Effect of ENV59-GP3 peptide on calcium influx following Ca2+ depletion in THP-1 cells
To measure the effects of ENV59-GP3 peptide and NS6180 blocker on calcium flux, THP-1 cells were loaded with the calcium sensitive dye Fluo-4 and incubated with either ENV59-GP3 peptide or NS6180 blocker, and/or with the calcium store depleting agent Thapsigargin, an inhibitor of the sarco/endoplasmic reticulum Ca2+ ATPase (SERCA) [25] (Figure 4A). Similarly, both ENV59-GP3 peptide and NS6180 reduced the influx-mediated Ca2+ accumulation induced by Thapsigargin. In comparison, an immune suppressive peptide isolated from the influenza hemagglutinin (Flu ISU) which has a similar immunosuppressive phenotype to ENV59-GP3 [21], but interacts with and inhibits dimerization of the stimulator of interferon genes (STING) [21] did not have a significant effect in the [Ca2+]I in this assay. Thus, the ENV59-GP3 peptide’s biological activity is more similar to the KCa3.1 inhibitor than it is to an influenza virus-derived immunosuppressive peptide.