Fig 1.
Quantitative comparison of protein expression between two P. vivax salivary gland sporozoite field isolates.
(A) Protein abundances based on spectral counts were estimated using the normalized spectral abundance factor (NSAF). Each point represents the natural log-transformed NSAF value of a protein, comparing its ln(NSAF) value in either sample. Deciles of relative abundance within each sample are shown (dashed gray lines). For each protein observed in both of the two P. vivax salivary gland sporozoite samples, the natural log of the protein ratio of the NSAF values observed in the VK210 sample and the VK247 sample was calculated as ln(NASF)VK210-ln(NSAF)VK247. The population of these values produced a normal distribution centered near zero, corresponding to a mean ratio of 1:1 (S1 Fig). Proteins identified in both samples are color-coded to indicate the deviation of their log-transformed protein ratio R from the population mean as determined from the fit curve. Deviation from the mean was low at high abundances and increased with decreasing spectral counts. The cyan and orange points represent proteins identified in only one isolate or the other. (B) Protein ratios were calculated based on the adjusted and normalized spectral counts used to calculate the G statistic. The population of log-transformed protein ratios of proteins detected in both samples assumed a Gaussian distribution with a mean near zero. The mean and standard deviation from this distribution were used to calculate p-values using the complementary error function (ERFC). (C) The protein ratios of all proteins detected in either sample are plotted with respect to the sum of the adjusted and normalized PSM from both samples used to calculate the ratio for each protein. Points in red are proteins with that were not significant at a 5.0% false discovery rate (FDR) according to the G-test. Points in green are proteins with ratios that were not significant at a 5.0% FDR according to the ERFC. Points in blue are proteins ratios that were significant by both cut-offs. Points inside of dashed boxes represent proteins detected in only one sample or the other. Protein ratios were estimated for these proteins by increasing all spectral counts by one in order to give all proteins non-zero values.
Fig 2.
Quantitative comparison of protein expression between P. vivax and P. falciparum salivary gland sporozoites.
(A) Protein abundances based on spectral counts were estimated using the normalized spectral abundance factor (NSAF). Each point represents the natural log-transformed NSAF value of a protein, comparing its ln(NSAF) value in either sample. P. vivax ln(NSAF) values are the average of the values observed in the two field isolate samples. Deciles of relative abundance within each sample are shown (dashed gray lines). For each protein with orthologs detected in both P. falciparum and P. vivax salivary gland sporozoites, the natural log of the protein ratio of the NSAF values observed in the P. falciparum sample and the P. vivax sample was calculated as ln(NASF)P.falciparum-ln(NSAF)P.vivax. The population of these values produced a normal distribution centered near zero, corresponding to a mean ratio of 1:1 (S2 Fig). Protein orthologs detected in both species are color-coded to indicate the deviation of their log-transformed protein ratio R from the population mean as determined from the fit curve. Deviation from the mean was low at high abundances and increased with decreasing spectral counts. The cyan and orange points represent protein orthologs identified in only one species or the other. Diamond points represent proteins with no ortholog in the other species. (B) Protein ratios were calculated based on the adjusted and normalized spectral counts used to calculate the G statistic. The population of log-transformed protein ratios of proteins detected in both samples assumed a Gaussian distribution with a mean near zero. The mean and standard deviation from this distribution were used to calculate p-values using the complementary error function (ERFC). (C) The protein ratios of all protein orthologs detected in either species are plotted with respect to the sum of the adjusted and normalized PSM from both samples used to calculate the ratio for each protein. Points in red are proteins with that were not significant at a 5.0% false discovery rate (FDR) according to the G-test. Points in green are proteins with ratios that were not significant at a 5.0% FDR according to the ERFC. Points in blue are proteins ratios that were significant by both cut-offs. Points inside of dashed boxes represent protein orthologs detected in only one species or the other. Protein ratios were estimated for these proteins by increasing all spectral counts by one in order to give all proteins non-zero values.
Table 1.
Highly abundant proteins identified in P. vivax salivary gland sporozoites.
Fig 3.
Glycosylation of CSP and TRAP in P. vivax and P. falciparum salivary gland sporozoites.
The glycosylated portions of the conserved thrombospondin repeat (TSR) domains of P. falciparum and P. vivax CSP and TRAP are aligned. The conserved glycosylation motif is highlighted. Residues that are putatively glycosylated according to the MS evidence are colored as shown in the legend. Information on glycosylation of P. falciparum is from [20]. Only TRAP in P. falciparum salivary gland sporozoites exhibited evidence for C-mannosylation. Both CSP and TRAP exhibited evidence for O-fucosylation in both species. In P. falciparum, a portion of CSP and TRAP also showed evidence for modification with a fucose-glucose disaccharide. No evidence for the disaccharide was observed in the P. vivax samples.
Table 2.
Proteins phosphorylated in sporozoites but not in blood stagesa.
Table 3.
Phosphorylated sporozoite-specific proteinsa.
Table 4.
Putatively surface-exposed proteins on P. vivax salivary gland sporozoites.
Fig 4.
Comparison Invasion-related proteins identified in P. vivax and P. falciparum salivary gland sporozoites.
Proteins identified from proteomic analyses of P. vivax and P. falciparum salivary gland sporozoites were compared against a compendium of known and putative invasion-related Plasmodium proteins (S10 Table). Proportional Venn diagrams show the number of proteins in each category identified only in P. vivax (cyan), only in P. falciparum (orange), in both species (green) and neither species (gray). The total number of proteins identified in each species and the total number of proteins in the category are listed below each diagram.