Recruitment of a SUMO isopeptidase to rDNA stabilizes silencing complexes by opposing SUMO targeted ubiquitin ligase activity.

Post-translational modification by SUMO (small ubiquitin-like modifier) plays important but still poorly understood regulatory roles in eukaryotic cells, including as a signal for ubiquitination by SUMO targeted ubiquitin ligases (STUbLs). Here, we delineate the molecular mechanisms for SUMO-dependent control of ribosomal DNA (rDNA) silencing through the opposing actions of a STUbL (Slx5:Slx8) and a SUMO isopeptidase (Ulp2). We identify a conserved region in the Ulp2 C terminus that mediates its specificity for rDNA-associated proteins and show that this region binds directly to the rDNA-associated protein Csm1. Two crystal structures show that Csm1 interacts with Ulp2 and one of its substrates, the rDNA silencing protein Tof2, through adjacent conserved interfaces in its C-terminal domain. Disrupting Csm1's interaction with either Ulp2 or Tof2 dramatically reduces rDNA silencing and causes a marked drop in Tof2 abundance, suggesting that Ulp2 promotes rDNA silencing by opposing STUbL-mediated degradation of silencing proteins. Tof2 abundance is rescued by deletion of the STUbL SLX5 or disruption of its SUMO-interacting motifs, confirming that Tof2 is targeted for degradation in a SUMO- and STUbL-dependent manner. Overall, our results demonstrate how the opposing actions of a localized SUMO isopeptidase and a STUbL regulate rDNA silencing by controlling the abundance of a key rDNA silencing protein, Tof2.

Assessment of the Clinical Relevance of BRCA2 Missense Variants by Functional and Computational Approaches.

Many variants of uncertain significance (VUS) have been identified in BRCA2 through clinical genetic testing. VUS pose a significant clinical challenge because the contribution of these variants to cancer risk has not been determined. We conducted a comprehensive assessment of VUS in the BRCA2 C-terminal DNA binding domain (DBD) by using a validated functional assay of BRCA2 homologous recombination (HR) DNA-repair activity and defined a classifier of variant pathogenicity. Among 139 variants evaluated, 54 had ?99% probability of pathogenicity, and 73 had ?95% probability of neutrality. Functional assay results were compared with predictions of variant pathogenicity from the Align-GVGD protein-sequence-based prediction algorithm, which has been used for variant classification. Relative to the HR assay, Align-GVGD significantly (p < 0.05) over-predicted pathogenic variants. We subsequently combined functional and Align-GVGD prediction results in a Bayesian hierarchical model (VarCall) to estimate the overall probability of pathogenicity for each VUS. In addition, to predict the effects of all other BRCA2 DBD variants and to prioritize variants for functional studies, we used the endoPhenotype-Optimized Sequence Ensemble (ePOSE) algorithm to train classifiers for BRCA2 variants by using data from the HR functional assay. Together, the results show that systematic functional assays in combination with in silico predictors of pathogenicity provide robust tools for clinical annotation of BRCA2 VUS.

Spreader Graft Placement: An Effective Procedure for Alleviation of Internal Nasal Valve Collapse.

