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. Author manuscript; available in PMC: 2019 May 6.
Published in final edited form as: Sci Signal. 2017 Oct 10;10(500):eaal4055. doi: 10.1126/scisignal.aal4055

Fig. 6. Development of remotely controlled TRPVFeRIC channels.

Fig. 6.

(A) TRPV channels were tagged with domain 5 of kininogenl (D5) and were cloned into the PLVX vector with an internal ribosomal entry site (IRES) for mCherry (fig. S12 and S13). (B) FeRIC channels were designed to recruit endogenous cellular ferritin to the modified TRPV channel at the cell membrane. (C) Cytoplasmic distribution of ferritin heavy chain fused with mCherry (FTH1mCherry) in HEK293T cells expressing TRPV1WT and membrane redistribution of FTH1mCherry (white arrowheads) in TRPV 1FeRIC-expressing cells. Images were representative of 2 independent experiments. (D) Representative immunoprecipitation western blot of HEK293T cells expressing FLAG tagged TRPV 1WT, TRPV1FeRIC, TRPV4WT, or TRPV4FeRIC. The blot was probed for FLAG and FTH1. 4 independent experiments were conducted using TRPV lFeRIC and 3 independent experiments were conducted using TRPV4FeRIC. (E) GCaMP6 fluorescence in TRPV 1WT-expressing (blue) or TRPV1FeRIC-expressing (red) HEK293T cells following RF (gray box) then 1μM capsaicin (bar). Bar graphs are DF/F0 averages of 4 experiments with 50–100 cells/group analyzed. (F) GCaMP6 fluorescence in TRPV4WT-expressing (blue) or TRPV4FeRIC-expressing (red) HEK293T cells following RF (gray box) then 1μM GSK101 (bar). Bar graphs are ΔF/F0 averages of 5 experiments with 106–123 cells/group analyzed. Significance was determined using unpaired t-test. *P<0.05.