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. 2019 Apr 4;15(4):e1007512. doi: 10.1371/journal.ppat.1007512

Fig 6. Over-expression of a dominant-negative form of TgDrpC leads to interconnected mitochondria.

Fig 6

A) Scheme of the plasmid DD-GFP-DrpCDN. (B) Western blot analysis with the indicated antibodies shows that Shield-1 induction efficiently regulates the expression of DD-GFP-DrpCDN as soon as 4 hours post induction. (C) Plaque assay shows that induction of DD-GFP-DrpCDN causes parasite death. Parasites were grown for 7 days on HFF cells in presence and absence of 1μM Shield-1. The experiment was performed in triplicate; representative images are shown. (D) The effects of DD-GFP-DrpCDN expression were assessed 24 hours after Shield-1 induction; immunofluorescence analysis of the specific markers Mic4, Rop13, HSP60 and Gap45 shows no effect on secretory organelles, apicoplast or IMC morphology. (E and F) Eight hours after stabilisation, DD-GFP-DrpCDN shows an accumulation at the basal part of the mitochondria, which increasingly appear interconnected (23% of the total); 24 hours after induction, more than half of the parasites show abnormally interconnected mitochondria, and DD-GFP-DrpCDN accumulates at the interconnections (scale bar: 5 μm). At least 100 parasitophorous vacuoles were counted in triplicate. Error bars represent SD from the three independent experiments. Asterisks indicate significant difference (P<0.001 multiple t-test).