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. 2021 Jan 12;34(2):108623. doi: 10.1016/j.celrep.2020.108623

Figure 4.

Figure 4

The VGLUT1-Specific NB9 Nanobody Abolishes Pi Transport by COS7-VGLUT1 Cells

(A) TetOn:mCherry-2A-NB9 expression was induced in COS7-VGLUT1-TetOn cells by overnight treatment with 500 ng/mL [doxycycline]. COS7-VGLUT1 cells transfected with a TetOn:luciferase construct served as MOCK control. EGFP and mCherry fluorescence signals were directly monitored by confocal microscopy as markers of VGLUT1 or NB9 expression, respectively. Scale bar, 25 μm.

(B) Pi content in COS7-VGLUT1-TetOn:mCherry-NB9 (left) or COS7-VGLUT1-TetOn:Luc (right) cells, preincubated in the absence (plain bars) or presence (striped bars) of doxycycline, shows that doxycycline-induced expression of NB9 reduced Pi uptake to levels comparable to those observed in the presence of RB (left). No effect of doxycycline was observed in the COS7-VGLUT1-TetOn:MOCK cell line (right).

All values are normalized to control (no doxycycline; no Pi) and represent mean values ± SEM from 4–13 experiments. Significance toward: (1) no Pi added; (2) Pi; and (8) no Dox.