Negative Regulation of the Mis17-Mis6 Centromere Complex by mRNA Decay Pathway and EKC/KEOPS Complex in Schizosaccharomyces pombe

The mitotic kinetochore forms at the centromere for proper chromosome segregation. Deposition of the centromere-specific histone H3 variant, spCENP-A/Cnp1, is vital for the formation of centromere-specific chromatin and the Mis17-Mis6 complex of the fission yeast Schizosaccharomyces pombe is required for this deposition. Here we identified extragenic suppressors for a Mis17-Mis6 complex temperature-sensitive (ts) mutant, mis17-S353P, using whole-genome sequencing. The large and small daughter nuclei phenotype observed in mis17-S353P was greatly rescued by these suppressors. Suppressor mutations in two ribonuclease genes involved in the mRNA decay pathway, exo2 and pan2, may affect Mis17 protein level, as mis17 mutant protein level was recovered in mis17-S353P exo2 double mutant cells. Suppressor mutations in EKC/KEOPS complex genes may not regulate Mis17 protein level, but restored centromeric localization of spCENP-A/Cnp1, Mis6 and Mis15 in mis17-S353P. Therefore, the EKC/KEOPS complex may inhibit Mis17-Mis6 complex formation or centromeric localization. Mutational analysis in protein structure indicated that suppressor mutations in the EKC/KEOPS complex may interfere with its kinase activity or complex formation. Our results suggest that the mRNA decay pathway and the EKC/KEOPS complex negatively regulate Mis17-Mis6 complex-mediated centromere formation by distinct and unexpected mechanisms

Whole-Genome Sequencing of Suppressor DNA Mixtures Identifies Pathways That Compensate for Chromosome Segregation Defects in Schizosaccharomyces pombe

Suppressor screening is a powerful method to identify genes that, when mutated, rescue the temperature sensitivity of the original mutation. Previously, however, identification of suppressor mutations has been technically difficult. Due to the small genome size of Schizosaccharomyces pombe, we developed a spontaneous suppressor screening technique, followed by a cost-effective sequencing method. Genomic DNAs of 10 revertants that survived at the restrictive temperature of the original temperature sensitive (ts) mutant were mixed together as one sample before constructing a library for sequencing. Responsible suppressor mutations were identified bioinformatically based on allele frequency. Then, we isolated a large number of spontaneous extragenic suppressors for three ts mutants that exhibited defects in chromosome segregation at their restrictive temperature. Screening provided new insight into mechanisms of chromosome segregation: loss of Ufd2 E4 multi-ubiquitination activity suppresses defects of an AAA ATPase, Cdc48. Loss of Wpl1, a releaser of cohesin, compensates for the Eso1 mutation, which may destabilize sister chromatid cohesion. The segregation defect of a ts histone H2B mutant is rescued if it fails to be deubiquitinated by the SAGA complex, because H2B is stabilized by monoubiquitination

Screening of potential biomarkers in the occurrence and development of type 1 diabetes mellitus based on transcriptome analysis

Introduction: The aim of the study was to reveal the mechanisms for the pathogenesis and progression of type 1 diabetes mellitus (T1DM). Material and methods: Two mRNA expression profiles and two miRNA expression profiles were downloaded from the Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs), differentially expressed miRNAs (DEMs), functional enrichment analyses, pathways, putative targets for DEMs and the miRNA-gene pairs, protein-protein pairs of DEGs, and PPI network were constructed. Results: Based on mRNA expression profiles, 37 and 110 DEGs were identified, and named as DEGs-short and DEGs-long, respectively. Based on miRNA expression profiles, 15 and six DEMs were identified, and named as DEMs-short and DEMs-long, respectively. DEGs-short were enriched in six GO terms and four pathways, and DEGs-long enriched in 40 GO terms and 10 pathways. Seventeen miRNA-gene pairs for DEMs-short were screened out; hisa-miR-181a and hisa-miR-181c were involved in the most pairs. Twenty pairs for DEMs-long were obtained; hsa-miR-338-3p was involved in all the pairs. KLRD1 was involved in more pairs in the network of DEGs-short. ACTA2 and USP9Y were involved in more pairs in the network of DEGs-long. Conclusions: KLRD1, hisa-miR-181a, and hisa-miR-181c might be pathogenic biomarkers for T1DM, ACTA2, USP9Y, and hsa-miR-338-3p progressive biomarkers of T1DM.

