DNA polymerase delta governs parental histone transfer to DNA replication lagging strand.

Chromatin replication is intricately intertwined with the recycling of parental histones to the newly duplicated DNA strands for faithful genetic and epigenetic inheritance. The transfer of parental histones occurs through two distinct pathways: leading strand deposition, mediated by the DNA polymerase ε subunits Dpb3/Dpb4, and lagging strand deposition, facilitated by the MCM helicase subunit Mcm2. However, the mechanism of the facilitation of Mcm2 transferring parental histones to the lagging strand while moving along the leading strand remains unclear. Here, we show that the deletion of Pol32, a nonessential subunit of major lagging-strand DNA polymerase δ, results in a predominant transfer of parental histone H3-H4 to the leading strand during replication. Biochemical analyses further demonstrate that Pol32 can bind histone H3-H4 both in vivo and in vitro. The interaction of Pol32 with parental histone H3-H4 is disrupted through the mutation of the histone H3-H4 binding domain within Mcm2. Our findings identify the DNA polymerase δ subunit Pol32 as a critical histone chaperone downstream of Mcm2, mediating the transfer of parental histones to the lagging strand during DNA replication.

Defective transfer of parental histone decreases frequency of homologous recombination by increasing free histone pools in budding yeast.

Recycling of parental histones is an important step in epigenetic inheritance. During DNA replication, DNA polymerase epsilon subunit DPB3/DPB4 and DNA replication helicase subunit MCM2 are involved in the transfer of parental histones to the leading and lagging strands, respectively. Single Dpb3 deletion (dpb3Δ) or Mcm2 mutation (mcm2-3A), which each disrupts one parental histone transfer pathway, leads to the other's predominance. However, the biological impact of the two histone transfer pathways on chromatin structure and DNA repair remains elusive. In this study, we used budding yeast Saccharomyces cerevisiae to determine the genetic and epigenetic outcomes from disruption of parental histone H3-H4 tetramer transfer. We found that a dpb3Δ mcm2-3A double mutant did not exhibit the asymmetric parental histone patterns caused by a single dpb3Δ or mcm2-3A mutation, suggesting that the processes by which parental histones are transferred to the leading and lagging strands are independent. Surprisingly, the frequency of homologous recombination was significantly lower in dpb3Δ, mcm2-3A and dpb3Δ mcm2-3A mutants, likely due to the elevated levels of free histones detected in the mutant cells. Together, these findings indicate that proper transfer of parental histones during DNA replication is essential for maintaining chromatin structure and that lower homologous recombination activity due to parental histone transfer defects is detrimental to cells.

FSH preserves the viability of hypoxic granulosa cells via activating the HIF-1α-GAS6-Axl-Akt pathway.

The developmental fate of ovarian follicles is primarily determined by the survival status (proliferation or apoptosis) of granulosa cells (GCs). Owing to the avascular environment within follicles, GCs are believed to live in a hypoxic niche. Follicle-stimulating hormone (FSH) has been reported to improve GCs survival by governing hypoxia-inducible factor-1α (HIF-1α)-dependent hypoxia response, but the underlying mechanisms remain poorly understood. Growth arrest-specific gene 6 (GAS6) is a secreted ligand of tyrosine kinase receptors, and has been documented to facilitate tumor growth. Here, we showed that the level of GAS6 was markedly increased in mouse ovarian GCs after the injection of FSH. Specifically, FSH-induced GAS6 expression was accompanied by HIF-1α accumulation under conditions of hypoxia both in vivo and in vitro, whereas inhibition of HIF-1α with small interfering RNAs/antagonist repressed both expression and secretion of GAS6. As such, Luciferase reporter assay and chromatin immunoprecipitation assay showed that HIF-1α directly bound to a hypoxia response element site within the Gas6 promoter and contributed to the regulation of GAS6 expression in response to FSH. Notably, blockage of GAS6 and/or its receptor Axl abrogated the pro-survival effects of FSH under hypoxia. Moreover, phosphorylation of Axl by GAS6 is required for FSH-mediated Akt activation and the resultant pro-survival phenotypes. Finally, the in vitro findings were verified in vivo, which showed that FSH-induced proliferative and antiapoptotic effects in ovarian GCs were diminished after blocking GAS6/Axl using HIF-1α antagonist. These findings highlight a novel function of FSH in preserving GCs viability against hypoxic stress by activating the HIF-1a-GAS6-Axl-Akt pathway.