PURPOSE: The internal nasal valve (INV) is one of the most commonly involved areas in patients with nasal obstruction, and surgeries such as turbinate reduction and septal recontouring affect the physiology of the nasal valve area. With this concept, a cohort study was conducted to evaluate spreader graft's effectiveness in recontouring INV and relieving nasal obstruction. MATERIALS AND METHODS: A prospective cohort study was performed, including patients with nasal obstruction with INV involvement. To quantify and compare the amount of nasal obstruction, Gertner Metal Plate (GMP) was used, and to assess the relief in associated symptoms, the Sino-Nasal Outcome Test (SNOT-22) was taken into consideration. GMP and SNOT 22 were performed preoperatively and postoperatively at the 1st, 3rd, and 6th months, and the data obtained were analyzed by SPSS 22 software using paired t-tests. The postoperative broadening of the nasal dorsum and patient satisfaction with the surgical outcome were also assessed using the visual analog scale . RESULTS: A total of 38 patients were included, with 28 males and 10 females. The data obtained were analyzed by using paired t-tests. Improvement in GMP assessment showed that t = 10.392305, 13.391485, and 15.985243 at the 1st, 3rd, and 6th months, respectively, with an overall P ≤ .00001 and relief in associated symptoms assessed by SNOT 22 computed t = 12.24228, 15.824486, and 18.046395 at the 1st, 3rd, and 6th month, respectively, with an overall P ≤ .00001. A total of 81.57% of patients did not perceive significant nasal dorsum broadening, and 86.84% of patients were highly satisfied by the surgical outcome based on the visual analog scale. CONCLUSION: We conclude that in patients with nasal obstruction associated with INV involvement, spreader grafting provides rapid relief of nasal obstruction and associated symptoms, does not cause any appreciable broadening, and provides a high degree of patient satisfaction. Hence, it should be preferred over septal recontouring and turbinate reduction.

The molecular basis of monopolin recruitment to the kinetochore.

The monopolin complex is a multifunctional molecular crosslinker, which in S. pombe binds and organises mitotic kinetochores to prevent aberrant kinetochore-microtubule interactions. In the budding yeast S. cerevisiae, whose kinetochores bind a single microtubule, the monopolin complex crosslinks and mono-orients sister kinetochores in meiosis I, enabling the biorientation and segregation of homologs. Here, we show that both the monopolin complex subunit Csm1 and its binding site on the kinetochore protein Dsn1 are broadly distributed throughout eukaryotes, suggesting a conserved role in kinetochore organisation and function. We find that budding yeast Csm1 binds two conserved motifs in Dsn1, one (termed Box 1) representing the ancestral, widely conserved monopolin binding motif and a second (termed Box 2-3) with a likely role in enforcing specificity of sister kinetochore crosslinking. We find that Box 1 and Box 2-3 bind the same conserved hydrophobic cavity on Csm1, suggesting competition or handoff between these motifs. Using structure-based mutants, we also find that both Box 1 and Box 2-3 are critical for monopolin function in meiosis. We identify two conserved serine residues in Box 2-3 that are phosphorylated in meiosis and whose mutation to aspartate stabilises Csm1-Dsn1 binding, suggesting that regulated phosphorylation of these residues may play a role in sister kinetochore crosslinking specificity. Overall, our results reveal the monopolin complex as a broadly conserved kinetochore organiser in eukaryotes, which budding yeast have co-opted to mediate sister kinetochore crosslinking through the addition of a second, regulatable monopolin binding interface.

Dual recognition of phosphoserine and phosphotyrosine in histone variant H2A.X by DNA damage response protein MCPH1.

Tyr142, the C-terminal amino acid of histone variant H2A.X is phosphorylated by WSTF (Williams-Beuren syndrome transcription factor), a component of the WICH complex (WSTF-ISWI chromatin-remodeling complex), under basal conditions in the cell. In response to DNA double-strand breaks (DSBs), H2A.X is instantaneously phosphorylated at Ser139 by the kinases ATM and ATR and is progressively dephosphorylated at Tyr142 by the Eya1 and Eya3 tyrosine phosphatases, resulting in a temporal switch from a postulated diphosphorylated (pSer139, pTyr142) to monophosphorylated (pSer139) H2A.X state. How mediator proteins interpret these two signals remains a question of fundamental interest. We provide structural, biochemical, and cellular evidence that Microcephalin (MCPH1), an early DNA damage response protein, can read both modifications via its tandem BRCA1 C-terminal (BRCT) domains, thereby emerging as a versatile sensor of H2A.X phosphorylation marks. We show that MCPH1 recruitment to sites of DNA damage is linked to both states of H2A.X.

Basal cell adenoma of the deep lobe of the parotid gland misdiagnosed as mucoepidermoid carcinoma - A case report.

