Visualizing the Assembly Pathway of Nucleolar Pre-60S Ribosomes.

Eukaryotic 60S ribosomal subunits are comprised of three rRNAs and ∼50 ribosomal proteins. The initial steps of their formation take place in the nucleolus, but, owing to a lack of structural information, this process is poorly understood. Using cryo-EM, we solved structures of early 60S biogenesis intermediates at 3.3 Å to 4.5 Å resolution, thereby providing insights into their sequential folding and assembly pathway. Besides revealing distinct immature rRNA conformations, we map 25 assembly factors in six different assembly states. Notably, the Nsa1-Rrp1-Rpf1-Mak16 module stabilizes the solvent side of the 60S subunit, and the Erb1-Ytm1-Nop7 complex organizes and connects through Erb1's meandering N-terminal extension, eight assembly factors, three ribosomal proteins, and three 25S rRNA domains. Our structural snapshots reveal the order of integration and compaction of the six major 60S domains within early nucleolar 60S particles developing stepwise from the solvent side around the exit tunnel to the central protuberance.

An essential thioredoxin-type protein of Trypanosoma brucei acts as redox-regulated mitochondrial chaperone.

Most known thioredoxin-type proteins (Trx) participate in redox pathways, using two highly conserved cysteine residues to catalyze thiol-disulfide exchange reactions. Here we demonstrate that the so far unexplored Trx2 from African trypanosomes (Trypanosoma brucei) lacks protein disulfide reductase activity but functions as an effective temperature-activated and redox-regulated chaperone. Immunofluorescence microscopy and fractionated cell lysis revealed that Trx2 is located in the mitochondrion of the parasite. RNA-interference and gene knock-out approaches showed that depletion of Trx2 impairs growth of both mammalian bloodstream and insect stage procyclic parasites. Procyclic cells lacking Trx2 stop proliferation under standard culture conditions at 27°C and are unable to survive prolonged exposure to 37°C, indicating that Trx2 plays a vital role that becomes augmented under heat stress. Moreover, we found that Trx2 contributes to the in vivo infectivity of T. brucei. Remarkably, a Trx2 version, in which all five cysteines were replaced by serine residues, complements for the wildtype protein in conditional knock-out cells and confers parasite infectivity in the mouse model. Characterization of the recombinant protein revealed that Trx2 can coordinate an iron sulfur cluster and is highly sensitive towards spontaneous oxidation. Moreover, we discovered that both wildtype and mutant Trx2 protect other proteins against thermal aggregation and preserve their ability to refold upon return to non-stress conditions. Activation of the chaperone function of Trx2 appears to be triggered by temperature-mediated structural changes and inhibited by oxidative disulfide bond formation. Our studies indicate that Trx2 acts as a novel chaperone in the unique single mitochondrion of T. brucei and reveal a new perspective regarding the physiological function of thioredoxin-type proteins in trypanosomes.

The Rise of the TROP2-Targeting Agents in NSCLC: New Options on the Horizon.

BACKGROUND: Lung cancer is the most common thoracic malignancy, representing the leading cause of cancer-related deaths worldwide with a 5-year survival rate of <10%. SUMMARY: The emergence of targeted therapy and immunotherapy has changed the treatment paradigm of advanced non-small cell lung cancer (NSCLC). However, for those who are not eligible for such therapy or currently have no available standard treatment options, new precision treatment approaches are needed. Human trophoblast cell-surface antigen 2 (TROP2) is a transmembrane glycoprotein that is highly expressed on several epithelial tumours including NSCLC. TROP2 is recognized as a promising molecular target for therapeutic development in various types of TROP2-expressing malignancies. As a result, several TROP2-targeted therapeutics have recently been developed for clinical use, such as anti-TROP2 antibodies and TROP2-targeted antibody-drug conjugates. Key Message: This review explores the literature data on the role of TROP2 in cancer development and the potential use of emerging TROP2 antibody-drug conjugates in NSCLC treatment.

DENR-MCTS1 heterodimerization and tRNA recruitment are required for translation reinitiation.

