Pyrrothiogatain

NK cell were supplemented with IL-12 (10 ng/mL) (Becton Dickinson, San Jose, CA, USA) to have a positive control for IFN-gamma secretion [18 (link)].
The Beas-2B (ATCC CRL-9609) bronchial epithelial cell line was grown in BEGM culture medium (BEGM Kit Catalog No. CC-3170; Lonza/Clonetics Corporation, USA). The K562 (ATC CCL-243) lymphoblastoid cell line was cultured in Roswell Park Memorial Institute medium (RPMI) (Gibco, Milano, Italy) supplemented with 10% of FCS (fetal calf serum, Euroclone, Pero, MI, Italy) and 100 U/mL penicillin and 100 µg/mL streptomycin. Cell cultures were maintained at 37 °C in a humidified atmosphere of 5% CO₂ in air.
The role of HLA-E and NKG2A was evaluated by incubating the cells with anti-HLA-E (clone MEM-E/08, Exbio, Praha, CZ) or anti-CD94/NKG2A (clone 131411, BD, Italy) antibodies (7.5 ng/mL).
GATA3 DNA-binding activity was inhibited by adding pyrrothiogatain (cat#sc-352288A, Santa Cruz, CA, USA) to cell cultures (10 μM) [19 (link)].

Pyrrothiogatain (3-(2,5-dimethyl-1H-pyrrol-1yl) thiophene-2- carboxylic acid) was purchased from Santa Cruz (cat #sc-352288A). Cells were treated with the indicated concentration of Pyrrothiogatain (50 mM stock dissolved in DMSO) for 48 h prior to infection with SARS-CoV-2. Infection and subsequent immunofluorescence imaging were carried out as described above.