Figure 2
Figure 2. PfPTP1 is localized on Maurer's clefts in P falciparum–infected RBCs. (A) Immunofluorescence assay on CS2/PfPTP1-GFP cell line: the GFP signal (green) colocalizes with that of the endogenous protein (PfPTP1; red); inset: enlargement of colocalization pattern. (B) Immunofluorescence assay on CS2/PfPTP1-HA cell line: the HA signal (green) colocalizes with the α-PfPTP1 signal (red); inset: enlargement of colocalization pattern. (C) Immunofluorescence assay on CS2/PfPTP1-HA cell line: the HA-signal (green) colocalizes partially with the MC resident protein SBP1 (red); inset: enlargement of colocalization pattern. (D) Immunofluorescence assay on CS2 parental cell line: the PfPTP2-signal (green) does not overlap with the PfPTP1-signal (red); inset: enlargement of area where MC (PfPTP1) and electron dense vesicles (PfPTP2) are in close proximity. (E) Immunofluorescence assay on CS2 parental cell line: the PfHsp70-x-signal (green) colocalizes partially with the PfPTP1-signal (red); inset: enlargement of partial colocalization on J-dots (PfHsp70-x). Scale bar: 1μm; same size of ROI in each image. (F) Western blot of solubility study on CS2/PfPTP1-HA cell line: top panel: parasite-infected RBC samples were treated with different detergents and supernatants (S) and pellets (P) detected on a western blot with α-HA antibodies; bottom panel; blot from top panel was stripped and probed with an antibody against the MC resident transmembrane protein SBP1. The 15 kDa is non-specific since it is found in the other blots treated with α-PfPTP1 antibodies. (G) Schematic and western blot of protease protection assay on CS2/PfPTP1-HA-infected RBCs; parasite-infected RBC samples were lysed and either treated with Tetanolysin O, saponin or Triton X-100 (1%). Each of the samples was then treated with either PBS or Proteinase K. The western blot was then either probed with an antibody recognizing the N-terminal (top panel) or C-terminal part (bottom panel) of PfPTP1. SBP 1 is a MC marker with its C-terminus exposed to the RBC cytosol, EXP 1 and EXP 2 are integral membrane proteins of the PVM and SERP is a soluble marker of the PV.

PfPTP1 is localized on Maurer's clefts in Pfalciparum–infected RBCs. (A) Immunofluorescence assay on CS2/PfPTP1-GFP cell line: the GFP signal (green) colocalizes with that of the endogenous protein (PfPTP1; red); inset: enlargement of colocalization pattern. (B) Immunofluorescence assay on CS2/PfPTP1-HA cell line: the HA signal (green) colocalizes with the α-PfPTP1 signal (red); inset: enlargement of colocalization pattern. (C) Immunofluorescence assay on CS2/PfPTP1-HA cell line: the HA-signal (green) colocalizes partially with the MC resident protein SBP1 (red); inset: enlargement of colocalization pattern. (D) Immunofluorescence assay on CS2 parental cell line: the PfPTP2-signal (green) does not overlap with the PfPTP1-signal (red); inset: enlargement of area where MC (PfPTP1) and electron dense vesicles (PfPTP2) are in close proximity. (E) Immunofluorescence assay on CS2 parental cell line: the PfHsp70-x-signal (green) colocalizes partially with the PfPTP1-signal (red); inset: enlargement of partial colocalization on J-dots (PfHsp70-x). Scale bar: 1μm; same size of ROI in each image. (F) Western blot of solubility study on CS2/PfPTP1-HA cell line: top panel: parasite-infected RBC samples were treated with different detergents and supernatants (S) and pellets (P) detected on a western blot with α-HA antibodies; bottom panel; blot from top panel was stripped and probed with an antibody against the MC resident transmembrane protein SBP1. The 15 kDa is non-specific since it is found in the other blots treated with α-PfPTP1 antibodies. (G) Schematic and western blot of protease protection assay on CS2/PfPTP1-HA-infected RBCs; parasite-infected RBC samples were lysed and either treated with Tetanolysin O, saponin or Triton X-100 (1%). Each of the samples was then treated with either PBS or Proteinase K. The western blot was then either probed with an antibody recognizing the N-terminal (top panel) or C-terminal part (bottom panel) of PfPTP1. SBP 1 is a MC marker with its C-terminus exposed to the RBC cytosol, EXP 1 and EXP 2 are integral membrane proteins of the PVM and SERP is a soluble marker of the PV.

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