matrix, oxidoreductase activity, and activity on paired donors, including incorporation or reduction of molecular oxygen, monooxygenase activity, and retinol metabolism. Analyses of protein-protein interaction networks with a high degree of connectivity identified significant modules containing 14 hub genes, including ANLN, ASPM, BUB1B, CCNB1, CDK1, CDKN3, ECT2, HMMR, NEK2, PBK, PRC1, RACGAP1, RRM2, and TOP2A, which were mainly associated with nuclear division, organelle fission, spindle formation, protein serine/threonine kinase activity, p53 signaling pathway, and cell cycle.ConclusionsThis study identified key genes and carcinogenic pathways that play essential roles in the development of HBV-associated HCC. This may provide important information for the development of diagnostic and therapeutic targets for HCC.

Glomerulosomes: morphologically distinct nuclear organelles of unknown nature.

The nucleus of some representatives of the genus Pelomyxa (Amoebozoa, Archamoebae, Pelobiontida) contains specific bodies (membrane-less organelles). They may be either embedded in the nucleolar mass or detached from the nucleolus. We termed these nuclear bodies the glomerulosomes for their characteristic ultrastructural appearance. The glomerulosomes are distinct nuclear bodies, about 1μm in diameter. The morphological and diagnostic unit of a glomerulosome is an electron-dense thread/string, about 30-40nm in thickness. These threads are not direct continuation of the nucleolar material. The threads create the unique geometric appearance of the glomerulosome by being organized into precisely parallel rows/cords. Each cord of the threads can curve at different angles within the glomerulosome body, but the threads themselves are not coiled. Nowadays, the glomerulosomes have been discovered in P. palustris, P.stagnalis, P.paradoxa, and Pelomyxa sp. Despite the unique appearance of glomerulosomes, their existence may be a more common phenomenon in eukaryotic cells than just a specific feature of the nucleus of elected pelomyxes.

Pan-cancer analysis combined with experiments explores the oncogenic role of spindle apparatus coiled-coil protein 1 (SPDL1)

BackgroundThe function of spindle apparatus coiled-coil protein 1 (SPDL1) as a cancer-promoting gene has been reported in a number of studies. However, the pan-cancer analysis of SPDL1 is still lacking. Here, we performed this pan-cancer analysis to evaluate the expression and prognostic value of SPDL1 and gain insights into the association between SPDL1 and immune infiltration.MethodsIn this study, based on the datasets of The cancer genome atlas (TCGA), Gene expression omnibus (GEO), The Genotype-Tissue Expression (GTEx) and Clinical Proteomic Tumor Analysis Consortium (CPTAC), we used R4.1.0 software and the online tools, including TIMER2.0, GEPIA2, cBioPortal, Modbase, UALCAN, MEXPRESS, STRING, Ensembl, NCBI, HPA, Oncomine, PhosphoNET and the Kaplan-Meier plotter, to explore the potential oncogenic roles of SPDL1. The expression of SPDL1 was also further verified by immunohistochemistry (IHC) in lung adenocarcinoma (LUAD) tissues.ResultsSPDL1 was overexpressed in most tumors compared with adjacent normal tissues, and SPDL1 expression was significantly correlated with the prognosis in most tumor types. The main type of genetic mutation of SPDL1 was missense mutation and the frequency of R318Q/W mutation was highest (4/119). The expression of SPDL1 was closely associated with genomic instability. The SPDL1 phosphorylation levels in S555 was enhanced in ovarian cancer. The SPDL1 expression was positively correlated with the immune infiltration of CD8+ T-cells and cancer-associated fibroblasts (CAFs) in most of the tumor types. Nuclear division, organelle fission and chromosome segregation were involved in the functional mechanisms of SPDL1.ConclusionsThese findings suggested that SPDL1 might serve as a biomarker for poor prognosis and immune infiltration in cancers, shedding new light on therapeutics of cancers.