INTRODUCTION: Basal cell adenoma is a rare, benign epithelial tumour of the salivary gland, comprising only 1-2 % of all salivary gland tumours. Predominantly found in the parotid gland, basal cell adenoma can also occur in minor salivary glands and are often confused with other benign and malignant salivary gland tumours. A thorough histopathological examination can provide a definitive diagnosis. PRESENTATION OF CASE: A 65-year-old woman presented with a painless mass in the right infra-auricular region. Imaging revealed a well-defined hypodense lesion in the deep lobe of the right parotid gland, initially suspected as mucoepidermoid carcinoma. Fine needle aspiration was inconclusive, leading to the decision to perform a total conservative parotidectomy. Histopathology confirmed basal cell adenoma, characterized by cystic areas filled with mucoid material and basaloid cells arranged in trabecular and tubular patterns. DISCUSSION: Basal cell adenoma was classified as a distinct entity by the WHO in 1991. Cytologically, they imitate both benign and malignant salivary as well as non-salivary gland tumours. The histological hallmark of basal cell adenoma involves basaloid cells with small round nuclei showing no atypia, scant pale cytoplasm, and distinct peripheral palisading. Treatment involves surgical removal, with a more radical approach for certain variants such as the membranous type. CONCLUSION: This case highlights the clinical, radiological, and histopathological features of basal cell adenoma, emphasizing the importance of accurate diagnosis and appropriate surgical management. Early detection and appropriate treatment are crucial for optimizing patient outcomes in basal cell adenoma management.

Optogenetics: Illuminating the Future of Hearing Restoration and Understanding Auditory Perception.

Hearing loss is a prevalent sensory impairment significantly affecting communication and quality of life. Traditional approaches for hearing restoration, such as cochlear implants, have limitations in frequency resolution and spatial selectivity. Optogenetics, an emerging field utilizing light-sensitive proteins, offers a promising avenue for addressing these limitations and revolutionizing hearing rehabilitation. This review explores the methods of introducing Channelrhodopsin- 2 (ChR2), a key light-sensitive protein, into cochlear cells to enable optogenetic stimulation. Viral- mediated gene delivery is a widely employed technique in optogenetics. Selecting a suitable viral vector, such as adeno-associated viruses (AAV), is crucial in efficient gene delivery to cochlear cells. The ChR2 gene is inserted into the viral vector through molecular cloning techniques, and the resulting viral vector is introduced into cochlear cells via direct injection or round window membrane delivery. This allows for the expression of ChR2 and subsequent light sensitivity in targeted cells. Alternatively, direct cell transfection offers a non-viral approach for ChR2 delivery. The ChR2 gene is cloned into a plasmid vector, which is then combined with transfection agents like liposomes or nanoparticles. This mixture is applied to cochlear cells, facilitating the entry of the plasmid DNA into the target cells and enabling ChR2 expression. Optogenetic stimulation using ChR2 allows for precise and selective activation of specific neurons in response to light, potentially overcoming the limitations of current auditory prostheses. Moreover, optogenetics has broader implications in understanding the neural circuits involved in auditory processing and behavior. The combination of optogenetics and gene delivery techniques provides a promising avenue for improving hearing restoration strategies, offering the potential for enhanced frequency resolution, spatial selectivity, and improved auditory perception.

Isolated Papillary Thyroid Carcinoma of the Pyramidal Lobe: A Case Report and Review of Literature.

The pyramidal lobe (PL) represents an embryological remnant of the thyroglossal duct. A solitary focus of papillary thyroid carcinoma (PTC) of the PL of thyroid gland is a rare entity. We present a case of a 33-year-old woman with PTC of the PL with lymph nodal involvement and further discuss the lines of surgical management for primary PTC arising from the PL of thyroid gland.

Molecular basis for the association of microcephalin (MCPH1) protein with the cell division cycle protein 27 (Cdc27) subunit of the anaphase-promoting complex.

