90K/LGALS3BP expression is upregulated in COVID-19 but may not restrict SARS-CoV-2 infection.

Glycoprotein 90K, encoded by the interferon-stimulated gene LGALS3BP, displays broad antiviral activity. It reduces HIV-1 infectivity by interfering with Env maturation and virion incorporation, and increases survival of Influenza A virus-infected mice via antiviral innate immune signaling. Its antiviral potential in SARS-CoV-2 infection remains largely unknown. Here, we analyzed the expression of 90K/LGALS3BP in 44 hospitalized COVID-19 patients at multiple levels. We quantified 90K protein concentrations in serum and PBMCs as well as LGALS3BP mRNA levels. Complementary, we analyzed two single cell RNA-sequencing datasets for expression of LGALS3BP in respiratory specimens and PBMCs from COVID-19 patients. Finally, we analyzed the potential of 90K to interfere with SARS-CoV-2 infection of HEK293T/ACE2, Calu-3 and Caco-2 cells using authentic virus. 90K protein serum concentrations were significantly elevated in COVID-19 patients compared to uninfected sex- and age-matched controls. Furthermore, PBMC-associated concentrations of 90K protein were overall reduced by SARS-CoV-2 infection in vivo, suggesting enhanced secretion into the extracellular space. Mining of published PBMC scRNA-seq datasets uncovered monocyte-specific induction of LGALS3BP mRNA expression in COVID-19 patients. In functional assays, neither 90K overexpression in susceptible cell lines nor exogenous addition of purified 90K consistently inhibited SARS-CoV-2 infection. Our data suggests that 90K/LGALS3BP contributes to the global type I IFN response during SARS-CoV-2 infection in vivo without displaying detectable antiviral properties in vitro.

SARS-CoV-2 variant Alpha has a spike-dependent replication advantage over the ancestral B.1 strain in human cells with low ACE2 expression.

Epidemiological data demonstrate that Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) Alpha and Delta are more transmissible, infectious, and pathogenic than previous variants. Phenotypic properties of VOC remain understudied. Here, we provide an extensive functional study of VOC Alpha replication and cell entry phenotypes assisted by reverse genetics, mutational mapping of spike in lentiviral pseudotypes, viral and cellular gene expression studies, and infectivity stability assays in an enhanced range of cell and epithelial culture models. In almost all models, VOC Alpha spread less or equally efficiently as ancestral (B.1) SARS-CoV-2. B.1. and VOC Alpha shared similar susceptibility to serum neutralization. Despite increased relative abundance of specific sgRNAs in the context of VOC Alpha infection, immune gene expression in infected cells did not differ between VOC Alpha and B.1. However, inferior spreading and entry efficiencies of VOC Alpha corresponded to lower abundance of proteolytically cleaved spike products presumably linked to the T716I mutation. In addition, we identified a bronchial cell line, NCI-H1299, which supported 24-fold increased growth of VOC Alpha and is to our knowledge the only cell line to recapitulate the fitness advantage of VOC Alpha compared to B.1. Interestingly, also VOC Delta showed a strong (595-fold) fitness advantage over B.1 in these cells. Comparative analysis of chimeric viruses expressing VOC Alpha spike in the backbone of B.1, and vice versa, showed that the specific replication phenotype of VOC Alpha in NCI-H1299 cells is largely determined by its spike protein. Despite undetectable ACE2 protein expression in NCI-H1299 cells, CRISPR/Cas9 knock-out and antibody-mediated blocking experiments revealed that multicycle spread of B.1 and VOC Alpha required ACE2 expression. Interestingly, entry of VOC Alpha, as opposed to B.1 virions, was largely unaffected by treatment with exogenous trypsin or saliva prior to infection, suggesting enhanced resistance of VOC Alpha spike to premature proteolytic cleavage in the extracellular environment of the human respiratory tract. This property may result in delayed degradation of VOC Alpha particle infectivity in conditions typical of mucosal fluids of the upper respiratory tract that may be recapitulated in NCI-H1299 cells closer than in highly ACE2-expressing cell lines and models. Our study highlights the importance of cell model evaluation and comparison for in-depth characterization of virus variant-specific phenotypes and uncovers a fine-tuned interrelationship between VOC Alpha- and host cell-specific determinants that may underlie the increased and prolonged virus shedding detected in patients infected with VOC Alpha.

Interaction of polycyclic musks and UV filters with the estrogen receptor (ER), androgen receptor (AR), and progesterone receptor (PR) in reporter gene bioassays.

Two important ingredients of personal care products, namely polycyclic musk fragrances and UV filters, can be found in the environment and in humans. In previous studies, several compounds of both classes have been tested for their interaction with the estrogen receptor. Two polycyclic musk fragrances, namely AHTN and HHCB, turned out to be anti-estrogenic both in vitro and in vivo in a transgenic zebrafish assay. Several UV filters have been shown to exert estrogenic effects in vitro and in some in vivo studies. Here, we assessed the interaction of five polycyclic musk compounds and seven UV filters with the estrogen receptor (ER), androgen receptor (AR), and progesterone (PR) receptor, using sensitive and specific reporter gene cell lines. Four polycyclic musks (AHTN, HHCB, AETT, and AHMI) were found to be antagonists toward the ERbeta, AR and PR. The UV filters that showed estrogenic effects (benzophenone-3, Bp-3; 3-benzylidene camphor, 3-BC; homosalate, HMS; and 4-methylbenzylidene camphor, 4-MBC) were found to be antagonists toward the AR and PR. The ERalpha agonistic UV filter octyl-dimethyl-p-aminobenzoic acid (OD-PABA) did not show activity toward the AR and PR. Octyl methoxy cinnamate (OMC) showed weak ERalpha agonism, but potent PR antagonism. Butyl methoxydibenzoylmethane (B-MDM) only showed weak ERalpha agonism and weak AR antagonism. Most effects were observed at relatively high concentrations (above 1 muM); however, the anti-progestagenic effects of the polycyclic musks AHMI and AHTN were detected at concentrations as low as 0.01 muM. The activity of anti-progestagenic xenobiotics at low concentrations indicates the need to undertake more research to find out about the potential endocrine disrupting effects of these compounds in vivo.