Molecular Mechanisms in Pentanucleotide Repeat Diseases.

The number of neurodegenerative diseases resulting from repeat expansion has increased extraordinarily in recent years. In several of these pathologies, the repeat can be transcribed in RNA from both DNA strands producing, at least, one toxic RNA repeat that causes neurodegeneration by a complex mechanism. Recently, seven diseases have been found caused by a novel intronic pentanucleotide repeat in distinct genes encoding proteins highly expressed in the cerebellum. These disorders are clinically heterogeneous being characterized by impaired motor function, resulting from ataxia or epilepsy. The role that apparently normal proteins from these mutant genes play in these pathologies is not known. However, recent advances in previously known spinocerebellar ataxias originated by abnormal non-coding pentanucleotide repeats point to a gain of a toxic function by the pathogenic repeat-containing RNA that abnormally forms nuclear foci with RNA-binding proteins. In cells, RNA foci have been shown to be formed by phase separation. Moreover, the field of repeat expansions has lately achieved an extraordinary progress with the discovery that RNA repeats, polyglutamine, and polyalanine proteins are crucial for the formation of nuclear membraneless organelles by phase separation, which is perturbed when they are expanded. This review will cover the amazing advances on repeat diseases.

Nuclear Organization in Response to Stress: A Special Focus on Nucleoli.

In this chapter, we discuss the nuclear organization and how it responds to different types of stress. A key component in these responses is molecular traffic between the different sub-nucleolar compartments, such as nucleoplasm, chromatin, nucleoli, and various speckle and body compartments. This allows specific repair and response activities in locations where they normally are not active and serve to halt sensitive functions until the stress insult passes and inflicted damage has been repaired. We focus on mammalian cells and their nuclear organization, especially describing the central role of the nucleolus in nuclear stress responses. We describe events after multiple stress types, including DNA damage, various drugs, and toxic compounds, and discuss the involvement of macromolecular traffic between dynamic, phase-separated nuclear organelles and foci. We delineate the key proteins and non-coding RNA in the formation of stress-responsive, non-membranous nuclear organelles, many of which are relevant to the formation of and utilization in cancer treatment.

A Transient Mystery: Nucleolar Channel Systems.

The nucleus is a complex organelle with functions beyond being a simple repository for genomic material. For example, its actions in biomechanical sensing, protein synthesis, and epigenomic regulation showcase how the nucleus integrates multiple signaling modalities to intricately regulate gene expression. This innate dynamism is underscored by subnuclear components that facilitate these roles, with elementsof the nucleoskeleton, phase-separated nuclear bodies, and chromatin safeguarding by nuclear envelope proteins providingexamplesof this functional diversity. Among these, one of the lesser characterized nuclear organelles is the nucleolar channel system (NCS), first reported several decades ago inhuman endometrial biopsies. This tubular structure, believed to be derived from the inner nuclear membrane of the nuclear envelope, was first observed in secretory endometrial cells during a specific phase of the menstrual cycle. Reported as a consistent, yet transient, nuclear organelle, current interpretations of existing data suggest that it serves as a marker of a window for optimal implantation. In spite of this available data, the NCS remains incompletely characterized structurally and functionally, due in part to its transient spatial and temporal expression. As a further complication, evidence exists showing NCS expression in fetal tissue, suggesting that it may not act exclusively as a marker of uterine receptivity, but rather as a hormone sensor sensitive to estrogen and progesterone ratios. To gain a better understanding of the NCS, current technologies can be applied to profile rare cell populations or capture transient structural dynamics, for example, at a level of sensitivity and resolution not previously possible. Moving forward, advanced characterization of the NCS will shed light on an uncharacterized aspect of reproductive physiology, with the potential to refine assisted reproductive techniques.