The succession of molecular events leading to eukaryotic translation reinitiation-whereby ribosomes terminate translation of a short open reading frame (ORF), resume scanning, and then translate a second ORF on the same mRNA-is not well understood. Density-regulated reinitiation and release factor (DENR) and multiple copies in T-cell lymphoma-1 (MCTS1) are implicated in promoting translation reinitiation both in vitro in translation extracts and in vivo. We present here the crystal structure of MCTS1 bound to a fragment of DENR. Based on this structure, we identify and experimentally validate that DENR residues Glu42, Tyr43, and Tyr46 are important for MCTS1 binding and that MCTS1 residue Phe104 is important for tRNA binding. Mutation of these residues reveals that DENR-MCTS1 dimerization and tRNA binding are both necessary for DENR and MCTS1 to promote translation reinitiation in human cells. These findings thereby link individual residues of DENR and MCTS1 to specific molecular functions of the complex. Since DENR-MCTS1 can bind tRNA in the absence of the ribosome, this suggests the DENR-MCTS1 complex could recruit tRNA to the ribosome during reinitiation analogously to the eukaryotic initiation factor 2 (eIF2) complex in cap-dependent translation.

YvcI from Bacillus subtilis has in vitro RNA pyrophosphohydrolase activity.

RNA degradation is one of several ways for organisms to regulate gene expression. In bacteria, the removal of two terminal phosphate moieties as orthophosphate (Bacillus subtilis) or pyrophosphate (Escherichia coli) triggers ribonucleolytic decay of primary transcripts by 5'-monophosphate-dependent ribonucleases. In the soil-dwelling firmicute species B. subtilis, the RNA pyrophosphohydrolase BsRppH, a member of the Nudix family, triggers RNA turnover by converting primary transcripts to 5'-monophospate RNA. In addition to BsRppH, a source of redundant activity in B. subtilis has been proposed. Here, using recombinant protein expression and in vitro enzyme assays, we provide evidence for several additional RNA pyrophosphohydrolases, among them MutT, NudF, YmaB, and YvcI in B. subtilis We found that in vitro, YvcI converts RNA 5'-di- and triphosphates into monophosphates in the presence of manganese at neutral to slightly acidic pH. It preferred G-initiating RNAs and required at least one unpaired nucleotide at the 5'-end of its substrates, with the 5'-terminal nucleotide determining whether primarily ortho- or pyrophosphate is released. Exchanges of catalytically important glutamate residues in the Nudix motif impaired or abolished the enzymatic activity of YvcI. In summary, the results of our extensive in vitro biochemical characterization raise the possibility that YvcI is an additional RNA pyrophosphohydrolase in B. subtilis.

Concerted removal of the Erb1-Ytm1 complex in ribosome biogenesis relies on an elaborate interface.

The complicated process of eukaryotic ribosome biogenesis involves about 200 assembly factors that transiently associate with the nascent pre-ribosome in a spatiotemporally ordered way. During the early steps of 60S subunit formation, several proteins, collectively called A3 cluster factors, participate in the removal of the internal transcribed spacer 1 (ITS1) from 27SA3 pre-rRNA. Among these factors is the conserved hetero-trimeric Nop7-Erb1-Ytm1 complex (or human Pes1-Bop1-Wdr12), which is removed from the evolving pre-60S particle by the AAA ATPase Rea1 to allow progression in the pathway. Here, we clarify how Ytm1 and Erb1 interact, which has implications for the release mechanism of both factors from the pre-ribosome. Biochemical studies show that Ytm1 and Erb1 bind each other via their ß-propeller domains. The crystal structure of the Erb1-Ytm1 heterodimer determined at 2.67Å resolution reveals an extended interaction surface between the propellers in a rarely observed binding mode. Structure-based mutations in the interface that impair the Erb1-Ytm1 interaction do not support growth, with specific defects in 60S subunit synthesis. Under these mutant conditions, it becomes clear that an intact Erb1-Ytm1 complex is required for 60S maturation and that loss of this stable interaction prevents ribosome production.

