The nature of the complexes of histones H1 and H5 and their globular domains (GH1 and GH5) with DNA suggested two DNA‐binding sites which are likely to be the basis of the preference of H1 and H5 for the nucleosome, compared with free DNA. More recently the X‐ray and NMR structures of GH5 and GH1, respectively, have identified two basic clusters on opposite sides of the domains as candidates for these sites. Removal of the positive charge at either location by mutagenesis impairs or abolishes the ability of GH5 to assemble cooperatively in ‘tramline’ complexes containing two DNA duplexes, suggesting impairment or loss of its ability to bind two DNA duplexes. The mutant forms of GH5 also fail to protect the additional 20 bp of nucleosomal DNA that are characteristically protected by H1, H5 and wild‐type recombinant GH5. They still bind to H1/H5‐depleted chromatin, but evidently inappropriately. These results confirm the existence of, and identify the major components of, two DNA‐binding sites on the globular domain of histone H5, and they strongly suggest that both binding sites are required to position the globular domain correctly on the nucleosome.

Accurate gene expression requires the targeting of transcription factors (TFs) to regulatory sequences often occluded within nucleosomes. The ability to target a TF binding site (TFBS) within a nucleosome has been the defining characteristic for a special class of TFs known as pioneer factors. Recent studies suggest TP53 functions as a pioneer factor that can target its TFBS within nucleosomes, but it remains unclear how TP53 binds to nucleosomal DNA. To comprehensively examine TP53 nucleosome binding, we competitively bound TP53 to multiple in vitro–formed nucleosomes containing a high- or low-affinity TP53 TFBS located at differing translational and rotational positions within the nucleosome. Stable TP53–nucleosome complexes were isolated and quantified using next-generation sequencing. Our results demonstrate TP53 binding is limited to nucleosome edges with significant binding inhibition occurring within 50 bp of the nucleosome dyad. Binding site affinity only affects TP53 binding for TFBSs located at the same nucleosomal positions; otherwise, nucleosome position takes precedence. Furthermore, TP53 has strong nonspecific nucleosome binding facilitating its interaction with chromatin. Our in vitro findings were confirmed by examining TP53-induced binding in a cell line model, showing induced binding at nucleosome edges flanked by a nucleosome-free region. Overall, our results suggest that the pioneering capabilities of TP53 are driven by nonspecific nucleosome binding with specific binding at nucleosome edges.

It has not been possible to generate naïve human pluripotent stem cells (hPSCs) that substantially contribute to mouse embryos. We found that a brief inhibition of mTOR with Torin1 converted hPSCs from primed to naïve pluripotency. The naïve hPSCs were maintained in the same condition as mouse embryonic stem cells and exhibited high clonogenicity, rapid proliferation, mitochondrial respiration, X chromosome reactivation, DNA hypomethylation, and transcriptomes sharing similarities to those of human blastocysts. When transferred to mouse blastocysts, naïve hPSCs generated 0.1 to 4% human cells, of all three germ layers, including large amounts of enucleated red blood cells, suggesting a marked acceleration of hPSC development in mouse embryos. Torin1 induced nuclear translocation of TFE3; TFE3 with mutated nuclear localization signal blocked the primed-to-naïve conversion. The generation of chimera-competent naïve hPSCs unifies some common features of naïve pluripotency in mammals and may enable applications such as human organ generation in animals.

Background Older adults have been reported to be a population with high-risk of death in the COVID-19 outbreak. Rapid detection of high-risk patients is crucial to reduce mortality in this population. The aim of this study was to evaluate the prognositc accuracy of the Modified Early Warning Score (MEWS) for in-hospital mortality in older adults with COVID-19. Methods A retrospective cohort study was conducted in Wuhan Hankou Hospital in China from 1 January 2020 to 29 February 2020. Receiver operating characteristic (ROC) analysis was used to evaluate the predictive value of MEWS, Acute Physiology and Chronic Health Evaluation II (APACHE II), Sequential Organ Function Assessment (SOFA), quick Sequential Organ Function Assessment (qSOFA), Pneumonia Severity Index (PSI), Combination of Confusion, Urea, Respiratory Rate, Blood Pressure, and Age ≥65 (CURB-65), and the Systemic Inflammatory Response Syndrome Criteria (SIRS) for in-hospital mortality. Logistic regression models were performed to detect the high-risk older adults with COVID-19. Results Among the 235 patients included in this study, 37 (15.74%) died and 131 (55.74%) were male, with an average age of 70.61 years (SD 8.02). ROC analysis suggested that the capacity of MEWS in predicting in-hospital mortality was as good as the APACHE II, SOFA, PSI and qSOFA (Difference in AUROC: MEWS vs. APACHE II, −0.025 (95% CI [−0.075 to 0.026]); MEWS vs. SOFA, −0.013 (95% CI [−0.049 to 0.024]); MEWS vs. PSI, −0.015 (95% CI [−0.065 to 0.035]); MEWS vs. qSOFA, 0.024 (95% CI [−0.029 to 0.076]), all P > 0.05), but was significantly higher than SIRS and CURB-65 (Difference in AUROC: MEWS vs. SIRS, 0.218 (95% CI [0.156–0.279]); MEWS vs. CURB-65, 0.064 (95% CI [0.002–0.125]), all P < 0.05). Logistic regression models implied that the male patients (≥75 years) had higher risk of death than the other older adults (estimated coefficients: 1.16, P = 0.044). Our analysis further suggests that the cut-off points of the MEWS score for the male patients (≥75 years) subpopulation and the other elderly patients should be 2.5 and 3.5, respectively. Conclusions MEWS is an efficient tool for rapid assessment of elderly COVID-19 patients. MEWS has promising performance in predicting in-hospital mortality and identifying the high-risk group in elderly patients with COVID-19.