Microcephalin (MCPH1), the first gene identified as causative for primary recessive autosomal microcephaly, is aberrantly expressed in autism-like disorders and human malignancy of breast and ovarian origin. MCPH1, the encoded protein product, has been implicated in various cellular processes including the DNA damage checkpoint, DNA repair, and transcription. Although our understanding of the cellular context in which MCPH1 operates continues to develop, a structural understanding of the C-terminal tandem BRCT domains of MCPH1 remains unexplored. Here, we identify cell division cycle protein 27 (Cdc27), a component of the anaphase-promoting complex (APC/C), as a novel interacting partner of MCPH1. We provide in vitro and in vivo evidence that the C-terminal tandem BRCT domains of MCPH1 (C-BRCTs) bind Cdc27 in a phosphorylation-dependent manner. To characterize this interaction further, we determined the structure of MCPH1 C-BRCTs in complex with a phosphorylated Cdc27 peptide (pCdc27) using x-ray crystallography. Based on this structure, we identified single amino acid mutations targeted at the binding interface that disrupted the MCPH1-pCdc27 interaction. Collectively, our data define the biochemical, structural, and cellular determinants of the novel interaction between MCPH1 and Cdc27 and suggest that this interaction may occur within the larger context of MCPH1-APC/C.

Radioactive Iodine in Differentiated Carcinoma of Thyroid: An Overview.

Thyroid cancer is the fifth most prevalent cancer in women and the fastest-growing malignancy. Although surgery is still the basis of treatment, internal radiation therapy [Brachytherapy] with radioactive iodine-131, which functions by releasing beta particles with low tissue penetration and causing DNA damage, is also a potential option. The three basic aims of RAI therapy in well-differentiated thyroid tumors are ablation of the remnant, adjuvant therapy, and disease management. Radioactive iodine dose is selected in one of two ways, empiric and dosimetric, which relies on numerous criteria. The dosage for ablation is 30-100 mCi, 30-150 mCi for adjuvant therapy, and 100-200 mCi for treatment. The RAI treatment effectively aids in the treatment to achieve complete removal of the disease and increase survival. The present review intends to emphasize the significance of radioactive iodine in the management of differentiated thyroid cancer and put forward the current breakthroughs in therapy.

Nuclear Imaging Modalities in the Diagnosis and Management of Thyroid Cancer.

In this review we have brought forward various nuclear imaging modalities used in the diagnosis, staging, and management of thyroid cancer. Thyroid cancer is the most common endocrine malignancy, accounting for approximately 3% of all new cancer diagnoses. Nuclear imaging plays an important role in the evaluation of thyroid cancer, and the use of radioiodine imaging, FDG imaging, and somatostatin receptor imaging are all valuable tools in the management of this disease. Radioiodine imaging involves the use of Iodine-123 [I-123] or Iodine-131 [I-131] to evaluate thyroid function and detect thyroid cancer. I-123 is a gamma-emitting isotope that is used in thyroid imaging to evaluate thyroid function and detect thyroid nodules. I-131 is a beta-emitting isotope that is used for the treatment of thyroid cancer. Radioiodine imaging is used to detect the presence of thyroid nodules and evaluate thyroid function. FDG imaging is a PET imaging modality that is used to evaluate the metabolic activity of thyroid cancer cells. FDG is a glucose analogue that is taken up by cells that are metabolically active, such as cancer cells. FDG PET/CT can detect primary thyroid cancer and metastatic disease, including lymph nodes and distant metastases. FDG PET/CT is also used to monitor treatment response and detect the recurrence of thyroid cancer. Somatostatin receptor imaging involves the use of radiolabeled somatostatin analogues to detect neuroendocrine tumors, including thyroid cancer. Radiolabeled somatostatin analogues, such as Indium-111 octreotide or Gallium-68 DOTATATE, are administered to the patient, and a gamma camera is used to detect areas of uptake. Somatostatin receptor imaging is highly sensitive and specific for the detection of metastatic thyroid cancer. Methods: A comprehensive search of relevant literature was done using online databases of PubMed, Embase, and Cochrane Library using the keywords "thyroid cancer," "nuclear imaging," "radioiodine imaging," "FDG PET/CT," and "somatostatin receptor imaging" to identify relevant studies to be included in this review. Conclusion: Nuclear imaging plays an important role in the diagnosis, staging, and management of thyroid cancer. The use of radioiodine imaging, thyroglobulin imaging, FDG imaging, and somatostatin receptor imaging are all valuable tools in the evaluation of thyroid cancer. With further research and development, nuclear imaging techniques have the potential to improve the diagnosis and management of thyroid cancer and other endocrine malignancies.