Low repetition rate 915†nm figure-9 ultrafast laser with all-fiber structure.

The advent of optical metrology applications has necessitated the development of compact, reliable, and cost-effective picosecond lasers operating around 900 nm, specifically catering to the requirements of precise ranging. In response to this demand, our work introduces an innovative solution-an all-fiber, all-polarization-maintaining (PM) figure-9 mode-locked laser operating at 915 nm. The proposed figure-9 Nd-doped fiber laser has a 69.2 m long cavity length, strategically designed and optimized to yield pulses with a combination of high pulse energy and low repetition rate. The laser can generate 915 nm laser pulses with a pulse energy of 4.65 nJ, a pulse duration of 15.2 ps under the repetition rate of 3.05 MHz. The 1064 nm amplified spontaneous emission (ASE) is deliberately filtered out, in order to prevent parasitic lasing and increase the spectral proportion of the 915 nm laser. The all-PM fiber configuration of this laser imparts exceptional mode-locking performance and environmental robustness, which is confirmed by long-term output power and spectral stability test. This compact and long-term reliable fiber laser could be a promising light source for applications like inter-satellite ranging.

SP1 impacts the primordial to primary follicle transition by regulating cholesterol metabolism in granulosa cells.

The primordial to primary follicle transition (PPT) in the ovary is critical to maintain sustainable reproductive resources in female mammals. However, it is unclear how granulosa cells (GCs) of the primary follicle participate in regulating PPT. This study focused on exploring the role of transcription factor Sp1 (SP1) in regulating PPT based on the fact that SP1 is pivotal for pregranulosa cell proliferation before primordial follicle formation. The results showed that mice fertility was prolonged when Sp1 was specifically depleted from GCs (GC- Sp1 -/- ). Besides, the PPT in GC- Sp1 -/- mice was reduced, resulting in more primordial follicles being preserved. Single-cell RNA-seq also indicated that the level of cholesterol metabolism was downregulated in GC- Sp1 -/- mice. Additionally, the PPT was promoted by either overexpression of ferredoxin-1 (FDX1), one of the key genes in mediating cholesterol metabolism or supplementing cholesterol for cultured fetal ovaries. Collectively, SP1 in GCs participates in the metabolism of cholesterol partially by regulating the transcription of Fdx1 during the PPT.

A higher dysregulation burden of brain DNA methylation in female patients implicated in the sex bias of Schizophrenia.

Sex differences are pervasive in schizophrenia (SCZ), but the extent and magnitude of DNA methylation (DNAm) changes underlying these differences remain uncharacterized. In this study, sex-stratified differential DNAm analysis was performed in postmortem brain samples from 117 SCZ and 137 controls, partitioned into discovery and replication datasets. Three differentially methylated positions (DMPs) were identified (adj.p < 0.05) in females and 29 DMPs in males without overlap between them. Over 81% of these sex-stratified DMPs were directionally consistent between sexes but with different effect sizes. Females experienced larger magnitude of DNAm changes and more DMPs (based on data of equal sample size) than males, contributing to a higher dysregulation burden of DNAm in females SCZ. Additionally, despite similar proportions of female-related DMPs (fDMPs, 8%) being under genetic control compared with males (10%), significant enrichment of DMP-related single nucleotide polymorphisms (SNPs) in signals of genome-wide association studies was identified only in fDMPs. One DMP in each sex connected the SNPs and gene expression of CALHM1 in females and CCDC149 in males. PPI subnetworks revealed that both female- and male-related differential DNAm interacted with synapse-related dysregulation. Immune-related pathways were unique for females and neuron-related pathways were associated with males. This study reveals remarkable quantitative differences in DNAm-related sexual dimorphism in SCZ and that females have a higher dysregulation burden of SCZ-associated DNAm than males.

Neuroimmune transcriptome changes in patient brains of psychiatric and neurological disorders.