УЛЬТРАСТРУКТУРНАЯ ПЕРЕСТРОЙКА МИОКАРДА НА 60-e СУТКИ ПОСЛЕ ДОКСОРУБИЦИН-ИНДУЦИРОВАННОЙ КАРДИОМИОПАТИИ У КРЫС

Background. To assess the cardiotoxic effects of doxorubicin, it is of great importance to elucidate the nature and severity of tissue and cell damage, their ultrastructural changes, as well as to elucidate the features of myocardial remodeling in such conditions. Objective. The aim of the study was a qualitative and quantitative analysis of ultrastructural changes in the myocardium on the 60th day after doxorubicin-induced cardiomyopathy in rats. Material and methods. Cardiomyopathy in rats was induced by doxorubicin given at a cumulative dose of 15 mg/kg for 14 days. The animals were removed from the experiment on the 60th day after last administered dose of the medicine. An electron microscopic research method was used. Ultrastructural analysis was performed using a JEM-100 CX microscope, and morphometric evaluation was done using the ImageJ data processing software. Results. Ultrastructural disorders of the myocardium of rats on the 60th day after doxorubicin-induced cardiomyopathy were characterized by: alterative changes of contractile apparatus and nuclear compartment of cardiomyocytes, activation of autophagic and necrotic cell death and compensatory mitochondria hyperplasia with T-system channels dilation; a variety of reactive changes of capillary endothelium in the form of nuclear hypertrophy and organelle hyperplasia in some endothelial cells along with intracellular edema and destruction of organelles in others, which was accompanied by a significant decrease in cross-sectional area of capillaries; activation of fibroblasts with a formation of interstitial and perivascular fibrosis; activation of autophagy processes in cardiomyocytes, fibroblastic cells and capillary endothelium. Conclusions. The established ultrastructural changes in the rat’s myocardium on the 60th day after the doxorubicin- induced cardiomyopathy indicate the complex effect of metabolic and angiogenic mechanisms of pathology development. Activation of autophagy processes plays an important role in myocardial reorganization in the late stages of doxorubicin myocardial damage.

The NF-κB Nucleolar Stress Response Pathway.

The nuclear organelle, the nucleolus, plays a critical role in stress response and the regulation of cellular homeostasis. P53 as a downstream effector of nucleolar stress is well defined. However, new data suggests that NF-κB also acts downstream of nucleolar stress to regulate cell growth and death. In this review, we will provide insight into the NF-κB nucleolar stress response pathway. We will discuss apoptosis mediated by nucleolar sequestration of RelA and new data demonstrating a role for p62 (sequestosome (SQSTM1)) in this process. We will also discuss activation of NF-κB signalling by degradation of the RNA polymerase I (PolI) complex component, transcription initiation factor-IA (TIF-IA (RRN3)), and contexts where TIF-IA-NF-κB signalling may be important. Finally, we will discuss how this pathway is targeted by aspirin to mediate apoptosis of colon cancer cells.

M6 A modification of HSATIII lncRNAs regulates temperature-dependent splicing.

Nuclear stress bodies (nSBs) are nuclear membraneless organelles formed around stress-inducible HSATIII architectural long noncoding RNAs (lncRNAs). nSBs repress splicing of hundreds of introns during thermal stress recovery, which are partly regulated by CLK1 kinase phosphorylation of temperature-dependent Ser/Arg-rich splicing factors (SRSFs). Here, we report a distinct mechanism for this splicing repression through protein sequestration by nSBs. Comprehensive identification of RNA-binding proteins revealed HSATIII association with proteins related to N6 -methyladenosine (m6 A) RNA modification. 11% of the first adenosine in the repetitive HSATIII sequence were m6 A-modified. nSBs sequester the m6 A writer complex to methylate HSATIII, leading to subsequent sequestration of the nuclear m6 A reader, YTHDC1. Sequestration of these factors from the nucleoplasm represses m6 A modification of pre-mRNAs, leading to repression of m6 A-dependent splicing during stress recovery phase. Thus, nSBs serve as a common platform for regulation of temperature-dependent splicing through dual mechanisms employing two distinct ribonucleoprotein modules with partially m6 A-modified architectural lncRNAs.