The velvet family of fungal regulators contains a DNA-binding domain structurally similar to NF-κB.

Morphological development of fungi and their combined production of secondary metabolites are both acting in defence and protection. These processes are mainly coordinated by velvet regulators, which contain a yet functionally and structurally uncharacterized velvet domain. Here we demonstrate that the velvet domain of VosA is a novel DNA-binding motif that specifically recognizes an 11-nucleotide consensus sequence consisting of two motifs in the promoters of key developmental regulatory genes. The crystal structure analysis of the VosA velvet domain revealed an unforeseen structural similarity with the Rel homology domain (RHD) of the mammalian transcription factor NF-κB. Based on this structural similarity several conserved amino acid residues present in all velvet domains have been identified and shown to be essential for the DNA binding ability of VosA. The velvet domain is also involved in dimer formation as seen in the solved crystal structures of the VosA homodimer and the VosA-VelB heterodimer. These findings suggest that defence mechanisms of both fungi and animals might be governed by structurally related DNA-binding transcription factors.

The Aspergillus nidulans MAPK module AnSte11-Ste50-Ste7-Fus3 controls development and secondary metabolism.

The sexual Fus3 MAP kinase module of yeast is highly conserved in eukaryotes and transmits external signals from the plasma membrane to the nucleus. We show here that the module of the filamentous fungus Aspergillus nidulans (An) consists of the AnFus3 MAP kinase, the upstream kinases AnSte7 and AnSte11, and the AnSte50 adaptor. The fungal MAPK module controls the coordination of fungal development and secondary metabolite production. It lacks the membrane docking yeast Ste5 scaffold homolog; but, similar to yeast, the entire MAPK module's proteins interact with each other at the plasma membrane. AnFus3 is the only subunit with the potential to enter the nucleus from the nuclear envelope. AnFus3 interacts with the conserved nuclear transcription factor AnSte12 to initiate sexual development and phosphorylates VeA, which is a major regulatory protein required for sexual development and coordinated secondary metabolite production. Our data suggest that not only Fus3, but even the entire MAPK module complex of four physically interacting proteins, can migrate from plasma membrane to nuclear envelope.

RNA synthesis and purification for structural studies.

RNAs play pivotal roles in the cell, ranging from catalysis (e.g., RNase P), acting as adaptor molecule (tRNA) to regulation (e.g., riboswitches). Precise understanding of its three-dimensional structures has given unprecedented insight into the molecular basis for all of these processes. Nevertheless, structural studies on RNA are still limited by the very special nature of this polymer. The most common methods for the determination of 3D RNA structures are NMR and X-ray crystallography. Both methods have their own set of requirements and give different amounts of information about the target RNA. For structural studies, the major bottleneck is usually obtaining large amounts of highly pure and homogeneously folded RNA. Especially for X-ray crystallography it can be necessary to screen a large number of variants to obtain well-ordered single crystals. In this mini-review we give an overview about strategies for the design, in vitro production, and purification of RNA for structural studies.

Advancing Research Training in Medical Education: Global Perspectives and Paradigms for Future Development.

BACKGROUND: This study delves into the dynamic field of medical education research, emphasizing the integration of research training within medical curricula. It seeks to understand the impact of such integration on the competencies of future medical professionals. OBJECTIVE: The primary aim is to systematically categorize and analyze the current trends and future directions in research training in medical education. This involves assessing the influence of research training on medical students' skills and the methodologies used in such research. METHODS: The research employs an extensive bibliographic literature review across multiple databases. It classifies studies like experiential or case studies, editorials, and original research articles. This classification is based on criteria such as geographical location, research objectives, theoretical frameworks, and methodologies. RESULTS: Findings reveal a diverse landscape in medical education research, with a significant emphasis on research training. The research showcases varying methodologies and approaches used globally, highlighting the thematic focus and geographical distribution of these studies. CONCLUSION: Research training in medical education is a globally expansive and evolving field. It underscores the importance of continuous investigation, particularly focusing on integrating research elements at curricular levels and exploring innovative educational strategies. The study also points out potential research gaps, especially in underrepresented regions, indicating directions for future research efforts.