Neuroinflammation has been implicated in multiple brain disorders but the extent and the magnitude of change in immune-related genes (IRGs) across distinct brain disorders has not been directly compared. In this study, 1275 IRGs were curated and their expression changes investigated in 2467 postmortem brains of controls and patients with six major brain disorders, including schizophrenia (SCZ), bipolar disorder (BD), autism spectrum disorder (ASD), major depressive disorder (MDD), Alzheimer's disease (AD), and Parkinson's disease (PD). There were 865 IRGs present across all microarray and RNA-seq datasets. More than 60% of the IRGs had significantly altered expression in at least one of the six disorders. The differentially expressed immune-related genes (dIRGs) shared across disorders were mainly related to innate immunity. Moreover, sex, tissue, and putative cell type were systematically evaluated for immune alterations in different neuropsychiatric disorders. Co-expression networks revealed that transcripts of the neuroimmune systems interacted with neuronal-systems, both of which contribute to the pathology of brain disorders. However, only a few genes with expression changes were also identified as containing risk variants in genome-wide association studies. The transcriptome alterations at gene and network levels may clarify the immune-related pathophysiology and help to better define neuropsychiatric and neurological disorders.

Discovery of novel HER2 targeting peptide-camptothecin conjugates with effective suppression for selective cancer treatment.

Due to the strong selectivity and permeability of tumor tissue, anti-cancer peptide-drug conjugates (PDCs) can accumulate high concentration of toxic payloads at the target, effectively killing tumor cells. This approach holds great promise for tumor-targeted treatment. In our previous study, we identified the optimal peptide P1 (NPNWGRSWYNQRFK) targeting HER2 from pertuzumab, a monoclonal antibody that blocks the HER2 signaling pathway. Here, a series of PDCs were constructed through connecting P1 and CPT with different linkers. Among these, Z8 emerged as the optimal compound, demonstrating good antitumor activity and targeting ability in biological activity tests. Z8 exhibited IC50 values of 1.04 ± 0.24 µM and 1.91 ± 0.71 µM against HER2-positive SK-BR-3 and NCI-N87 cells, respectively. Moreover, superior antitumor activity and higher biosafety of Z8 were observed compared to the positive control CPT in vivo, suggesting a novel idea for the construction of PDCs.

Alcohol reshapes a liver premetastatic niche for cancer by extra- and intrahepatic crosstalk-mediated immune evasion.

Cancer metastatic organotropism is still a mystery. The liver is known to be susceptible to cancer metastasis and alcoholic injury. However, it is unclear whether and how alcohol facilitates liver metastasis and how to intervene. Here, we show that alcohol preferentially promotes liver metastasis in colon-cancer-bearing mice and post-surgery pancreatic cancer patients. The mechanism is that alcohol triggers an extra- and intrahepatic crosstalk to reshape an immunosuppressive liver microenvironment. In detail, alcohol upregulates extrahepatic IL-6 and hepatocellular IL-6 receptor expression, resulting in hepatocyte STAT3 signaling activation and downstream lipocalin-2 (Lcn2) upregulation. Furthermore, LCN2 promotes T cell-exhaustion neutrophil recruitment and cancer cell epithelial plasticity. In contrast, knocking out hepatocellular Stat3 or systemic Il6 in alcohol-treated mice preserves the liver microenvironment and suppresses liver metastasis. This mechanism is reflected in hepatocellular carcinoma patients, in that alcohol-associated signaling elevation in noncancerous liver tissue indicates adverse prognosis. Accordingly, we discover a novel application for BBI608, a small molecular STAT3 inhibitor that can prevent liver metastasis. BBI608 pretreatment protects the liver and suppresses alcohol-triggered premetastatic niche formation. In conclusion, under extra- and intrahepatic crosstalk, the alcoholic injured liver forms a favorable niche for cancer cell metastasis, while BBI608 is a promising anti-metastatic agent targeting such microenvironments.

A mechanism for Rad53 to couple leading- and lagging-strand DNA synthesis under replication stress in budding yeast.

