Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90.

SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) is the causative agent of the COVID19 pandemic. The SARS-CoV-2 genome encodes for a small accessory protein termed Orf9b, which targets the mitochondrial outer membrane protein TOM70 in infected cells. TOM70 is involved in a signaling cascade that ultimately leads to the induction of type I interferons (IFN-I). This cascade depends on the recruitment of Hsp90-bound proteins to the N-terminal domain of TOM70. Binding of Orf9b to TOM70 decreases the expression of IFN-I; however, the underlying mechanism remains elusive. We show that the binding of Orf9b to TOM70 inhibits the recruitment of Hsp90 and chaperone-associated proteins. We characterized the binding site of Orf9b within the C-terminal domain of TOM70 and found that a serine in position 53 of Orf9b and a glutamate in position 477 of TOM70 are crucial for the association of both proteins. A phosphomimetic variant Orf9bS53E showed drastically reduced binding to TOM70 and did not inhibit Hsp90 recruitment, suggesting that Orf9b-TOM70 complex formation is regulated by phosphorylation. Eventually, we identified the N-terminal TPR domain of TOM70 as a second binding site for Orf9b, which indicates a so far unobserved contribution of chaperones in the mitochondrial targeting of the viral protein.

Antibiotic prescription rate for upper respiratory tract infections in general practice - a Croatian study.

Aim Acute upper respiratory tract infections are the most common indication for antibiotic prescribing in primary care. The frequency of antibiotics prescribed to patients with upper respiratory tract infections has been studied. Methods A cross-sectional survey has been conducted from October to December 2017 with participation of 17 General Practices in Croatia. Visits of patients with symptoms of upper respiratory tract infections made the survey framework. A specially structured questionnaire was a tool for the assessment of reasons for encounters, clinical statuses and practitioners' decisions. According to the registered clinical status, two independent doctors classified patients using Centor criteria, which were used to evaluate appropriate indication for antibiotic prescription. Results The survey analysed 709 patients' visits. Seventy three percent of general practice visits due to acute respiratory tract infection were initial. Antibiotics had been prescribed in the total of 32.6% of cases. Analysing upper respiratory tract infections with Centor score 1 and 2, antibiotics had been prescribed in 22.8% cases of the first visit and in 31.7% of the second visit, and with the Centor score 3 and 4 antibiotics had been prescribed in 85.1% of cases during the initial visit, and 66.7% of cases during the second visit. Logistic regression analysis about the contribution of nonclinical variables to antibiotic prescribing showed that most often antibiotics had been prescribed on Mondays. The most prescribed group of antibiotics was the penicillin group (74.8%), followed by macrolides (17.6%) and cephalosporins (4.0%). Conclusion General practitioners prescribe antibiotics for acute respiratory tract infections more than recommended by evidence based medicine and professional criteria.