Integrating Research and Teaching in Medical Education: Challenges, Strategies, and Implications for Healthcare.

Introduction: The integration of research and teaching in medical education offers numerous benefits, fostering critical thinking and analytical skills in students. Institutions worldwide have recognized the significance of this nexus and have implemented initiatives to link teaching with discipline-based research, promoting interdisciplinary collaboration. This article aims to explore the challenges and recommendations for integrating research and teaching in medical schools and provide recommendations to overcome these challenges. Methods: We conducted a comprehensive review of the literature to identify the common challenges faced by medical institutions in integrating research and teaching. PubMed, Scopus, Web of Science, ERIC, and Google Scholar databases were searched to assess the literature that met the study objectives with explicit inclusion and exclusion criteria. We also examined successful strategies employed by some institutions to promote research-teaching integration. Results: The challenges identified include limited resources, the need to balance research and curriculum requirements, and the importance of cultivating a research-oriented institutional culture. Successful strategies involve curriculum updates, faculty motivation, and cross-disciplinary collaboration. Implementing strategies involve vertically and horizontally integrating research methodology throughout the undergraduate curriculum and cross-integrating traditional medical courses with other disciplines. Collaboration between universities, enterprises, and schools can enhance comprehensive cooperation. Conclusion: To create a research-oriented learning environment, medical institutions should address these challenges and implement effective strategies. This approach will not only nurture research-oriented healthcare professionals but also advance medical knowledge for the benefit of patient care. By addressing these challenges and implementing appropriate strategies, medical institutions can create a research-oriented learning environment, nurturing research-oriented healthcare professionals and advancing medical knowledge to improve patient care.

Enhancing Cancer Care Amid Conflict: A Proposal for Optimizing Oncology Services During Wartime.

The landscape of cancer care within armed conflict zones is characterized by intricate challenges arising from disrupted health care systems, scarcity of resources, and population displacement. During times of war, the provision of cancer services is often disrupted, leading to significant challenges for oncologists and other health care providers. To optimize cancer services during wartime, several key priorities must be addressed. Focusing on needs assessment, treatment prioritization, drug supply chain, telemedicine, mobile clinics, cross-border collaborations, health care staff support, and continuity of care will enable health care systems to provide essential cancer services and mitigate the adverse impact of conflict on patients with cancer. This article delineates the pivotal key priorities for optimizing cancer services during wartime. It calls for collaborative action, the integration of technology, and holistic care approaches to safeguard the rights, well-being, and dignity of individuals confronting the dual challenges of cancer and conflict. By addressing these priorities, health care providers, policymakers, and stakeholders can collectively ensure that cancer services remain steadfast and compassionate even amid the turmoil of war. Thus, it may be possible to optimize cancer services during wartime, ensuring that patients with cancer continue to receive the care they need.

Utilization of ChatGPT in Medical Education: Applications and Implications for Curriculum Enhancement.

Background: The integration of artificial intelligence (AI) into medical education has sparked a paradigm shift in pedagogical approaches, reshaping the way medical knowledge is accessed, processed, and applied. Medical education is a dynamic field that demands continuous adaptation to the evolving healthcare landscape. ChatGPT, an advanced AI language model, with its natural language understanding and generation capabilities, offers a multifaceted toolset that enhances various aspects of medical education. Objective: The objective of this paper is to explore how ChatGPT, an advanced AI language model, is transforming medical education by serving as a dynamic information resource and driving curriculum reform. It aims to highlight the multifaceted uses of ChatGPT and its potential to reshape the pedagogical landscape in medical education. Methods: PubMed, Scopus, Web of Science, ERIC, and Google Scholar databases were searched to assess the literature that met the study objectives from 2019 to August 2023 with explicit inclusion and exclusion criteria. Results: The results demonstrate that ChatGPT's applications in medical education are diverse and encompass real-time curriculum adaptation, personalized learning, and collaborative learning. Its capacity to provide immediate and contextually relevant information has the potential to enhance the quality of medical education significantly. Conclusion: ChatGPT's integration into medical education represents a transformative shift in educational approaches. It offers a wide range of capabilities, from serving as a repository of medical knowledge to facilitating collaborative learning. As medical education continues to evolve, ChatGPT emerges as a powerful tool that can reshape pedagogy and drive meaningful curriculum reform to meet the needs of modern healthcare practice.ChatGPT emerges as a transformative tool that holds the potential to reshape the landscape of medical pedagogy and drive meaningful curriculum reform.