In response to DNA replication stress, DNA replication checkpoint kinase Mec1 phosphorylates Mrc1, which in turn activates Rad53 to prevent the generation of deleterious single-stranded DNA, a process that remains poorly understood. We previously reported that lagging-strand DNA synthesis proceeds farther than leading strand in rad53-1 mutant cells defective in replication checkpoint under replication stress, resulting in the exposure of long stretches of the leading-strand templates. Here, we show that asymmetric DNA synthesis is also observed in mec1-100 and mrc1-AQ cells defective in replication checkpoint but, surprisingly, not in mrc1∆ cells in which both DNA replication and checkpoint functions of Mrc1 are missing. Furthermore, depletion of either Mrc1 or its partner, Tof1, suppresses the asymmetric DNA synthesis in rad53-1 mutant cells. Thus, the DNA replication checkpoint pathway couples leading- and lagging-strand DNA synthesis by attenuating the replication function of Mrc1-Tof1 under replication stress.

Redistribution of Epithelium and Stroma and Their Relationship With Corneal Curvature After 1-Month Cessation of Overnight Orthokeratology.

OBJECTIVES: To investigate the changes in the thickness of epithelium and stroma and their relationship with corneal curvature following the cessation of overnight orthokeratology for a period of 1 month. METHODS: This prospective study consecutively included 20 juveniles (20 right eyes) who had undergone overnight orthokeratology for a minimum of one year and were willing to discontinue the treatment. The study measured and compared epithelial and corneal curvature using optical coherence tomography and Medmont topographer at the first day of cessation and 1 month after cessation. In addition, changes in uncorrected visual acuity and refractive error before and after the cessation of the treatment were analyzed. RESULTS: The study found a significant increase in the thickness of the epithelium in the central 2-mm area after the cessation of the treatment (t = -4.807, P <0.001). Moreover, the stroma in the paracentral area (2-5 mm) and peripheral area (5-6 mm) showed a general thinning trend ( P =0.016, P =0.016). Regarding the correlation analysis, the change in central epithelial thickness (ΔCET) was significantly correlated with the change in paracentral corneal curvature (ΔPCCC) (r=0.610, P =0.007) and the change in peripheral corneal curvature (ΔPCC) (r=0.597, P =0.009). Similarly, the change in central stromal thickness (ΔCST) was significantly correlated with the change in central corneal curvature (ΔCCC) (r=0.500, P =0.035), ΔPCCC (r=0.700, P =0.001), and ΔPCC (r=0.635, P =0.005). CONCLUSIONS: The study found that the corneal remodeling induced by orthokeratology was reversible after the cessation of the treatment. Specifically, changes in the epithelium were found to be more prominent in the central area, while changes in the stroma were more pronounced in the paracentral and peripheral areas. In addition, the study established a significant correlation between central corneal remodeling and changes in curvature.

Structural insights into the Oryza sativa cation transporters HKTs in salt tolerance.

The high-affinity potassium transporters (HKTs), selectively permeable to either Na+ alone or Na+/K+, play pivotal roles in maintaining plant Na+/K+ homeostasis. Although their involvement in salt tolerance is widely reported, the molecular underpinnings of Oryza sativa HKTs remain elusive. In this study, we elucidate the structures of OsHKT1;1 and OsHKT2;1, representing two distinct classes of rice HKTs. The dimeric assembled OsHKTs can be structurally divided into four domains. At the dimer interface, a half-helix or a loop in the third domain is coordinated by the C-terminal region of the opposite subunit. Additionally, we present the structures of OsHKT1;5 salt-tolerant and salt-sensitive variants, a key quantitative trait locus associated with salt tolerance. The salt-tolerant variant of OsHKT1;5 exhibits enhanced Na+ transport capability and displays a more flexible conformation. These findings shed light on the molecular basis of rice HKTs and provide insights into their role in salt tolerance.

Integrated genetic analysis of diabetic complications: Bioinformatics insights into foot ulcers, neuropathy and peripheral artery disease.

