Estimation of seroprevalence of novel coronavirus disease (COVID-19) using preserved serum at an outpatient setting in Kobe, Japan: A cross-sectional study.

OBJECTIVES: Coronavirus disease 2019 (COVID-19) pandemic caused by SARS-CoV-2 has been affecting many people on earth and our society. Japan is known to have relatively smaller number of its infections as well as deaths among developed nations. However, accurate prevalence of COVID-19 in Japan remains unknown. Therefore, we conducted a cross-sectional study to estimate seroprevalence of SARS-CoV-2 infection. METHODS: We conducted a cross-sectional serologic testing for SARS-CoV-2 antibody using 1000 samples from patients at outpatient settings who visited the clinic from March 31 to April 7, 2020, stratified by the decade of age and sex. RESULTS: There were 33 positive IgG among 1000 serum samples (3.3%, 95%CI: 2.3-4.6%). By applying this figure to the census of Kobe City (population: 1,518,870), it is estimated that the number of people with positive IgG be 50,123 (95%CI: 34,934-69,868). Age and sex adjusted prevalence of positivity was calculated 2.7% (95%CI: 1.8-3.9%), and the estimated number of people with positive IgG was 40,999 (95%CI: 27,333-59,221). These numbers were 396 to 858-fold more than confirmed cases with PCR testing in Kobe City. CONCLUSIONS: Our cross-sectional serological study suggests that the number of people with seropositive for SARS-CoV-2 infection in Kobe, Japan is far more than the confirmed cases by PCR testing.

Src-dependent phosphorylation of the EGF receptor Tyr-845 mediates Stat-p21waf1 pathway in A431 cells.

BACKGROUND: Cell surface receptor for the epidermal growth factor (EGFR) and cytoplasmic tyrosine kinase c-Src co-operate in several cellular functions such as proliferation and apoptosis. Our previous studies have shown that ectopic expression of the adaptor protein p52shc or p66shc, but not p46shc, and EGF stimulation lead to the activation of c-Src that is accompanied by phosphorylation of signal transducers and activators of transcription (Stat) in A431 cells. RESULTS: Here, we show that by using A431 cells as a model system, expression of p52shc, or cell stimulation with EGF or H2O2 leads to phosphorylation of EGFR on Tyr 845 that is located to the activation segment of the catalytic domain. The phosphorylation of Tyr 845 can be inhibited by PP2, but not by AG1478, and is associated with Src activation and Stat 3/5 phosphorylation, but not with MAP (mitogen-activated protein) kinase phosphorylation. Phosphorylation of Stat 3/5 in response to p52shc expression, EGF or H2O2 could also be inhibited by introduction into cells of phospho-Tyr 845-specific antibody or by expression of dominant-negative version of c-Src. Co-incubation of purified c-Src and EGFR results in phosphorylation of Tyr 845 in vitro, indicating that c-Src can directly phosphorylate EGFR on Tyr 845. CONCLUSION: These results indicate that multiple signals for c-Src activation can promote Stat 3/5 phosphorylations through Src-dependent phosphorylation of EGFR on Tyr 845.

Inhibition and activation of c-Src: the head and tail of a coin.

Protein-tyrosine kinases (PTKs) play pivotal roles in many cell systems. The Src family kinases (SFKs) are the most characterized PTKs shown to be coupled with various cell surface receptors. However, their mode of activation and regulating partners are largely unknown. Here we describe a novel mechanism of inhibition and activation of c-Src, a representative of the SFKs. Both directions of regulation take place at the same site in the catalytic domain of c-Src via a peptide- or protein-protein interaction. Our results highlight a novel and general mode of kinase regulation that may be applied not only to SFKs, but to other PTKs and Ser/Thr kinases as well.

Adaptor protein Shc is an isoform-specific direct activator of the tyrosine kinase c-Src.

The activity of c-Src protein-tyrosine kinase is up-regulated under a number of receptor signaling pathways. However, the activation mechanism of c-Src under physiological conditions has remained unclear. We show here that the Shc adaptor protein is a novel direct activator of c-Src in epidermal growth factor receptor signaling in A431 human epidermoid carcinoma cells. Among the three Shc isoforms, P66 and P52, but not P46, were found to interact with and activate c-Src in vitro and in vivo. Activation of c-Src accompanied autophosphorylation of c-Src in the activation segment, but the carboxyl-terminal dephosphorylation was not observed. We have identified the interaction sites between Shc and c-Src and constructed a point mutant of Shc that abolishes the c-Src activation. Using this mutant, we have confirmed that the Shc-mediated c-Src activation triggers Stat-p21/WAF1/Cip1 pathway that has been implicated in the cell cycle arrest and apoptosis of epidermal growth factor-stimulated A431 cells.