Journal Pre-proof J o u r n a l P r e -p r o o f Journal Pre-proof J o u r n a l P r e -p r o o f 3 Abstract Factors associated with negative conversion of SARS-CoV-2 RNA in hospitalized patients have not yet been systematically determined. We conducted a retrospective cohort study of COVID-19 patients in Qingdao, China. Both univariate and multivariate analysis were performed to identify independent factors for time to viral RNA negative conversion. Data on patients with re-detectable viral RNA after showing negative on RT-PCR test (intermittent negative status) were also analyzed. A total of 59 patients confirmed with COVID-19 were included in this study, with a median duration of 1 (interquartile range, IQR: 0-2) day from symptom onset to hospital admission. Median communicable period (from first day of positive nucleic acid test to first day of consecutive negative results) was 14 (IQR: 10-18) days, and 7 (IQR: 6-10) days for 10 patients with intermittent negative results. Age older than 45 years (hazard ratio, HR: 0.378; 95% confidence interval, CI: 0.205-0.698) and chest tightness (HR: 0.290; 95%CI: 0.091-0.919) were factors indepently affecting negative conversion of SARS-CoV-2 RNA. Headache (odds ratio: 7.553; 95%CI: 1.011-28.253)was significantly associated with intermittent negative status, with a predicted probability of 60%. Older age and chest tightness were independently associated with delayed clearance of SARS-CoV-2 RNA in hospitalized patients. These predictors would provide a new perspective on early identification of patients with prolonged viral shedding and facilitate optimal isolation protocols and treatment strategies.

MIND–Mis12 bridges the microtubule receptor complex and the inner kinetochore (see also a related paper by Petrovic et al. in this issue).

SummaryThe Ndc80 complex establishes end-on attachment of kinetochores to microtubules, which is essential for chromosome segregation. The Ndc80 subunit is characterized by an N-terminal region that binds directly to microtubules, and a long coiled-coil region that interacts with Nuf2. A loop region in Ndc80 that generates a kink in the structure disrupts the long coiled-coil region but the exact function of this loop, has until now, not been clear. Here we show that this loop region is essential for end-on attachment of kinetochores to microtubules in human cells. Cells expressing loop mutants of Ndc80 are unable to align the chromosomes, and stable kinetochore fibers are absent. Through quantitative mass spectrometry and immunofluorescence we found that the binding of the spindle and kinetochore associated (Ska) complex depends on the loop region, explaining why end-on attachment is defective. This underscores the importance of the Ndc80 loop region in coordinating chromosome segregation through the recruitment of specific proteins to the kinetochore.

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Tocilizumab has been reported to attenuate the “cytokine storm” in COVID-19 patients. We attempted to verify the effectiveness and safety of tocilizumab therapy in COVID-19 and identify patients most likely to benefit from this treatment. We conducted a randomized, controlled, open-label multicenter trial among COVID-19 patients. The patients were randomly assigned in a 1:1 ratio to receive either tocilizumab in addition to standard care or standard care alone. The cure rate, changes of oxygen saturation and interference, and inflammation biomarkers were observed. Thirty-three patients were randomized to the tocilizumab group, and 32 patients to the control group. The cure rate in the tocilizumab group was higher than that in the control group, but the difference was not statistically significant (94.12% vs. 87.10%, rate difference 95% CI–7.19%–21.23%, P = 0.4133). The improvement in hypoxia for the tocilizumab group was higher from day 4 onward and statistically significant from day 12 ( P = 0.0359). In moderate disease patients with bilateral pulmonary lesions, the hypoxia ameliorated earlier after tocilizumab treatment, and less patients (1/12, 8.33%) needed an increase of inhaled oxygen concentration compared with the controls (4/6, 66.67%; rate difference 95% CI–99.17% to–17.50%, P = 0.0217). No severe adverse events occurred. More mild temporary adverse events were recorded in tocilizumab recipients (20/34, 58.82%) than the controls (4/31, 12.90%). Tocilizumab can improve hypoxia without unacceptable side effect profile and significant influences on the time virus load becomes negative. For patients with bilateral pulmonary lesions and elevated IL-6 levels, tocilizumab could be recommended to improve outcome. Electronic Supplementary Material Supplementary material is available in the online version of this article at 10.1007/s11684-020-0824-3 and is accessible for authorized users.

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BackgroundBirt-Hogg-Dubé syndrome (BHD) is an autosomal dominant disorder, the main manifestations of which are fibrofolliculomas, renal tumors, pulmonary cysts and recurrent pneumothorax. The known causative gene for BHD syndrome is the folliculin (FLCN) gene on chromosome 17p11.2. Studies of the FLCN mutation for BHD syndrome are less prevalent in Chinese populations than in Caucasian populations. Our study aims to investigate the genotype spectrum in a group of Chinese patients with BHD.MethodsWe enrolled 51 patients with symptoms highly suggestive of BHD from January 2014 to February 2017. The FLCN gene was examined using PCR and Sanger sequencing in every patient, for those whose Sanger sequencing showed negative mutation results, multiplex ligation-dependent probe amplification (MLPA) testing was conducted to detect any losses of large segments.Main resultsAmong the 51 patients, 27 had FLCN germline mutations. In total, 20 mutations were identified: 14 were novel mutations, including 3 splice acceptor site mutations, 2 different deletions, 6 nonsense mutations, 1 missense mutation, 1 small insertion, and 1 deletion of the whole exon 8.ConclusionsWe found a similar genotype spectrum but different mutant loci in Chinese patients with BHD compared with European and American patients, thus providing stronger evidence for the clinical molecular diagnosis of BHD in China. It suggests that mutation analysis of the FLCN gene should be systematically conducted in patients with cystic lung diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s13023-017-0656-7) contains supplementary material, which is available to authorized users.

The functions of cohesin are central to genome integrity, chromosome organization and transcription regulation through its prevention of premature sister-chromatid separation and the formation of DNA loops. The loading of cohesin onto chromatin depends on the Scc2–Scc4 complex; however, little is known about how it stimulates the cohesion-loading activity. Here we determine the large ‘hook' structure of Scc2 responsible for catalysing cohesin loading. We identify key Scc2 surfaces that are crucial for cohesin loading in vivo. With the aid of previously determined structures and homology modelling, we derive a pseudo-atomic structure of the full-length Scc2–Scc4 complex. Finally, using recombinantly purified Scc2–Scc4 and cohesin, we performed crosslinking mass spectrometry and interaction assays that suggest Scc2–Scc4 uses its modular structure to make multiple contacts with cohesin.