Diabetic foot ulcers (DFU), diabetic peripheral neuropathy (DPN) and peripheral arterial disease (PAD) are common complications of diabetes mellitus, while diabetic peripheral neuropathy and peripheral arterial disease contribute to the pathogenesis of diabetic foot ulcers, and the pathogenic mechanisms between these three diseases still need further investigation. The keywords 'diabetic foot ulcer', 'diabetic peripheral neuropathy' and 'atherosclerosis' were used to search for related gene sets in the GEO database. Differentially expressed genes (DEGs) were screened and analysed for GO, KEGG and enrichR functional enrichment. Potential three disease biomarkers were identified by SVM-SVM-RFE and LASSO regression analysis. The results were also validated using external datasets and discriminability was measured by area under the ROC curve (AUC). Finally, biomarkers and co-upregulated genes were analysed through the GSEA and Attie Laboratories diabetes databases. A total of 11 shared genes (KRT16, CD24, SAMD9L, SRGAP2, FGL2, GPR34, DDIT4, NFE2L3, FBLN5, ANXA3 and CPA3), two biomarkers (SAMD9L and FGL2) and one co-upregulated gene (CD24) were screened. GO and KEGG pathway analysis of DEGs, enrichr enrichment analysis of shared differential genes and GSEA analysis of biomarkers showed that these significant genes were mainly focused on vasoregulatory, inflammatory-oxidative stress and immunomodulatory pathways. In this study, we used bioinformatics to investigate the intrinsic relationship and potential mechanisms of three common lower extremity complications of diabetes and identified two pivotal genes using the LASSO model and the SVM-RFE algorithm, which will further help clinicians to understand the relationship between diabetic complications, improve the diagnosis and treatment of diabetic foot problems and help doctors to identify the potential risk factors of diabetic foot.

Development of novel self-assembled vaccines based on tumour-specific antigenic peptide and TLR2 agonist for effective breast cancer immunotherapy via activating CD8+ T cells and enhancing their function.

Vaccines based on tumour-specific antigens are a promising approach for immunotherapy. However, the clinical efficacy of tumour-specific antigens is still challenging. Twelve conjugates with self-assembly properties were designed and synthesized using MAGE-A1 peptide and TLR2 agonist, combined with different covalent bonds. All the developed conjugates formed spherical nanoparticles with a diameter of approximately 150 nm, and enhanced the efficacy of the peptide vaccines with the better targeting of lymph nodes. All the conjugates could well bind to serum albumin and improve the plasma stability of the individual antigenic peptides. In particular, conjugate 6 (N-Ac PamCS-M-6) had a more significant ability to promote dendritic cell maturation, CD8+ T cell activation, and subsequent killing of tumour cells, with an in vivo tumour inhibition rate of 70 ± 2.9%. The interaction between specific response and the different conjugation modes was further explored, thereby providing a fundamental basis for novel immune anti-tumour molecular platforms.

Oocytes and hypoxanthine orchestrate the G2-M switch mechanism in ovarian granulosa cells.

In mammalian growing follicles, oocytes are arrested at the diplotene stage (which resembles the G2/M boundary in mitosis), while the granulosa cells (GCs) continue to proliferate during follicular development, reflecting a cell cycle asynchrony between oocytes and GCs. Hypoxanthine (Hx), a purine present in the follicular fluid, has been shown to induce oocytes meiotic arrest, although its role in GC proliferation remains ill-defined. Here, we demonstrate that Hx indiscriminately prevents G2-to-M phase transition in porcine GCs. However, oocyte-derived paracrine factors (ODPFs), particularly GDF9 and BMP15, maintain the proliferation of GCs, partly by activating the ERK1/2 signaling and enabling the G2/M transition that is suppressed by Hx. Interestingly, GCs with lower expression of GDF9/BMP15 receptors appear to be more sensitive to Hx-induced G2/M arrest and become easily detached from the follicular wall. Importantly, Hx-mediated inhibition of G2/M progression instigates GC apoptosis, which is ameliorated in the presence of GDF9 and/or BMP15. Therefore, our data indicate that the counterbalance of intrafollicular factors, particularly Hx and oocyte-derived GDF9/BMP15, fine-tunes the development of porcine follicles by regulating the cell cycle progression of GCs.

EPSD: a well-annotated data resource of protein phosphorylation sites in eukaryotes.

