Fig. 2: Mass spectrometric confirmation of PA-mTOR binding in vivo. | Communications Biology

Fig. 2: Mass spectrometric confirmation of PA-mTOR binding in vivo.

From: Phosphatidic acid directly activates mTOR and then regulates SREBP to promote ganoderic acid biosynthesis under heat stress in Ganoderma lingzhi

Fig. 2

a Isolation of mTOR by immunoprecipitation. mTOR was immunoprecipitated using an anti-mTOR antibody from cell lysates of mycelia, and was probed by immunoblotting using the same antibody. + and - indicate with and without antibody (lines 1 and 2), with and without HS for 30 min (lines 2 and 3), respectively, in the immunoprecipitation. Line 4 indicated mycelia were pre-incubation with 100 nM rapamycin for 30 min before being exposed to HS in the immunoprecipitation. Middle and bottom blot shows protein input (mTOR and β-Tubulin) used for the immunoprecipitation. b Coprecipitation of total PA with mTOR. Total PA extracted from mTOR immunoprecipitated in a was quantified by ESI-MS/MS. c Coprecipitation of PA species with mTOR. PA species from total PA extracted in (b). The values in (b) and (c) are the means ± SD (n = 3 independent experiments). Different letters in (b) and (c) indicate significant differences between the lines in the total PA and same PA species, respectively (P < 0.05, according to Duncan’s multiple range test).

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