Cohesin ATPase activities regulate DNA binding and coiled-coil configuration

The cohesin complex is required for sister chromatid cohesion and genome compaction. Cohesin coiled coils (CCs) can fold at break sites near midpoints to bring head and hinge domains, located at opposite ends of coiled coils, into proximity. Whether ATPase activities in the head play a role in this conformational change is yet to be known. Here, we dissected functions of cohesin ATPase activities in cohesin dynamics in Schizosaccharomyces pombe. Isolation and characterization of cohesin ATPase temperature-sensitive (ts) mutants indicate that both ATPase domains are required for proper chromosome segregation. Unbiased screening of spontaneous suppressor mutations rescuing the temperature lethality of cohesin ATPase mutants identified several suppressor hotspots in cohesin that located outside of ATPase domains. Then, we performed comprehensive saturation mutagenesis targeted to these suppressor hotspots. Large numbers of the identified suppressor mutations indicated several different ways to compensate for the ATPase mutants: 1) Substitutions to amino acids with smaller side chains in coiled coils at break sites around midpoints may enable folding and extension of coiled coils more easily; 2) substitutions to arginine in the DNA binding region of the head may enhance DNA binding; or 3) substitutions to hydrophobic amino acids in coiled coils, connecting the head and interacting with other subunits, may alter conformation of coiled coils close to the head. These results reflect serial structural changes in cohesin driven by its ATPase activities potentially for packaging DNAs

Effect of Heat Shock Protein A6 on Changes in Meat Quality of Qinchuan Cattle during Postmortem Maturation

In order to investigate the effect of differential expression of heat shock protein family A (Hsp70) member 6 (HSPA6) on the meat quality of Qinchuan cattle during postmortem maturation, the Longissimus dorsi muscle of Qinchuan cattle was stored at 4 ℃ and evaluated for meat quality at 0, 2, 4, 6 and 8 days postmortem. 4D-Label free quantification (4D-LFQ) was used to analyze the expression of HSPA6 and related differential proteins. The results showed that the expression of HSPA6 decreased with increasing storage time, the contents of adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP) and reduced form of nicotinamide-adenine dinucleotide (NADH) decreased, the myofibril fragmentation index (MFI) increased, and the shear force and centrifugal loss increased and then decreased. Correlation analysis showed that HSPA6 expression was positively correlated with the contents of ATP and NADH and shear force (P < 0.05) and negatively correlated with MFI and centrifugal loss (P < 0.01), indicating that HSPA6 degradation was closely related to energy metabolism and water retention in beef during storage. Proteomics identified 24 significantly differentially expressed proteins (DEPs) associated with HSPA6 expression, which could cause proteasome-mediated ubiquitin-dependent proteolytic metabolism and cellular proteolysis through carbohydrate derivative binding and purine nucleoside triphosphate binding, in turn causing metabolic disorders or imbalances. These DEPs could also control protein degradation and cell apoptosis, in turn affecting structural changes in muscle and reducing tissue energy levels, and ultimately affecting the tenderness, MFI and water retention of Qinchuan beef through resistance to cellular structural protein degradation

Advanced HCC with amplified mesenchymal epithelial transition factor receptor responds well to savolitinib: a case report

ObjectiveCurrent treatment agents for HCC are mostly immune checkpoint inhibitors (ICIs) plus bevacizumab and multitarget tyrosine kinase inhibitors (TKIs); however, their limited overall response rate and shorter median progression-free survival (PFS) discourage their frequent use. The development of Mesenchymal Epithelial Transition Factor receptor (MET) Tyrosine Kinase Inhibitors (MET-TKI) has transformed the treatment pattern in MET-altered solid tumors and improved their prognosis. However, the benefits of MET-TKIs in MET-amplified hepatocellular carcinoma (HCC) remain unclear.MethodsHere, we present a case of advanced HCC amplified with MET treated with savolitinib, a MET-TKI, after progression from first-line treatment with bevacizumab plus sintilimab.ResultsThe patient achieved a partial response (PR) to savolitinib in the second line setting. The progression-free survival (PFS) of first-line of bevacizumab plus sintilimab and sequential second-line treatment with MET-TKI, savolitinib, are 3 and over 8 months, respectively. furthermore, the patient still had continuous PR status with manageable toxicities.ConclusionsThe present case report provides first-hand evidence that savolitinib may be beneficial for patients with advanced MET-amplified HCC and offers a promising treatment option

Seabed Politics and International Law: A Review of Seabed Politics by Barry Buzan -And New Developments in International Seabed Politics