The zinc-finger protein Red1 orchestrates MTREC submodules and binds the Mtl1 helicase arch domain.

Cryptic unstable transcripts (CUTs) are rapidly degraded by the nuclear exosome in a process requiring the RNA helicase Mtr4 and specific adaptor complexes for RNA substrate recognition. The PAXT and MTREC complexes have recently been identified as homologous exosome adaptors in human and fission yeast, respectively. The eleven-subunit MTREC comprises the zinc-finger protein Red1 and the Mtr4 homologue Mtl1. Here, we use yeast two-hybrid and pull-down assays to derive a detailed interaction map. We show that Red1 bridges MTREC submodules and serves as the central scaffold. In the crystal structure of a minimal Mtl1/Red1 complex an unstructured region adjacent to the Red1 zinc-finger domain binds to both the Mtl1 KOW domain and stalk helices. This interaction extends the canonical interface seen in Mtr4-adaptor complexes. In vivo mutational analysis shows that this interface is essential for cell survival. Our results add to Mtr4 versatility and provide mechanistic insights into the MTREC complex.

Colonic Metastasis of Primary Lung Cancer.

The colon is an uncommon secondary site for metastasis of lung adenocarcinoma. Distinguishing primary colonic carcinoma from metastatic spread of lung carcinoma can be difficult. We present a case of a patient with lung adenocarcinoma who, on abdominal computed tomography scan examination, was found to have a sigmoid tumor that was thought to represent a synchronous primary colorectal adenocarcinoma. Histological examination of endoscopic sigmoid tumor biopsies confirmed this to be metastasis from the lung adenocarcinoma. The patient subsequently developed major rectal bleeding and deteriorated significantly. This case also illustrates the poor prognosis association with colorectal metastasis of lung cancer.

Correction: Mpp10 represents a platform for the interaction of multiple factors within the 90S pre-ribosome.

[This corrects the article DOI: 10.1371/journal.pone.0183272.].

Does Curriculum Analysis in Clinical Residency Training Need to be Different?

OBJECTIVES: Curriculum analysis is an essential process in exploring the validity of assumptions behind the curriculum and judging its perspectives, goals, and objectives. This study analysed the curriculum of the Internal Medicine Residency Programme at the Sudan Medical Specialisation Board, exploring the programme's strengths, weaknesses, and areas for improvement. MATERIALS AND METHODS: A qualitative descriptive analysis method using Harden Ten Questions framework was used, implemented through document analysis. Thematic analysis was used to categorise the findings following the Harden Ten Questions framework. RESULTS: The analysis of this curriculum using Harden Ten Questions framework revealed that it is built on needs assessment, with clear objectives and contents, good organisation of the contents and a precise management process. However, there is a need to improve the areas related to training strategies, training methods, assessment methods and the learning environment. CONCLUSIONS: This study showed that the curriculum is built to meet the needs of the Sudanese community. The contents fit for internal medicine clinical residency and the methods of training are likely to foster long-life learning. The curriculum needs some improvement particularly in the following areas such as clinical teaching, trainee assessments, and learning environments. Harden Ten Questions framework for curricula was practical to some degree in analysing the postgraduate curriculum. However, some suggestions have been made regarding the original framework to make it more adaptable to curriculum analysis at postgraduate level. The systematic analysis is transferable to the analysis of other undergraduate or postgraduate medical curricula in Sudan.