Solid/trabecular subtype of papillary thyroid carcinoma on cytology with focal differentiated high-grade thyroid carcinoma on histology: a cyto-histologic correlation.

Solid/trabecular subtype of papillary thyroid carcinoma (S/T PTC) is a rare entity that has been shown to have higher tumour recurrence and mortality rates. A definite diagnosis on fine needle aspiration cytology is often not easy. Rather, this entity may be misdiagnosed in cytology due to a lack of widespread features of classic PTC. We present a case of S/T PTC in a 61-year-old female, showing a focus on differentiated high-grade thyroid carcinoma (DHGTC) on histology. We discuss cytological features with the histologic correlation of S/T PTC and briefly discuss the newly introduced entity, DHGTC.

COVID-19 Associated Rhino-Orbital-Cerebral Mucormycosis: A Proposed Classification and Treatment Strategies.

OBJECTIVES: Mucormycosis is a rare fungal disease, which was known to affect only immunocompromised hosts, but during the COVID-19 pandemic, a surge in the cases of rhino- orbital-cerebral mucormycosis have been reported; however, the cause is still unknown. As the disease was a rare entity, there was no classification considering the spread and proper management at various stages. METHODS: Extensive literature search with the terms "mucormycosis," "invasive fungal sinusitis," "COVID-19 associated mucormycosis," and "mucormycosis in COVID-19" was conducted on Pubmed, Scopus, and Embase database, taking into consideration case histories, revealing the site of involvement and treatment based on the disease's extent. RESULTS: Relevant articles were analyzed, and it was found that there is no specific classification of the disease entity and no proper surgical and medical management guidelines to date, Conclusion: This review is an attempt to elaborate on the pathophysiology of mucormycosis and its spread and propose a classification that will help determine policies for the control and prevention of complications, morbidity, and mortality.

An Insight into Novel Sperm Cell Proteins as Bio-markers for Male Infertility: A Review.

Male infertility is rising nowadays and accounts for a major part of infertility cases worldwide. Novel tests are being developed for better detection and management of male infertility. Though there are many tests available for diagnosing male infertility like acrosome reaction rate, hemizona assay, in vivo or in vitro sperm penetration assay, sperm DNA damage tests, however, a semen analysis is the most commonly used initial test for male infertility. It is usually associated with failure to detect the cause in many cases, as seminal composition gets affected by a number of factors and can give false reports. Furthermore, it does not give any information about defects in capacitation, sperm-zona pellucida interaction, and sperm's ability to fertilize oocytes. This results in failure of detection and delayed management of male infertility. Hence, the present review was conducted to identify various sperm proteins that play a significant role in spermatogenesis, sperm motility, sperm-zona pellucida interaction, and fertilization. These proteins can be used in the future as markers of male infertility and will aid in better detection and management of male infertility. Methodology: Search for literature was made from 1970 to 2020 from various databases like PUBMED, SCOPUS, Google Scholar on sperm proteins and their role in male fertility using keywords: "sperm protein as bio-markers", "novel sperm proteins as markers of infertility", "Sperm proteins essential for capacitation, sperm motility and oocyte fertilization". Inclusion criteria: All full-length research articles, systematic reviews, meta-analyses, or abstracts on sperm proteins and male infertility published in the English language in peer-reviewed journals were considered.

