PXA domain of Mdm1 recruits FA-CoA ligase Faa1 to LD budding sites. (A) Left: Light microscopy of GFP-tagged Mdm1^ER-PM in yeast stably expressing mCherry-tagged Faa1 in the presence of 0.2% oleate. Scale bar, 2 µm. LDs (gray) were visualized by using MDH. Arrows indicate co-enrichment of Mdm1^ER-PM, Faa1, and LDs. Scale bar, 2 µm. (B) Line tracing of light microscopy images in (A). (C) Left: Light microscopy of different fragments of Mdm1^ER-PM in yeast stably expressing mCherry-tagged Faa1 in the presence of 0.2% oleate. LDs (gray) were visualized by using MDH. Scale bar, 2 µm. Arrows indicate co-enrichment of Mdm1^ER-PM, Faa1, and LDs. Right: Illustrated representation of the topology of different Mdm1^ER-PM constructs being expressed. (D) Quantification of Mdm1^ER-PM foci associated with Faa1 from images in A and C. Data represent the number of LD-associated foci over the total Mdm1^ER-PM foci. Mean ± SD; n > 50 cells; *, P < 0.01; **, P < 0.005; Student’s t test. (E) Diagram depicting the pathway of neutral lipid synthesis. (F) Quantification of BODIPY-C16 incorporation into neutral lipid (NL) BODIPY-DAG in mdm1Δ and faa1Δ relative to WT yeast (1-h incubation). n = 3; **, P < 0.005; Student’s t test. Raw data are shown in Fig. S3G. (G) Quantification of BODIPY-C16 incorporation into neutral lipid (NL) BODIPY-TAG in mdm1Δ and faa1Δ relative to WT yeast (1-h incubation). n = 3; **, P < 0.005; Student’s t test. Raw data are shown in Fig. S3 G (high exposure). (H) Quantification of NBD-C16-CoA incorporation into NBD-TAG in mdm1Δ and faa1Δ relative to WT yeast (1-h incubation). n = 3; ns, nonsignificant; Student’s t test. Raw data are shown in Fig. S3 H.