Extended Data Fig. 3 |. mTOR is not required for the activation of TMEM175 by AKT.

a–c, Lysosomal I_K currents were recorded from TMEM175-tranfected HEK293T cells with mTOR (a component of both mTORC1 and mTORC2 complexes) (a), Raptor (an mTORC1 component) (b) or Rictor (an mTORC2 component) (c) knocked down with short hairpin (sh)RNA lentivirus¹⁰. d, Lysosomal I_K recorded from TMEM175-tranfected HEK293T cells treated with rapamycin (2 μM, 1 h). Figure 1f provides comparisons with those with no knockdown or rapamycin treatment. *P ≤ 0.05. P values (unpaired two-tailed t-tests; K vs K+SC79 groups) are: P < 0.0001 (mTor shRNA), P = 0.0013 (Raptor shRNA), P < 0.0001 (Rictor shRNA). e–h, Control for mTOR knockdown. Lysosomal Na⁺ currents were recorded from HEK293T cells transfected with TPC2 as positive controls for mTOR knockdown. TPC2 is a lysosomal Na⁺ channel that is inhibited by ATP. The ATP sensitivity requires mTORC1 but not mTORC2; knocking down mTOR or Raptor—but not Rictor—reduces the ATP sensitivity¹⁰. In f, arrows are used to indicate traces that overlap and are not easily distinguished. Black, recorded before 1 mM ATP was added to the bath; red, recorded after 1 mM ATP was added to the bath. Data are mean ± s.e.m. Numbers of recordings are in parentheses.