Physiology to Disease Transmission of Respiratory Tract Infection: A Narrative Review.

INTRODUCTION: In the current scenario of the COVID 19 pandemic, the protective reflexes, namely sneeze and cough, have received great importance. However, it is not in terms of protection but in terms of the spread of infection. The present review tries to bring out the correlation between the physiology of sneeze and cough, taking into consideration the various receptors that initiate the two reflexes, then correlating it with the formation of expelled droplets and the significance of various aspects of droplets that lead to the spread of infection. MATERIAL AND METHODS: For the compilation of the present review, we searched the terms "Physiology of cough", "Physiology of sneeze", "droplets", "aerosols" and "Aerosols in COVID 19". The above-mentioned terms were extensively searched on PubMed, Google Scholar, and google search engine. After reviewing the various available material, the most significant research has been considered for this review. CONCLUSION: Through this review, we conclude that there are various factors responsible for the initiation of sneeze and cough, but in the case of infection, it is mainly the inflammatory reaction that directly stimulates the receptors to produce the reflex outburst air. As the flow of air during expiration is turbulent, it causes damage to the Epithelial Lining Fluid present in the respiratory conduit. In addition, it gets admixed with the saliva in the oropharynx and oral cavity and mucus in the nose to form droplets of various sizes. Large droplets settle close and are responsible for droplet and fomite transmission, but the smaller droplets remain suspended in the air and travel farther distances to cause airborne transmission. The spread of droplet cloud in sneezing may range to 6m or more as compared to cough; hence the concept of 1m to 2m of social distancing does not hold reliable if the patient is sneezing.

"Emerging role of Novel Seminal Plasma Bio-markers in Male Infertility: A Review".

Male infertility has emerged as an important cause of infertility worldwide. There are many factors affecting male fertility and research is going on to know impact of various factors on sperm functions. Semen analysis is gold standard diagnostic test for male infertility, but it is crude method for estimation of male infertility as seminal composition gets affected by environmental factors, infections, other pathologies, hence, results of semen analysis either becomes normal/ambiguous, leading to failure of diagnosis and delayed treatment. Hence, with need of newer, better tests for assessing male factor infertility, seminal plasma is being tested for biomarkers. Seminal plasma is considered gold mine for male fertility as it contains molecules from male reproductive glands which play important role in sperm function. Study of seminal plasma molecules can give an idea about sperm concentration, motility, morphology and cause of infertility and can serve as biomarkers for male infertility. Present review briefs on some of these novel seminal plasma biomarkers which may play significant role in male fertility and can be used in future for better identification, assessment of infertile males. METHODOLOGY: Literature from 1985 to 2019 was searched from various databases including PUBMED, SCOPUS, Google Scholar on seminal plasma biomarkers using keywords: "seminal plasma protein biomarkers", "novel seminal plasma markers and male infertility", "hormones in seminal plasma and male infertility", "oxidative stress and male infertility", "Reactive Oxygen Species and sperm DNA", "immunoinfertility". INCLUSION CRITERIA: All full length original or review articles or abstracts on seminal plasma markers and male infertility published in English language in various peer-reviewed journals were considered. EXCLUSION CRITERIA: Articles published in languages other than English were excluded from the study. RESULTS: Seminal plasma is a big reservoir of molecules derived from the various male reproductive glands which can be used as potential biomarkers of male fertility. CONCLUSION: Hence, seminal plasma biomarkers can be used in future for better assessment of male factor infertility, its causes and may play an important role in management of male factor infertility.

The budding-yeast RWD protein Csm1 scaffolds diverse protein complexes through a conserved structural mechanism.

