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Recently, Maxim A. Yurkin commented on our paper “Is the nuclear refractive index lower than cytoplasm? Validation of phase measurements and implications for light scattering technologies” as well as on a complementary study “Cell nuclei have lower refractive index and mass density than cytoplasm” from Schürmann et al. In his comment, Yurkin concluded that quantitative phase images of cells with nuclei that are less optically dense than the cytoplasm must exhibit a characteristic concavity, the absence of which is evidence against our conclusion of a less-dense nucleus. In this response, we suggest that Yurkin’s conclusion is reached through an oversimplification of the spatial refractive index distribution within cells, which does not account for high index inclusions such as the nucleolus. We further cite recent studies in 3-dimensional refractive index imaging, in which the preponderance of studies supports our conclusion. Finally, we comment on the current state of knowledge regarding subcellular refractive index distributions in living cells.

Circulating hemocytes from larval stages of the paper wasp Polistes dominulus were characterized by light and transmission electron microscopy. Three types were identified: prohemocytes, plasmatocytes and granulocytes. The first two are agranular cells while the latter present typical cytoplasmic inclusions called granules. Plasmatocytes differ from prohemocytes being larger, showing lower nucleus/cytoplasm ratio and they possess many phagolysosomes. The substantial uniformity of most subcellular features and the presence of "intermediate forms" support the "single-cell theory" i.e., there is only one cell line that originates from the prohemocyte and leads to the granular cell passing through the plasmatocyte. This hypothesis seems to be confirmed by functional tests. Indeed, most part of cells adheres to the glass and is able to phagocytize fluorescent microspheres.

Complex structures are generated and maintained through energy flux. Structures embody information, and biological information is stored in nucleic acids. The progressive increase in biological complexity over geologic time is thus the consequence of the information-generating power of energy flow plus the information-accumulating capacity of DNA, winnowed by natural selection. Consequently, the most important component of the biological environment is energy flow: the availability of calories and their use for growth, survival, and reproduction. Animals can exploit and adapt to available energy resources at three levels. They can evolve different anatomical forms through nuclear DNA (nDNA) mutations permitting exploitation of alternative energy reservoirs, resulting in new species. They can evolve modified bioenergetic physiologies within a species, primarily through the high mutation rate of mitochondrial DNA (mtDNA)–encoded bioenergetic genes, permitting adjustment to regional energetic environments. They can alter the epigenomic regulation of the thousands of dispersed bioenergetic genes via mitochondrially generated high-energy intermediates permitting individual accommodation to short-term environmental energetic fluctuations. Because medicine pertains to a single species, Homo sapiens, functional human variation often involves sequence changes in bioenergetic genes, most commonly mtDNA mutations, plus changes in the expression of bioenergetic genes mediated by the epigenome. Consequently, common nDNA polymorphisms in anatomical genes may represent only a fraction of the genetic variation associated with the common “complex” diseases, and the ascent of man has been the product of 3.5 billion years of information generation by energy flow, accumulated and preserved in DNA and edited by natural selection.

More than 30 years of research on nuclear RNA polymerases (RNAP I, II, and III) has uncovered numerous factors that regulate the activity of these enzymes during the transcription reaction. However, very little is known about the machinery that regulates the fate of RNAPs before or after transcription. In particular, the mechanisms of biogenesis of the 3 nuclear RNAPs, which comprise both common and specific subunits, remains mostly uncharacterized and the proteins involved are yet to be discovered. Using protein affinity purification coupled to mass spectrometry (AP–MS), we recently unraveled a high-density interaction network formed by nuclear RNAP subunits from the soluble fraction of human cell extracts. Validation of the dataset using a machine learning approach trained to minimize the rate of false positives and false negatives yielded a high-confidence dataset and uncovered novel interactors that regulate the RNAP II transcription machinery, including a set of proteins we named the RNAP II-associated proteins (RPAPs). One of the RPAPs, RPAP3, is part of an 11-subunit complex we termed the RPAP3/R2TP/prefoldin-like complex. Here, we review the literature on the subunits of this complex, which points to a role in nuclear RNAP biogenesis.