刘中民，教育部人文社会科学重点研宄基地中国海洋大学海洋发展研宄院研究员，中国海洋大学法学院教授。本文是2004年教育部人文社会科学重点研究基地重大项目 “世界海洋政治的发展与中国海洋战略选择”（04JJDZH017)、2003年教育部重大攻 关项目子课题“中国海洋政治战略研究”（03JZD0024)、2004年山东省社科规划项目 “21世纪中国海洋政治战略研究”（04BMZ01)的部分研究成果。 黎兴亚，中国海洋大学法学院2004级研究生。【中文摘要】本文介绍了加拿大学者巴里•布赞的《海底政治》的主要内容及研究方法。该书以翔实的史料、历史的方法分析了国际海底制度发生、发展的政治进程，通过海底政治考察经济价值与政治进程的互动关系，以及国家、国际组织的政治行动与国际法律制度的相互作用、相互影响。巴里•布赞的思想对于研究现 今的国际海底制度以及海洋法甚至整个国际法制度，都有着重要意义。在评介巴 里•布赞思想的基础上，本文对该书出版后30年来国际海底制度的新发展进行了论述。 【Abstract】This article introduces the main contents and research methods of the book Seabed Politics by the Canadian scholar Barry Buzan. The book analyzes the occurrence and the development of the political process of the international seabed regime by reference to full and accurate historical data, adopting a historical approach. The book, through seabed politics, reviews the interactive relationship between economic value and political processes, as well as the interaction and interplay between political actions taken by countries and international organizations and the international legal regime. Barry Buzan’s thinking is significant for both research into the present international seabed regime and the law of the sea, and even the whole international legal regime. Based on the review of Barry Buzan’s thinking, this article discusses the new development of the international seabed regime over the 30 years since that book was published

The efficacy of furmonertinib in untreated advanced NSCLC patients with sensitive EGFR mutations in a real-world setting: a single institutional experience

BackgroundFurmonertinib is the standard treatment option in the first-line setting for advanced non-small cell lung cancer (NSCLC) with sensitive epidermal growth factor receptor (EGFR) mutations in China. However, there are limited real-world data available.MethodsWe conducted a retrospective study at a single center, analyzing a cohort of 73 NSCLC patients who tested positive for EGFR mutations and were treated with furmonertinib as their initial therapy between August 2022 and December 2023. The primary endpoint was progression-free survival (PFS), with secondary endpoints including objective response rate (ORR), overall survival (OS), and safety profile.ResultsThe median observation period was 9 months (95% confidence interval [CI], 8.0–20.0). The median PFS was 19.5 months (95% CI, 14.6–24.4). OS data were not yet mature. Univariate analysis showed no significant correlation between PFS and factors such as Eastern Cooperative Oncology Group performance status (ECOG PS) score, presence of brain or liver metastases, sex, age, EGFR mutation status, or number of metastatic sites. However, multivariate analysis indicated a potential trend toward extended PFS in patients younger than 65 years (p = 0.053, 95% CI, 0.10–1.02), although the p-value was only marginally significant. The most common adverse events were diarrhea (24%), anemia (36%), and liver injury (32%); however, only four cases experienced severe adverse events.ConclusionIn a real-world setting, furmonertinib appears to be a favorable treatment option for EGFR-mutated patients. The manageable nature of adverse events further supports its use in clinical practice

Suppressor mutation analysis combined with 3D modeling explains cohesin’s capacity to hold and release DNA

Cohesin is a fundamental protein complex that holds sister chromatids together. Separase protease cleaves a cohesin subunit Rad21/SCC1, causing the release of cohesin from DNA to allow chromosome segregation. To understand the functional organization of cohesin, we employed next-generation whole-genome sequencing and identified numerous extragenic suppressors that overcome either inactive separase/Cut1 or defective cohesin in the fission yeast Schizosaccharomyces pombe. Unexpectedly, Cut1 is dispensable if suppressor mutations cause disorders of interfaces among essential cohesin subunits Psm1/SMC1, Psm3/SMC3, Rad21/SCC1, and Mis4/SCC2, the crystal structures of which suggest physical and functional impairment at the interfaces of Psm1/3 hinge, Psm1 head–Rad21, or Psm3 coiled coil–Rad21. Molecular-dynamics analysis indicates that the intermolecular β-sheets in the cohesin hinge of cut1 suppressor mutants remain intact, but a large mobility change occurs at the coiled coil bound to the hinge. In contrast, suppressors of rad21-K1 occur in either the head ATPase domains or the Psm3 coiled coil that interacts with Rad21. Suppressors of mis4-G1326E reside in the head of Psm3/1 or the intragenic domain of Mis4. These may restore the binding of cohesin to DNA. Evidence is provided that the head and hinge of SMC subunits are proximal, and that they coordinate to form arched coils that can hold or release DNA by altering the angles made by the arched coiled coils. By combining molecular modeling with suppressor sequence analysis, we propose a cohesin structure designated the “hold-and-release” model, which may be considered as an alternative to the prevailing “ring” model