RWD domains mediate protein-protein interactions in a variety of pathways in eukaryotes. In budding yeast, the RWD domain protein Csm1 is particularly versatile, assembling key complexes in the nucleolus and at meiotic kinetochores through multiple protein interaction surfaces. Here, we reveal a third functional context for Csm1 by identifying a new Csm1-interacting protein, Dse3. We show that Dse3 interacts with Csm1 in a structurally equivalent manner to its known binding partners Mam1 and Ulp2, despite these three proteins' lack of overall sequence homology. We theorize that the unique "clamp" structure of Csm1 and the loose sequence requirements for Csm1 binding have led to its incorporation into at least three different structural/signaling pathways in budding yeast.

Identification of H3K4me1-associated proteins at mammalian enhancers.

Enhancers act to regulate cell-type-specific gene expression by facilitating the transcription of target genes. In mammalian cells, active or primed enhancers are commonly marked by monomethylation of histone H3 at lysine 4 (H3K4me1) in a cell-type-specific manner. Whether and how this histone modification regulates enhancer-dependent transcription programs in mammals is unclear. In this study, we conducted SILAC mass spectrometry experiments with mononucleosomes and identified multiple H3K4me1-associated proteins, including many involved in chromatin remodeling. We demonstrate that H3K4me1 augments association of the chromatin-remodeling complex BAF to enhancers in vivo and that, in vitro, H3K4me1-marked nucleosomes are more efficiently remodeled by the BAF complex. Crystal structures of the BAF component BAF45C indicate that monomethylation, but not trimethylation, is accommodated by BAF45C's H3K4-binding site. Our results suggest that H3K4me1 has an active role at enhancers by facilitating binding of the BAF complex and possibly other chromatin regulators.

Effect of formative evaluation using direct observation of procedural skills in assessment of postgraduate students of obstetrics and gynecology: Prospective study.

INTRODUCTION: Direct Observation of Procedural Skills (DOPS) is a way of evaluating procedural skills through observation in the workplace. The purpose of this study was to assess the role of DOPS in teaching and assessment of postgraduate students and to know the effect of repeated DOPS on improvement of the skills and confidence of the students. METHODS: In both phases, significant difference was observed between the two groups on first DOPS comparison (1st phase: p=0.000; 2nd phase: p=0.002), with simulation group performing better. Comparison of sixth DOPS in the two groups revealed no difference in both phases, but significant difference on first and sixth DOPS comparison in each group (p=0.000). RESULTS: Repeated DOPS results in improved skills and confidence of students in managing real life obstetric emergencies irrespective of the teaching modality. CONCLUSION: Repeated DOPS results in improved skills and confidence of students in managing real life obstetric emergencies irrespective of the teaching modality.

Structural basis for dimerization and RNA binding of avian infectious bronchitis virus nsp9.

The potential for infection by coronaviruses (CoVs) has become a serious concern with the recent emergence of Middle East respiratory syndrome and severe acute respiratory syndrome (SARS) in the human population. CoVs encode two large polyproteins, which are then processed into 15-16 nonstructural proteins (nsps) that make significant contributions to viral replication and transcription by assembling the RNA replicase complex. Among them, nsp9 plays an essential role in viral replication by forming a homodimer that binds single-stranded RNA. Thus, disrupting nsp9 dimerization is a potential anti-CoV therapy. However, different nsp9 dimer forms have been reported for alpha- and beta-CoVs, and no structural information is available for gamma-CoVs. Here we determined the crystal structure of nsp9 from the avian infectious bronchitis virus (IBV), a representative gamma-CoV that affects the economy of the poultry industry because it can infect domestic fowl. IBV nsp9 forms a homodimer via interactions across a hydrophobic interface, which consists of two parallel alpha helices near the carboxy terminus of the protein. The IBV nsp9 dimer resembles that of SARS-CoV nsp9, indicating that this type of dimerization is conserved among all CoVs. This makes disruption of the dimeric interface an excellent strategy for developing anti-CoV therapies. To facilitate this effort, we characterized the roles of six conserved residues on this interface using site-directed mutagenesis and a multitude of biochemical and biophysical methods. We found that three residues are critical for nsp9 dimerization and its abitlity to bind RNA.