Summary Topological cytonuclear discordance is commonly observed in plant phylogenetic and phylogeographic studies, yet few studies have attempted to detect two other forms of cytonuclear discordance (branch length and geographical) and to uncover the causes of the discordance.We used the whole nuclear and chloroplast genome data from 80 individual Asian butternuts to reveal the pattern and processes of cytonuclear discordance.Our findings indicate that the chloroplast genome had substantially deeper divergence (branch‐length discordance) and a steeper cline in the contact zone (geographic discordance) compared with the nuclear genome. After various hypothesis have been tested, the results suggest that incomplete lineage sorting, positive selection and cytonuclear incompatibility are probably insufficient to explain this pattern. However, isolation‐by‐distance analysis and gene flow estimation point to a much higher level of gene flow by pollen compared with by seeds, which may have slowed down lineage divergence and mediated wider contact for nuclear genome compared with the chloroplast genome.Altogether, this study highlights a critical role of sex‐biased dispersal in causing discordance between the nuclear and plastid genome of Asian butternuts. Given its ubiquity among plants, asymmetric gene flow should be given a high priority in future studies of cytonuclear discordance.

Osteoarthritis (OA) is a cartilage degenerative disease commonly observed in the elderly population and is pathologically characterized by the degradation of the cartilage extracellular matrix (ECM). Matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTSs) are critical enzymes involved in the degradation of ECM. Olmesartan is an inhibitor of the angiotensin II receptor developed for the treatment of hypertension, and recent studies show that it exerts anti-inflammatory effects in arthritis. The present study aimed to investigate the mechanism of the protective effect of Olmesartan on cartilage ECM degradation. Interleukin-29 (IL-29) is a novel inflammatory mediator involved in the inflammation and degradation of cartilage in OA, and human T/C-28a2 cells treated with it were the inflammatory model in vitro. We found that the degradation of type 2 collagens and aggrecans was induced by IL-29, accompanied by the upregulation of MMPs and ADAMTSs, but the presence of Olmesartan significantly ameliorated these increases. In addition, Olmesartan abolished IL-29- induced oxidative stress and elevated the expression level of TNF receptor-associated factor 6 (TRAF-6). Mechanistically, we showed that Olmesartan suppressed IL-29- caused inhibitor kappa B α (IκBα) expression and nuclear translocation of nuclear factor kappa-B (NF-κB) p65, indicating it suppressed the activation of the NF-κB pathway. Collectively, our data reveal that Olmesartan exerted a protective function on IL-29- induced type 2 collagen degradation in human chondrocytes.

Accumulating evidence suggests that long non-coding RNAs (lncRNAs) participate in the formation and development of keloids, a benign tumor. In addition, lncRNA H19 has been shown to act on the biological processes of keloids. This study aimed to identify other important mechanisms of the effect of lncRNA H19 on keloid formation. The H19, miR-196b-5p, and SMAD family member 5 (SMAD5) expression levels were detected using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and Western blotting. Subcellular localization of lncRNA H19 was detected using a nuclear–cytoplasmic separation assay. Cell viability and proliferation were measured using counting kit-8 and colony formation assays. Bax and Bcl-2 levels were examined using Western blot analysis. The interaction between H19 and miR-196b-5p or SMAD5 was verified using a dual-luciferase reporter assay. H19 and SMAD5 expression was upregulated in keloid tissue and fibroblasts, whereas miR-196b-5p expression was downregulated. Knockdown of H19, overexpression of miR-196b-5p, or knockdown of SMAD5 inhibited the viability and proliferation of keloid fibroblasts and promoted apoptosis. Overexpression of H19 or SMAD5 and knockdown of miR-196b-5p promoted viability and proliferation and inhibited apoptosis. miR-196b-5p was identified as a H19 sponge, and SMAD5 was identified as a miR-196b-5p target. The combination of lncRNA H19 and miR-196b-5p regulates SMAD5 expression and promotes keloid formation, thus providing a new direction for keloid treatment.