Environmentally induced ribosomal DNA (rDNA) instability in human cells and populations exposed to hexavalent chromium [Cr (VI)

Hexavalent Chromium [Cr (VI)] is an established toxicant, carcinogen, and a significant source of public health concern. The multicopy ribosomal DNA (rDNA) array is mechanistically linked to aging and cancer, is the most evolutionarily conserved segment of the human genome, and gives origin to nucleolus, a nuclear organelle where ribosomes are assembled. Here we show that exposure to Cr (VI) induces instability in the rDNA, triggering cycles of rapid, specific, and transient amplification and contraction of the array in human cells. The dynamic of environmentally responsive rDNA copy number (CN) amplification and contraction occurs at doses to which millions of individuals are regularly exposed. Finally, analyses of human populations occupationally exposed to Cr (VI) indicate that environmental exposure history and drinking habits but not age shape extensive naturally occurring rDNA copy number variation. Our observations identify a novel pathway of response to hexavalent chromium exposure and raise the prospect that a suite of environmental determinants of rDNA copy number remain to be discovered.

A first insight into the genome of Prototheca wickerhamii, a major causative agent of human protothecosis

BackgroundColourless microalgae of the Prototheca genus are the only known plants that have consistently been implicated in a range of clinically relevant opportunistic infections in both animals and humans. The Prototheca algae are emerging pathogens, whose incidence has increased importantly over the past two decades. Prototheca wickerhamii is a major human pathogen, responsible for at least 115 cases worldwide. Although the algae are receiving more attention nowadays, there is still a substantial knowledge gap regarding their biology, and pathogenicity in particular. Here we report, for the first time, the complete nuclear genome, organelle genomes, and transcriptome of the P. wickerhamii type strain ATCC 16529.ResultsThe assembled genome size was of 16.7 Mbp, making it the smallest and most compact genome sequenced so far among the protothecans. Key features of the genome included a high overall GC content (64.5%), a high number (6081) and proportion (45.9%) of protein-coding genes, and a low repetitive sequence content (2.2%). The vast majority (90.6%) of the predicted genes were confirmed with the corresponding transcripts upon RNA-sequencing analysis. Most (93.2%) of the genes had their putative function assigned when searched against the InterProScan database. A fourth (23.3%) of the genes were annotated with an enzymatic activity possibly associated with the adaptation to the human host environment. The P. wickerhamii genome encoded a wide array of possible virulence factors, including those already identified in two model opportunistic fungal pathogens, i.e. Candida albicans and Trichophyton rubrum, and thought to be involved in invasion of the host or elicitation of the adaptive stress response. Approximately 6% of the P. wickerhamii genes matched a Pathogen-Host Interaction Database entry and had a previously experimentally proven role in the disease development. Furthermore, genes coding for proteins (e.g. ATPase, malate dehydrogenase) hitherto considered as potential virulence factors of Prototheca spp. were demonstrated in the P. wickerhamii genome.ConclusionsOverall, this study is the first to describe the genetic make-up of P. wickerhamii and discovers proteins possibly involved in the development of protothecosis.

