RESUMO
The genome of SARS-CoV-2 (SARS2) encodes for two viral proteases (NSP3/ papain-like protease and NSP5/ 3C-like protease or major protease) that are responsible for cleaving viral polyproteins for successful replication. NSP3 and NSP5 of SARS-CoV (SARS1) are known interferon antagonists. Here, we examined whether the protease function of SARS2 NSP3 and NSP5 target proteins involved in the host innate immune response. We designed a fluorescent based cleavage assay to rapidly screen the protease activity of NSP3 and NSP5 on a library of 71 human innate immune proteins (HIIPs), covering most pathways involved in human innate immunity. By expressing each of these HIIPs with a genetically encoded fluorophore in a cell-free system and titrating in the recombinant protease domain of NSP3 or NSP5, we could readily detect cleavage of cognate HIIPs on SDS-page gels. We identified 3 proteins that were specifically and selectively cleaved by NSP3 or NSP5: IRF-3, and NLRP12 and TAB1, respectively. Direct cleavage of IRF3 by NSP3 could explain the blunted Type- I IFN response seen during SARS-CoV-2 infections while NSP5 mediated cleavage of NLRP12 and TAB1 point to a molecular mechanism for enhanced production of IL-6 and inflammatory response observed in COVID-19 patients. Surprisingly, both NLRP12 and TAB1 have each two distinct cleavage sites. We demonstrate that in mice, the second cleavage site of NLRP12 is absent. We pushed this comparative alignment of IRF-3 and NLRP12 homologs and show that the lack or presence of cognate cleavage motifs in IRF-3 and NLRP12 could contribute to the presentation of disease in cats and tigers, for example. Our findings provide an explanatory framework for in-depth studies into the pathophysiology of COVID-19 and should facilitate the search or development of more effective animal models for severe COVID-19. Finally, we discovered that one particular species of bats, Davids Myotis, possesses the five cleavage sites found in humans for NLRP12, TAB1 and IRF3. These bats are endemic from the Hubei province in China and we discuss its potential role as reservoir for the evolution of SARS1 and SASR2.
RESUMO
The non-invasive quantification of human tissue fat fraction using easily scalable and accessible imaging technologies is crucial for the diagnosis of many diseases including liver steatosis. Here, we propose a non-invasive quantification of fat content using a highly accessible ultrasonic imaging technology. Ultrasonic echoes backscattered from human liver tissues are recombined to synthetize echoes of a virtual point-like reflector within the organs. This virtual point-like reflector is an ultrasonic analogue of artificial stars generated by laser beams in the field of astronomy, which are used to estimate the aberrations induced in the propagation medium. Here, the ultrasonic echoes from the point-like reflector provide an estimate of the Green's function relating the ultrasonic array and the reflector location and consequently represent a measurement of the aberrations induced along the ultrasonic beam travel path. Maximizing the spatial coherence of echoes backscattered from this targeted region provides an estimate of the acoustic sound speed while iteratively making the reflector more echogenic. The acoustic sound speed is dependent of the organ fat content, and we derive and cross-validate a theoretical equation relating acoustic sound speed and fat content both in phantom experiments and humans. An ultrasound-based fat fraction was found to be highly correlated with the oil paraffin concentration (R 2 = 0.985) in phantoms and well correlated with the gold standard magnetic resonance imaging proton density fat fraction measurements (R 2 = 0.73) in patients.