Dexmedetomidine (DEX) has been reported to attenuate cecal ligation perforation (CLP)-stimulated acute lung injury (ALI) by downregulating HMGB1 and RAGE. This study aimed to further investigate the specific mechanisms of RAGE and its potential-related mechanisms of DEX on ALI models in vitro and in vivo. The in vitro and in vivo ALI models were established by lipopolysaccharide treatment in MLE-12 cells and CLP in mice, respectively. The effect of DEX on pathological alteration was investigated by HE staining. Thereafter, the myeloperoxidase (MPO) activity and inflammatory cytokine levels were respectively detected to assess the lung injury of mice using commercial kits. The expression levels of HMGB1, RAGE, NF-κB, and pyroptosis-related molecules were detected by RT-qPCR and Western blot. HE staining showed that lung injury, increased inflammatory cell infiltration, and lung permeability was found in the ALI mice, and DEX treatment significantly attenuated lung tissue damage induced by CLP. The MPO activity and inflammatory cytokines (TNF-α, IL-1β, and NLRP3) levels were also significantly reduced after DEX treatment compared with those in the ALI mice. Moreover, DEX activated the HMGB1/RAGE/NF-κB pathway and upregulated the pyroptosis-related proteins. However, the protective DEX effect was impaired by RAGE overexpression in ALI mice and MLE-12 cells. Additionally, DEX treatment significantly suppressed HMGB1 translocation from the nucleus region to the cytoplasm, and this effect was reversed by RAGE overexpression. These findings suggested that DEX may be a useful ALI treatment, and the protective effects on ALI mice may be through the inhibition of HMGB1/RAGE/NF-κB pathway and cell pyroptosis., Graphical abstractDexmedetomidine (DEX) has protective effects on acute lung injury (ALI) in vitro and in vivo. The possible mechanisms may be closely associated with inflammatory response, caspase-1-mediated cell pyroptosis, and high-mobility group protein 1 (HMGB1)/receptor for advanced glycation end products (RAGE)/nuclear factor-κB pathway. Moreover, DEX could promote the HMGB1 translocation from the cytoplasm to the nucleus in lipopolysaccharide-activated MLE-12 cells, whereas the action of RAGE overexpression was opposite.

Curcumin suppressed ultraviolet (UV) induced skin carcinogenesis and activated the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. However, whether curcumin protects skin injury caused by UV is still unknown. A vitro model was established and curcumin effects on Hacat cells were detected. Nrf2 was knocked down in Hacat cells to verify the Nrf2 role in the protective effect of curcumin. Results indicated that ultraviolet A (UVA) (or ultraviolet B (UVB)) irradiation would lead to decreased cell proliferation, increased cell apoptosis, decreased catalase, heme oxygenase 1, and superoxide dismutase expression, and increased levels of protein carbonylation and malondialdehyde (p

Subcellular localization of transcripts is highly associated with regulation of gene expression, synthesis of protein, and also the development of the human brain cortex. Although many mechanisms are prevalent in the occurrence of neuroinflammation, the mechanisms based on differences in subcellular localization of transcripts have not been explored. To characterize the dynamic profile of nuclear and cytoplasmic transcripts during the progress of haemorrhage-induced neuroinflammation, we isolated nucleo-cytoplasmic RNA fractions of oxyhaemoglobin (oxy-Hb) treated microglia cells and sequenced both fractions. We discovered that cytoplasmic retained genes were the major forces to maintain the neuroinflammatory microenvironment with 10 hub genes and 40 conserved genes were identified. Moreover, antisense RNA Gm44096 and lincRNA Gm47270, which co-expressed with a crowd of inflammatory genes in the cytoplasm, were discovered as regulatory strategies for sustaining the neuroinflammatory microenvironment. Thus, our study provides a new perspective on understanding haemorrhage-induced neuroinflammation and also reveals a mechanism of lncRNA responsible for maintaining the neuroinflammatory microenvironment.

Super-resolution microscopy revealed that the plant nuclear lamina proteins NMCP1 and NMCP2 organize into filamentous networks with filament bundles., Plant genomes lack genes encoding intermediate filament proteins, including lamins; however, functional lamin analogues are presumed to exist in plants. Plant-specific coiled-coil proteins, that is, nuclear matrix constituent proteins (NMCPs), are the most likely candidates as the structural elements of the nuclear lamina because they exhibit a lamin-like domain arrangement. They are exclusively localized at the nuclear periphery and have functions that are analogous to those of lamins. However, their assembly into filamentous polymers has not yet been confirmed. In this study, we examined the higher-order structure of NMCP1 and NMCP2 in Apium graveolens cells by using stimulated emission depletion microscopy combined with immunofluorescence cell labelling. Our analyses revealed that NMCP1 and NMCP2 form intricate filamentous networks, which include thick segments consisting of filament bundles, forming a dense filamentous layer extending across the nuclear periphery. Furthermore, the outermost chromatin distribution was found to be in the nucleoplasm-facing region of the nuclear lamina. Recombinant Daucus carota NMCP1 with a His-tag produced in Escherichia coli refolded into dimers and self-assembled into filaments and filament bundles. These results suggest that NMCP1 and NMCP2 organize into the nuclear lamina by forming a filamentous network with filament bundles that localize at the nuclear periphery.