Background ΔNp63 is a master transcriptional regulator playing critical roles in epidermal development and other cellular processes. Recent studies suggest that ΔNp63 functions as a pioneer factor that can target its binding sites within inaccessible chromatin and induce chromatin remodeling. Methods In order to examine if ΔNp63 can bind to inaccessible chromatin and to determine if specific histone modifications are required for binding, we induced ΔNp63 expression in two p63-naïve cell lines. ΔNp63 binding was then examined by ChIP-seq and the chromatin at ΔNp63 targets sites was examined before and after binding. Further analysis with competitive nucleosome binding assays was used to determine how ΔNp63 directly interacts with nucleosomes. Results Our results show that before ΔNp63 binding, targeted sites lack histone modifications, indicating ΔNp63’s capability to bind at unmodified chromatin. Moreover, the majority of the sites that are bound by ectopic ΔNp63 expression exist in an inaccessible state. Once bound, ΔNp63 induces acetylation of the histone and the repositioning of nucleosomes at its binding sites. Further analysis with competitive nucleosome binding assays reveal that ΔNp63 can bind directly to nucleosome edges with significant binding inhibition occurring within 50 bp of the nucleosome dyad. Conclusion Overall, our results demonstrate that ΔNp63 is a pioneer factor that binds nucleosome edges at inaccessible and unmodified chromatin sites and induces histone acetylation and nucleosome repositioning.

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Treatment options for gestational diabetes (GDM) are limited. In order to better understand mechanisms and improve treatments, appropriate animal models of GDM are crucial. Heterozygous db mice (db/+) present with glucose intolerance, insulin resistance, and increased weight gain during, but not prior to, pregnancy. This makes them an ideal model for GDM. However, several recent studies have reported an absence of GDM phenotype in their colony. We investigated several hypotheses for why the phenotype may be absent, with the aim of re-establishing it and preventing further resources being wasted on an ineffective model. Experiments were carried out across two laboratories in two countries (New Zealand and China), and were designed to assess type of control strain, diet, presence of the misty allele, and parity as potential contributors to the lost phenotype. While hyperleptinemia and pre-pregnancy weight gain were present in all db/+mice across the four studies, we found no consistent evidence of glucose intolerance or insulin resistance during pregnancy. In conclusion, we were unable to acquire the GDM phenotype in any of our experiments, and we recommend researchers do not use the db/+ mouse as a model of GDM unless they are certain the phenotype remains in their colony.

The exosome is a complex with exoribonuclease activity that regulates RNA surveillance and turnover. The exosome also plays a role in regulating the degradation of precursor mRNAs to maintain the expression of splicing variants. In Neurospora, the silencing of rrp44, which encodes the catalytic subunit of the exosome, changed the expression of a set of spliceosomal snRNA, snRNP genes and SR protein related genes. The knockdown of rrp44 also affected the assembly of the spliceosome. RNA-seq analysis revealed a global change in bulk splicing events. Exosome-mediated splicing may regulate alternative splicing of NCU05290, NCU07421 and the circadian clock gene frequency (frq). The knockdown of rrp44 led to an increased ratio of splicing variants without intron 6 (I-6) and shorter protein isoform small FRQ (s-FRQ) as a consequence. These findings suggest that the exosome controls splicing events by regulating the degradation of precursor mRNAs and the gene expression, assembly and function of the spliceosome.