Extended Data Fig. 4: Vps34 inactivation does not impact Bmal1 protein levels.
From: Class 3 PI3K coactivates the circadian clock to promote rhythmic de novo purine synthesis
a, Densitometric analysis of Vps15 and Vps34 levels normalized to Actin in total protein extracts of synchronized control (GFP) and Vps34-depleted (CRE) MEFs collected 16 h after synchronization with dexamethasone. Data are the mean ± s.e.m. (n = 3). #P < 0.05 versus GFP; two-tailed unpaired Student’s t-test. b, Immunofluorescence microscopy analyses of p62 in Vps34-depleted (CRE) and GFP-transduced MEFs 5 d post infection. Experiment repeated three times, representative field shown. Scale bar, 20 µm. c, Relative transcript levels of the indicated genes in dexamethasone-synchronized control (GFP) and Vps34-depleted (CRE) MEFs. Data collected in three independent experiments presented as the fold change ± s.e.m. over GFP-treated cells (n = 5 GFPCT16 and n = 6 for all other groups). #P < 0.05 versus GFP; two-tailed unpaired Student’s t-test. Rhythmicity was determined using JTK_CYCLE (Supplementary Table 1). d, Immunoblot analysis, using the indicated antibodies, of cytosolic and soluble nuclear protein extracts in dexamethasone-synchronized control and Vps34-depleted MEFs. Densitometric analyses of Bmal1 protein normalized to tubulin (cytosolic fraction) and β-catenin (soluble nuclear fraction) presented as the fold change over GFP-treated cells. Data collected in three independent experiments are the mean ± s.e.m. (n = 8). Source data and unprocessed blots are provided.