Expansion microscopy (ExM) has been successfully used to improve the spatial resolution when imaging tissues by optical microscopy. In ExM, proteins of a fixed sample are crosslinked to a swellable acrylamide gel, which expands when incubated in water. Therefore, ExM allows enlarged subcellular structures to be resolved that would otherwise be hidden to standard confocal microscopy. Herein, we aim to validate ExM for the study of peroxisomes, mitochondria, nuclei and the plasma membrane. Upon comparison of the expansion factors of these cellular compartments in HEK293 cells within the same gel, we found significant differences, of a factor of above 2, in expansion factors. For peroxisomes, the expansion factor differed even between peroxisomal membrane and matrix marker; this underlines the need for a thorough validation of expansion factors of this powerful technique. We further give an overview of possible quantification methods for the determination of expansion factors of intracellular organelles, and we highlight some potentials and challenges., In Expansion Microscopy (ExM) subcellular structures are imaged in isotropically expanded fixed samples, consequently allowing to resolve enlarged subcellular structures, otherwise hidden to standard microscopy. Upon comparison of the expansion factors of different cellular compartments in cells within the same gel, we found significant differences in expansion factors of a factor of above 2.

The vitamin D receptor (VDR) and aryl hydrocarbon receptor (AHR) are two nuclear receptors that exert their effects by binding with ligands and forming a molecular complex. These complexes translocate to the nucleus and activate the expression of a series of genes which have a response element to VDR or AHR. Both receptors have been identified in the pathogenesis of endometriosis, a common disease characterized by the formation of endometrium-like tissue in ectopic zones. Despite numerous therapies, there is no definitive cure for endometriosis at the pharmacological level. Our study aims to describe the location and the expression of VDR and AHR at the protein level. For this purpose, an evaluation was performed using tissue from the three normal phases of the endometrium (proliferative, early, and late secretory) and in endometriosis by immunohistochemistry, using anti-VDR and anti-AHR antibodies. We demonstrate that in the nuclei of glandular cells in endometriosis, the expression of VDR and AHR is mutually exclusive—when the expression of one receptor is high, the other one is low—suggesting a possible target in the treatment of endometriosis. We also identify a significant change in the expression of glandular cytoplasmic AHR between the proliferative and late secretory endometrium. Supplementary Information The online version contains supplementary material available at 10.1007/s00418-021-02005-9.

The occurrence of hypoxia-ischemia (HI) in the developing brain is closely associated with neuronal injury and even death. However, the underlying molecular mechanism is not fully understood. This study was designed to investigate phosphatase and tensin homolog (PTEN) nuclear translocation and its possible role in rat cortical neuronal damage following oxygen-glucose deprivation (OGD) in vitro. An in vitro OGD model was established using primary cortical neurons dissected from newborn Sprague-Dawley rats to mimic HI conditions. The PTENK13R mutant plasmid, which contains a lysine-to-arginine mutation at the lysine 13 residue, was constructed. The nuclei and cytoplasm of neurons were separated. Neuronal injury following OGD was evidenced by increased lactate dehydrogenase (LDH) release and apoptotic cell counts. In addition, PTEN expression was increased and the phosphorylation of extracellular signal-regulated kinase 1/2 (p-ERK1/2) and activation of nuclear factor kappa B (NF-κB) were decreased following OGD. PTENK13R transfection prevented PTEN nuclear translocation; attenuated the effect of OGD on nuclear p-ERK1/2 and NF-κB, apoptosis, and LDH release; and increased the expression of several anti-apoptotic proteins. We conclude that PTEN nuclear translocation plays an essential role in neuronal injury following OGD via modulation of the p-ERK1/2 and NF-κB pathways. Prevention of PTEN nuclear translocation might be a candidate strategy for preventing brain injury following HI.