Noninvasive Measurement of Interphase Chromatin Rheology In Vivo

The material properties of the genome are critical for its proper function and organization inside the cell nucleus. Moreover, they directly impact all cellular processes via the central dogma of biology by setting the length and timescales of nuclear processes. Chromatin, the functional form of DNA in cells, consists of DNA and associated proteins, forming long linear fibers in the interphase nucleus of eukaryotic cells. Traditionally, the rheology of cellular components has been studied by tracking microparticles injected inside the cell [1]. Recently, we developed an injection-free noninvasive approach to study chromatin rheology using nuclear organelles as native probes [2]. Here, we show an alternative noninvasive experimental strategy using intrinsic dynamics to measure chromatin rheology across a large range of timescales, and elucidate the viscoelastic nature of chromatin in live cells. Furthermore, by spatially resolving the dynamics in different regions of the cell nucleus, our method reveals local rheology of the chromatin polymer. The measured rheology is captured by a surprisingly simple model whose few parameters have clear physical interpretations for this complex active material, and highlight the spatial and dynamical heterogeneity of the underlying chromatin organization. This work was supported by the National Institutes of Health Grant R00-GM104152, the National Science Foundation (NSF) Grants CAREER PHY-1554880 and CMMI-1762506, New York University (NYU) MRSEC NSF Grant DMR-1420073 and NYU Whitehead Fellowship for Junior Faculty in Biomedical and Biological Sciences (to AZ). 1. Wirtz, Annu. Rev. Biophys, 2009. 2. Caragine et. al., Phys. Rev. Lett., 2018.

Dissecting single-cell dynamics leveraging micro/nano electroporation

We here introduce approaches, SINC-seq (single-cell integrated nuclear and cytoplasmic RNA-sequencing) and NanoSINC-seq (nanopore-based SINC-seq), that allow us to study the correlated and anti-correlated expressions between cytoplasmic and nuclear transcripts in single cells at unprecedented resolution. We have developed microfluidic approaches that enable fractionation of cytoplasmic versus nuclear RNA in single cells, leveraging microflow and electric field control in a micro/nanometer scale. Our microfluidic approaches electrically and selectively lyse plasma membranes of eukaryotic cells while retaining nuclear membranes relatively intact, electrophoretically extract cytoplasmic components, physically fractionate cytoplasmic versus nuclear molecules and organelle, and automatically output the fractionated sample to individual off-chip analyses within 5 min. We demonstrated the integration of our approaches with off-chip next-generation sequencing to dissect the dynamical processes of post-transcriptional regulation.

Methoxyeugenol regulates the p53/p21 pathway and suppresses human endometrial cancer cell proliferation

Ethnopharmacological relevancePlant-derived compounds are a reservoir of natural chemicals and can act as drug precursors or prototypes and pharmacological probes. Methoxyeugenol is a natural compound found in plant extracts, such as nutmeg (Myristica fragrans), and it presents anthelmintic, antimicrobial, anti-inflammatory activities. Recently, interest in the anticancer activity of plant extracts is increasing and the therapeutic activity of methoxyeugenol against cancer has not yet been explored. Aim of the studyThe present study aimed to evaluate the cancer-suppressive role and the molecular signaling pathways of methoxyeugenol in human endometrial cancer (Ishikawa) cell line. Materials and methodsProliferation, viability, and cell toxicity were assessed by direct counting, MTT assay, and LDH enzyme release assay, respectively. Antiproliferative effect were evaluated by nuclear morphological changes along with the cellular mechanisms of apoptosis and senescence by flow cytometry. The underlying molecular and cellular mechanisms were investigated by RT-qPCR, reactive oxygen species (ROS) levels, mitochondrial dysfunction, and proliferative capacity. Results and conclusionsMethoxyeugenol treatment significantly inhibited the proliferation and viability of Ishikawa cells. Probably triggered by the higher ROS levels and mitochondrial dysfunction, the gene expression of p53 and p21 increased and the gene expression of CDK4/6 decreased in response to the methoxyeugenol treatment. The rise in nuclear size and acidic vesicular organelles corroborate with the initial senescence-inducing signals in Ishikawa cells treated with methoxyeugenol. The antiproliferative effect was not related to cytotoxicity and proved to effectively reduce the proliferative capacity of endometrial cancer cells even after treatment withdrawal. These results demonstrated that methoxyeugenol has a promising anticancer effect against endometrial cancer by rising ROS levels, triggering mitochondrial instability, and modulating cell signaling pathways leading to an inhibition of cell proliferation.