Extended Data Fig. 8: Dense foci of mTOR in GCs.
From: Subcellular transcriptomes and proteomes of developing axon projections in the cerebral cortex
a, Data relate to Fig. 5. Biochemical analysis of GC enrichment of mTOR pathway proteins and controls, shown in triplicate western blots of homogenate (input) and GC fraction pairs, derived from six independent preps. GAP43 is positive control for enrichment; GM130 is a negative control. Quantification as in Fig. 5. b, Close-ups of GCs from callosal projection neurons immunelabelled for endogenous mTOR pathway proteins (red in overlays, heat-mapped in underlying panels). Five example GCs are shown per sample to capture the representative range. Neurons were labelled via in utero electroporation at E15 with membrane-GFP (green in overlays, outlined in underlying panels), cultured at P0, fixed and stained at DIV2–DIV3. mTOR, LARP1, TSC1 and raptor (mTORC1 marker) appear in dense local foci within GCs. RICTOR (mTORC2 marker) and LAMP1 (lysosome marker) appear in fine granules distinct from GC foci. Bar (bottom right) indicates heat-map colour range, as well as 10-μm scale. 83 GCs imaged for mTOR, 47 for LARP1, 42 for LAMP1, 49 for TSC1, 26 for raptor, and 30 for RICTOR, from a minimum of n = 3 biological replicates from independent in utero electroporations.