Crystal structures of Rea1-MIDAS bound to its ribosome assembly factor ligands resembling integrin-ligand-type complexes.

The Rea1 AAA+-ATPase dislodges assembly factors from pre-60S ribosomes upon ATP hydrolysis, thereby driving ribosome biogenesis. Here, we present crystal structures of Rea1-MIDAS, the conserved domain at the tip of the flexible Rea1 tail, alone and in complex with its substrate ligands, the UBL domains of Rsa4 or Ytm1. These complexes have structural similarity to integrin α-subunit domains when bound to extracellular matrix ligands, which for integrin biology is a key determinant for force-bearing cell-cell adhesion. However, the presence of additional motifs equips Rea1-MIDAS for its tasks in ribosome maturation. One loop insert cofunctions as an NLS and to activate the mechanochemical Rea1 cycle, whereas an additional ß-hairpin provides an anchor to hold the ligand UBL domains in place. Our data show the versatility of the MIDAS fold for mechanical force transmission in processes as varied as integrin-mediated cell adhesion and mechanochemical removal of assembly factors from pre-ribosomes.

Students' perception of the learning environment and its relation to their study year and performance in Sudan.

OBJECTIVES: To evaluate students' perceptions of the learning environment and to assess any differences in perception related to students' performance and their year of study. METHODS: A descriptive cross-sectional study was performed of 638 students from the second, sixth and tenth semesters at the Faculty of Medicine at Gezira University, Sudan. This study employed the Arabic-translated Dundee Ready Education Environment Measure. The main predictor variables were the study year and academic performance. Descriptive statistics and one-way analysis of variance with a post hoc Tukey-Kramer multiple comparisons test were used for data analysis. RESULTS: The overall score for this study was 122/200 (SD=16.6), indicating a positive perception of the learning environment. The overall mean score was 109.94/200 (SD=21.2) for Semester 2 students, 122.9/200 (SD=20.29) for Semester 6 students, and 116.53 (SD=20.12) for Semester 10 students, reflecting a significant difference in students' perceptions in different years of study (F (2,2422) = 3.21, p=0.04). There was also a significant difference between the mean overall scores with respect to academic performance. High-achieving students' mean DREEM score was 126 (SD=24.4); while low-achieving students' mean DREEM score was 102 (SD=26.25) (F(2,2453) = 3.53, p=0.029). CONCLUSIONS: High achievers' perceptions of the learning environment are significantly better than those of low achievers. A significant difference was observed between students in different years of study. The differences in students' academic performance should be further investigated, targeting specific domains. A large-scale study is required to differentiate between the weakness and the strength of each academic level.

Unusual Sites of High-Grade Neuroendocrine Carcinomas: A Case Series and Review of the Literature.

BACKGROUND Neuroendocrine tumors (NETs) encompass a diverse group of varying clinicopathological entities arising from cells of the endocrine and nervous systems. The presentation of these unique tumors can range from occult disease discovered incidentally to hyperactive, metastatic secretory tumors. NETs most commonly originate in the gastrointestinal and respiratory tract, although they may occur at any site in the body due to the wide distribution of neuroendocrine cells. Their classification system is complex and continues to evolve, and the current system uses histological grade in defining these subtypes. Neuroendocrine carcinomas (NECs), or high-grade, poorly-differentiated NETs, are the most aggressive subtype. Surgical resection remains the primary treatment modality and may be curative, thus early diagnosis is paramount. Management of advanced NETs remains both a diagnostic and therapeutic challenge; however, advances in our understanding of these unique neoplasms as well as an evolving classification system has led to the development of adjunctive therapeutic approaches aimed to minimize morbidity and improve patient outcomes. CASE REPORT We present 6 cases of unusual sites of high-grade neuroendocrine carcinomas involving the cervix, gallbladder, oesophagus, ovary, prostate, and urinary bladder. CONCLUSIONS Our case series highlights the heterogenous and aggressive nature of this subtype of NETs as well as their diagnostic and therapeutic difficulties. We also review the evolution of the NET classification system and its impact on the management of these malignancies.