Fig. 5.

Use of CRISPR/Cas9 to generate BSG knockout in JK-1 cells. (A) The ΔBSG clone has no surface expression of BSG but retains expression of known host receptors (GypA, GypC, and CR1) at levels comparable to the levels in jkRBCs. (Scale bars: 10 μm.) (B) Quantitative surface proteomics analysis comparing the abundance of 237 surface-membrane proteins in WT jkRBCs and ΔBSG jkRBCs confirms the specific loss of BSG in the ΔBSG cells. (C) The ΔBSG-knockout JK-1 cell line is refractory to P. falciparum Invasion. Invasion of the sialic acid-independent P. falciparum strain 3D7 and sialic acid-dependent P. falciparum strain Dd2 was completely inhibited in two independent clones of ΔBSG. Data are normalized to the invasion efficiency of WT JK-1 cells and are representative of three biological replicates. Error bars represent SDs from three biological replicates. P values were calculated using a two-tailed t test (GraphPad Prism version 7.01).