Figure 5.

The propeller domain of B9 is required for sporozoite infectivity but does not restrict host receptor usage

(A) Strategy used to genetically complement PbΔb9 with different versions of B9 (indicated as B9∗) by double crossover homologous recombination.

(B) Schematic representation of the B9 constructs used for genetic complementation. SP, signal peptide, GPI, glycosylphosphatidylinositol.

(C) Infection rates were determined by quantification of EEFs (GFP-positive cells) 24 h after infection of HepG2 cell cultures with sporozoites of PbGFP, PbΔb9, or PbΔb9 complemented with PbB9, Δprop, ΔpropΔ6cys1, PyProp, PyProp6cys1, or PfProp constructs. Data are represented as % of GFP-positive cells (mean ± SEM). Each dot represents the mean value in one experiment. ∗∗p < 0.01 as compared to PbGFP (one-way ANOVA followed by Dunnett’s multiple comparisons test).

(D) Infection rates in HepG2 or HepG2/CD81 cells infected with PbΔb9 or PbΔb9 complemented with PyProp or Pyprop6cys1 constructs were determined 24 h postinfection. The results show the percentage of invaded (GFP-positive) cells as determined by FACS (mean ± SEM). ns, nonsignificant, ∗p < 0.05 (two-way ANOVA followed by Sidak’s multiple comparisons test).

(E) Immunofluorescence analysis of sporozoites from PbGFP and PbΔb9 complemented with Pfprop-Flag or Δprop-Flag constructs, labeled with anti-Flag antibodies (red). Parasites express GFP (green) and nuclei were stained with Hoechst 33342 (blue). Scale bars, 5 μm.

(F) Infection rates were determined 24 h after infection of HepG2 cell cultures with sporozoites of PbB9-Flag or PbΔb9 complemented with Δprop-Flag or Pfprop-Flag constructs. Results are expressed as % of control (PbB9-Flag). See also Figures S10 and S11, and Table S6 for quantitative source data.