Cataract is the leading cause of visual impairment globally. Racemization of lens proteins may contribute to cataract formation in aging individuals. As a special type of age-related cataract (ARC), diabetic cataract (DC) is characterized by the early onset of cortical opacification and finally developed into a mixed type of cortical and nuclear opacification. We compared racemization of Asp 58 residue, a hotspot position in αA-crystallin, from the cortex and nucleus of diabetic and age-matched senile cataractous lenses, by identifying L-Asp/L-isoAsp/D-Asp/D-isoAsp by mass spectrometry. Compared to nondiabetic cataractous lenses, DC lenses showed a significantly increased cortex/nucleus ratio of D-Asp 58, which originated primarily from an increased percentage of D-Asp 58 in the lens cortex of DC. Moreover, patients diagnosed with diabetes for over 10 years showed a lower cortex/nucleus ratio of D-isoAsp 58 in the lens compared with those who had a shorter duration of diabetes, which originated mainly from an increased percentage of D-isoAsp 58 in the lens nucleus of DC with increasing time of hyperglycemia. Further analysis confirmed decreased protein solubility in diabetic cataractous lenses. The different racemization pattern in DC may be distinguished from ARC and influence its phenotype over the protracted duration of diabetes.

cDNA microarray data conducted by our group revealed overexpression of CXCL2 and CXCL8 in ovarian cancer (OC) microenvironment. Herein, we have proven that the chemokine receptor, CXCR2, is a pivotal molecule in re-sensitizing OC to cisplatin, and its inhibition decreases cell proliferation, viability, tumor size in cisplatin-resistant cells, as well as reversed the overexpression of mesenchymal epithelium transition markers. Altogether, our study indicates a central effect of CXCR2 in preventing tumor progression, due to acquisition of cisplatin chemoresistant phenotype by tumor cells, and patients’ high lethality rate. We found that the overexpression of CXCR2 by OC cells is persistent and anomalously confined to the cellular nuclei, thus pointing to an urge in developing highly lipophilic molecules that promptly permeate cells, bind to and inhibit nuclear CXCR2 to fight OC, instead of relying on the high-cost genetic engineered cells.

Suppression of type I interferon (IFN) response is one pathological outcome of the infection of highly pathogenic human coronaviruses. To effect this, severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2 encode multiple IFN antagonists. In this study, we reported on the IFN antagonism of SARS-CoV-2 main protease NSP5. NSP5 proteins of both SARS-CoV and SARS-CoV-2 counteracted Sendai virus-induced IFN production. NSP5 variants G15S and K90R commonly seen in circulating strains of SARS-CoV-2 retained the IFN-antagonizing property. The suppressive effect of NSP5 on IFN-β gene transcription induced by RIG-I, MAVS, TBK1 and IKKϵ suggested that NSP5 likely acts at a step downstream of IRF3 phosphorylation in the cytoplasm. NSP5 did not influence steady-state expression or phosphorylation of IRF3, suggesting that IRF3, regardless of its phosphorylation state, might not be the substrate of NSP5 protease. However, nuclear translocation of phosphorylated IRF3 was severely compromised in NSP5-expressing cells. Taken together, our work revealed a new mechanism by which NSP5 proteins encoded by SARS-CoV and SARS-CoV-2 antagonize IFN production by retaining phosphorylated IRF3 in the cytoplasm. Our findings have implications in rational design and development of antiviral agents against SARS-CoV-2.

Cellular senescence is the irreversible cell cycle arrest in response to DNA damage. Because senescent cells accumulate with age and contribute to chronic inflammation, they are promising therapeutic targets for healthspan extension. The senescent phenotype can vary depending on cell type and on the specific insults that induce senescence. This variability is also reflected in the extensive remodeling of the genome organization within the nucleus of senescent cells. Here, we give an overview of the nuclear changes that occur in different forms of senescence, including changes to chromatin state and composition and to the three-dimensional organization of the genome, as well as alterations to the nuclear envelope and to the accessibility of repetitive genomic regions. Many of these changes are shared across all forms of senescence, implicating nuclear organization as a fundamental driver of the senescent state and of how senescent cells interact with the surrounding tissue.