As an important post-translational modification (PTM), protein phosphorylation is involved in the regulation of almost all of biological processes in eukaryotes. Due to the rapid progress in mass spectrometry-based phosphoproteomics, a large number of phosphorylation sites (p-sites) have been characterized but remain to be curated. Here, we briefly summarized the current progresses in the development of data resources for the collection, curation, integration and annotation of p-sites in eukaryotic proteins. Also, we designed the eukaryotic phosphorylation site database (EPSD), which contained 1 616 804 experimentally identified p-sites in 209 326 phosphoproteins from 68 eukaryotic species. In EPSD, we not only collected 1 451 629 newly identified p-sites from high-throughput (HTP) phosphoproteomic studies, but also integrated known p-sites from 13 additional databases. Moreover, we carefully annotated the phosphoproteins and p-sites of eight model organisms by integrating the knowledge from 100 additional resources that covered 15 aspects, including phosphorylation regulator, genetic variation and mutation, functional annotation, structural annotation, physicochemical property, functional domain, disease-associated information, protein-protein interaction, drug-target relation, orthologous information, biological pathway, transcriptional regulator, mRNA expression, protein expression/proteomics and subcellular localization. We anticipate that the EPSD can serve as a useful resource for further analysis of eukaryotic phosphorylation. With a data volume of 14.1 GB, EPSD is free for all users at http://epsd.biocuckoo.cn/.

Generation of 1.3†µm femtosecond pulses by cascaded nonlinear optical gain modulation in phosphosilicate fiber.

Nonlinear optical gain modulation (NOGM) is a simple and effective approach to generate highly coherent ultrafast pulses with a flexible wavelength. In this work, we demonstrate 34 nJ, 170 fs pulse generation at 1319 nm through a piece of phosphorus-doped fiber by two-stage cascaded NOGM with a 1064 nm pulsed pump. Beyond the experiment, numerical results show that 668 nJ, 391 fs pulses at 1.3 µm can be produced with up to 67% conversion efficiency by increasing the pump pulse energy and optimizing the pump pulse duration. This would offer an efficient method to obtain high-energy sub-picosecond laser sources for applications such as multiphoton microscopy.

Generation of 978†nm dispersion-managed solitons from a polarization-maintaining Yb-doped figure-of-9 fiber laser.

Restricted by the narrow gain bandwidth of Yb3+ near 980 nm, it is challenging to generate dispersion-managed (DM) solitons at this wavelength. In this work, we demonstrate the generation of DM solitons at 978 nm in a polarization-maintaining (PM) figure-of-9 fiber laser. Highly coherent pulses with 14.4 nm spectral bandwidth and 175 fs pulse duration are experimentally obtained. To the best of our knowledge, this is the shortest ∼980 nm pulse ever reported in an Yb-doped mode-locked fiber laser. Numerical simulations are performed to reveal the DM solitons' temporal and spectral evolution inside the figure-of-9 cavity under the condition of a narrow gain bandwidth. This robust and cost-effective 978 nm femtosecond laser is a promising light source for applications such as underwater communication and biophotonics.

Intermolecular Charge Transfer Induced Fragmentation of Formic Acid Dimers.

We investigate the intermolecular nonradiative charge transfer process in a double hydrogen-bonded formic acid (FA) dimer, initiated by electron-collision induced double ionization of one FA molecule. Through fragment ions and electron coincident momentum measurements and ab initio calculations, we obtain direct evidence that electron transfer from the neighboring FA molecule to fill one of the two vacancies occurs by a potential energy curve crossing of FA^{++}+FA with FA^{+}+FA^{+*} curves, forming an electronic excited state of dicationic dimers. This process causes the breaking of two hydrogen bonds and subsequently the cleavage of C─H and C─O covalent bonds in the dimers, which is expected to be a general phenomenon occurring in molecular complexes and can have important implications for radiation damage to biological matter.

Structural and dynamical studies of CH-πbonded CH4-C6H6dimer by ultrafast intermolecular Coulombic decay.

The inner-valence ionization and fragmentation dynamics of CH4-C6H6dimer induced by 200 eV electron impact is studied utilizing a multi-particle coincidence momentum spectroscopy. The three-dimensional momentum vectors and kinetic energy release (KER) of the CH4++C6H6+ion pairs are obtained by coincident momentum measurement. Our analysis on the absolute cross sections indicates that the intermediate dication CH4+-C6H6+is preferentially produced by the removal of an inner-valence electron from CH4or C6H6and subsequent relaxation of ultrafast intermolecular Coulombic decay followed by two-body Coulomb explosion. Combining withab initiomolecular dynamics (AIMD) simulations, the real-time fragmentation dynamics including translational, vibrational and rotational motions are presented as a function of propagation time. The revealed fragmentation dynamics are expected to have a potential implication for crystal structure imaging with various radiation sources.