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Crimean-Congo hemorrhagic fever virus (CCHFV) is a biosafety level-4 and World Health Organization-recognized priority pathogen requiring urgent Research & Development attention. The glycoproteins of CCHFV, Gn and Gc, are considered to play multiple roles in the viral life cycle by interactions with host cells; however, these interactions remain largely unclear to date. Here, we analyzed the cellular interactomes of CCHFV glycoproteins by affinity purification-quantitative mass spectrometry and identified 45 host proteins as high-confidence Gn/Gc interactors. These host molecules are involved in transmembrane transport, metabolism, oxidative stress, protein folding and glycosylation, etc., in line with the potential physiological actions of the viral glycoproteins. As a proof of principle, we detailedly elucidated the role of an identified cellular protein, HAX1, and therefore uncovered an interesting host antiviral strategy. HAX1 interacts with Gn by its C-terminus, while its N-terminal region leads to mitochondrial localization. By the strong interaction, HAX1 sequestrates Gn to mitochondria, thus depriving Gn of its normal Golgi localization that is indispensable for functional glycoprotein-mediated progeny virion packaging. Consistently, the notable inhibitory activity of HAX1 against viral packaging and hence propagation was further elucidated in the contexts of pseudotyped and authentic CCHFV infections by series of loss/gain-of-function assays in cellular and animal infection models. Together, the findings not only provide a systematic CCHFV Gn/Gc-host protein-protein interaction map, but also unravel a novel cellular antiviral mechanism, which may facilitate further studies on CCHFV biology and therapeutic approaches.

Enterovirus 71 (EV71), a small, single-stranded, positive-sense RNA virus belonging to the enterovirus genus in the family Picornaviridae, causes hand, foot, and mouth disease. Although EV71 seriously threatens to public health, no effective antiviral drugs are available for treating this disease. In this study, we found that ML390, a dihydroorotate dehydrogenase inhibitor, has potential anti-EV71 activity. ML390 dose-dependently inhibited EV71 replication with IC

Protein phosphorylation is one of the most common and important post-translational modifications and is involved in many biological processes, including DNA damage repair, transcriptional regulation, signal transduction, and apoptosis regulation. The use of antibodies targeting phosphorylated protein is a convenient method to detect protein phosphorylation. Therefore, high-quality antibodies are essential, and uniform and effective standards are urgently needed to evaluate the quality of these phosphorylation-specific antibodies. In this study, we established a simple, broad-spectrum system for the preparation of phosphorylation-positive samples. The positive samples for evaluation of phosphorylation-specific antibodies were then validated in cells from different species and tissues, and also been proven effectively in western blot, enzyme-linked immunosorbent assays, LC-MS/MS and immunofluorescence analysis. Overall, our findings established a novel approach for evaluation of the quality of phosphorylation-specific antibodies and may have applications in various biomedical fields.

Leaf color mutants are ideal materials for chloroplast development and photosynthetic mechanism research. Here, we characterized an EMS (ethyl methane sulfonate)-mutagenized sorghum (Sorghum bicolor) mutant, sbe6-a1, in which the severe disruption in chloroplast structure and a chlorophyll deficiency promote an albino leaf phenotype and lead to premature death. The proteomic analyses of mutant and its progenitor wild-type (WT) were performed using a Q Exactive plus Orbitrap mass spectrometer and 4,233 proteins were accurately quantitated. The function analysis showed that most of up-regulated proteins in mutant sbe6-a1 had not been well characterized. GO-enrichment analysis of the differentially abundant proteins (DAPs) showed that up-regulated DAPs were significantly enriched in catabolic process and located in mitochondria, while down regulated DAPs were located in chloroplasts and participated in photosynthesis and some other processes. KEGG pathway-enrichment analyses indicated that the degradation and metabolic pathways of fatty acids, as well as some amino acids and secondary metabolites, were significantly enhanced in the mutant sbe6-a1, while photosynthesis-related pathways, some secondary metabolites' biosynthesis and ribosomal pathways were significantly inhibited. Analysis also shows that some DAPs, such as FBAs, MDHs, PEPC, ATP synthase, CABs, CHLM, PRPs, pathogenesis-related protein, sHSP, ACP2 and AOX may be closely associated with the albino phenotype. Our analysis will promote the understanding of the molecular phenomena that result in plant albino phenotypes.

This study describes a new method for protein fixation that has high, stable, and good repeatable fixation effect without deactivating protein. This method involves intercalating bovine serum albumin (BSA) into montmorillonite (MMT) under different experimental conditions, i.e., the concentration of coupling agent KH570, pH values, ionic strength, and the impact of buffers. The study showed the interaction of buffer systems with pH values had the most significant influence on adsorption quantity. The existence of the buffer increased the adsorption quantity and helped with slow protein releasing, while the phosphate buffer worked better than acetate buffer. More BSA was adsorbed on KH570 treated than untreated MMT, and among the treated samples the most protein released was on MMT treated with KH570 in 100% ethanol and in the acetate buffer solution. Adsorption quantity of the BSA initially increased and then decreased with the increase of the ionic strength. The research on the preparation of MMT/BSA materials provides further information on the study of protein composites.