Skip to main content
. Author manuscript; available in PMC: 2018 Jul 15.
Published in final edited form as: Nat Genet. 2018 Jan 15;50(2):219–228. doi: 10.1038/s41588-017-0026-3

Figure 4. Nuclear PDC regulates the expression of lipid biosynthesis genes independently of mitochondrial PDC.

Figure 4

(a) Western blot analysis of indicated proteins in nuclear and cytoplasmic fractions of shPDHA1 22Rv1 cells infected with NES-PDHA1 and NLS-PDHA1 alone or in combination. Uncropped images are in Supplementary Figure 13. (n=3, independent cell cultures). (b) Western blot analysis of indicated proteins in shPDHA1 22Rv1 cells infected NES-PDHA1 and NLS-PDHA1 alone or in combination (see full panel in Supplementary Fig. 5b). Uncropped images are in Supplementary Figure 13. (n=3, independent cell cultures). (c-e) Quantitative real-time PCR analysis of mRNA expression for ACLY (c) and SQLE (d) and determination of citrate levels (e) in shRNA control and shPDHA1 22Rv1 and PC3 infected with NES-PDHA1, NLS-PDHA1 alone or in combination (n=3, independent cell cultures). (f) Representative confocal images and quantification of lipid droplets (average lipid droplets per cell) in shPDHA1 22Rv1 infected with NES-PDHA1, NLS-PDHA1 alone or in combination (n=3, independent cell cultures, Scale bar represents 10 µm, 5 fields acquired for each group). (g) Upper panel, representative images of crystal violet staining of shPDHA1 22Rv1 infected with infected with NES-PDHA1, NLS-PDHA1 alone or in combination (n=3, independent cell cultures). Lower panel, relative cell number quantification by crystal violet staining in shRNA control and shPDHA1 22Rv1 and PC3 infected with NES-PDHA1, NLS-PDHA1 alone or in combination (n=3, independent cell cultures). (h) Upper panel, representative micrographs in histopathological analysis of Ki-67 of these xenograft tumours. Lower panel, evaluation of tumour formation in xenotransplantation experiments of shPDHA1 22Rv1 infected with NES-PDHA1, NLS-PDHA1 alone or in combination (n=6 animals; 12 injections, Scale bar represents 50 µm, also see Supplementary Fig. 7d for the full panel). Error bars indicate s.e.m. *P < 0.05; **P < 0.01; ***P < 0.001. n.s, not significant.