FIG 7.

VAPB association with the inclusion membrane is dependent on IncD and the FFAT motif of CERT. (A, B, and D) Confocal fluorescence micrographs of inclusions of a C. trachomatis strain, expressing mCherry (red) and IncD3×FLAG under the control of an aTc-inducible promoter, in HeLa cells coexpressing a CFP-VAPB (blue) construct and either a wild-type YFP-CERT construct (YFP-CERT; yellow) (A and B) or a version displaying a mutation in the FFAT motif (YFP-CERT FFATmut; yellow) (D). At 1 h postinfection, the infected cells were incubated in the absence (No aTc) or presence (+ aTc) of 20 ng/ml aTc. The cells were fixed at 23 h post-aTc induction and imaged using a confocal microscope. An extended-focus view combining all the confocal planes spanning an entire inclusion (A) or views of a single plane crossing the middle of the inclusion (B and D) are shown. Merged images are shown on the left. Bar, 10 μm. (C and E) Quantifications of the proportions of the inclusion membrane covered by the indicated markers. The quantification method is described in Materials and Methods.