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. Author manuscript; available in PMC: 2015 Aug 1.
Published in final edited form as: Pathog Dis. 2014 May 16;71(3):336–351. doi: 10.1111/2049-632X.12179

Figure 4. Evidence for CPAF mediated processing of vimentin in intact cells.

Figure 4

A) CPAF is not active in 1% SDS buffer. HeLa cell lysates prepared under denaturing conditions in 1% SDS buffer were incubated with the 0.5, 2.5, and 5 μg recombinant CPAF (rCPAF) for 20 minutes at 37°C. As a positive control, rCPAF was also incubated with HeLa lysates prepared under non-denaturing conditions in TNEX buffer. Where indicated, 100 μM CPAF inhibitor peptide (CIP) was included as a control. CPAF activity was assessed by monitoring the generation of vimentin cleavage products by immunoblot analysis. B) Proteolytic processing of vimentin at later stages of infection is dependent on CPAF. The bottom panel represents an increased exposure (OE) of the middle panel. FL: full-length vimentin; C: cleavage product C) LDH release by cells infected with wild type and CPAF-deficient C. trachomatis strains. The supernatants of infected cells were collected at the indicated time points and the amount of LDH release was measured and compared to total LDH levels. Error bars represent standard deviation, n=3.