Ferredoxin reductase and p53 are necessary for lipid homeostasis and tumor suppression through the ABCA1-SREBP pathway

Abstract

p53 is known to modulate metabolism and FDXR is required for steroidogenesis. Given that FDXR is a target/regulator of p53, the FDXR-p53 axis may play a unique role in lipid metabolism. Here, we found that expression of ABCA1, a cholesterol-efflux pump, was suppressed by loss of FDXR and/or p53, leading to activation of master lipogenic regulators SREBP1/2. Accordingly, lipid droplets, cholesterol, and triglycerides were increased by loss of FDXR or p53, which were further increased by loss of both FDXR and p53. To explore the biological significance of the FDXR-p53 axis, we generated a cohort of mice deficient in Fdxr and/or Trp53. We found that Fdxr^+/-, Trp53^+/-, and Fdxr^+/-;Trp53^+/- mice had a short life span and were prone to spontaneous tumors and liver steatosis. Moreover, the levels of serum cholesterol and triglycerides were significantly increased in Fdxr^+/- and Trp53^+/- mice, which were further increased in Fdxr^+/-;Trp53^+/- mice. Interestingly, loss of Fdxr but not p53 led to accumulation of serum low-density lipoprotein. Together, our findings reveal that the FDXR-p53 axis plays a critical role in lipid homeostasis and tumor suppression.

Fig. 1. Lack of Fdxr, Trp53, or both leads to altered lipid metabolism though the ABCA1–SREBP pathway in MEFs.

A The levels of ABCA1, SREBP1/2, MVD, MVK, and actin were measured in WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− MEFs cultured in serum-free media for 4 h. B WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− MEFs were cultured in serum-free media for 8 h and then were stained with Nile Red (ex: 488 nm, em: 565 nm). DAPI (ex: 358 nm, em: 461 nm) was used to stain nuclei. C Quantitative measurement of intracellular cholesterol. WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− MEFs were cultured in a 96-well plate. After 4 h of fasting, the level of total cholesterol was measured with Cholesterol/Cholesterol Ester-Glo^TM assay kit according to the manufacturer’s instruction. Data represent the mean ± SD. D Quantitative measurement of intracellular triglyceride. WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− MEFs were cultured in a 96-well plate. After 4 h of fasting, the level of total triglycerides was measured with Triglyceride-Glo^TM assay kit according to the manufacturer’s instruction. Data represent the mean ± SD.

Fig. 2. Lack of FDXR, p53, or both leads to deregulation of lipid metabolism through the ABCA1–SREBP pathway in HCC cells.

A The levels of ABCA1, SREBP1/2, MVD, MVK, and actin proteins were measured in HepG2 cells transfected with scrambled siRNA (Scr) or siRNAs against FDXR and/or p53 for three days, followed by culturing in serum-free media for 4 h. B The levels of ABCA1, MVD, MVK, and actin mRNAs were measured in HepG2 cells treated as in A. C HepG2 cells transfected with scrambled siRNA (Scr) or siRNAs against FDXR and/or p53 were cultured in serum-free media for 4 h and then stained for lipid droplets with Nile Red (ex: 488 nm, em: 565 nm). DAPI (ex: 358 nm, em: 461 nm) was used to stain nuclei. D HepG2 cells transfected with scrambled siRNA (Scr) or siRNAs against FDXR and/or p53 were cultured in a 96-well plate. After 4 h of fasting, the level of total cholesterol was measured with Cholesterol/Cholesterol Ester-Glo^TM assay kit according to the manufacturer’s instruction. Data represent the mean ± SD. E HepG2 cells transfected with scrambled siRNA (Scr) or siRNAs against FDXR and/or p53 were cultured in a 96-well plate. After 4 h of fasting, the level of total triglycerides was measured with Triglyceride-Glo^TM assay kit according to the manufacturer’s instruction. Data represent the mean ± SD.

Fig. 3. Mice deficient in Fdxr, Trp53, or both have a short life span and are prone to spontaneous tumors.

A Kaplan–Meier survival curve for WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− mice. B Tumor spectra and penetrance in WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− mice. C Representative images of hematoxylin and eosin (H&E)-stained HCC in Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− mice.

Fig. 4. A deficiency in Fdxr, Trp53, or both leads to liver steatosis and chronic inflammation in mouse liver tissues.

A Representative images of hematoxylin and eosin (H&E)-stained liver in WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− mice. B The percentage of WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− mice with liver steatosis. C The percentage of WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−; Trp53^+/− mice with liver inflammation.

Fig. 5. Mice deficient in Fdxr, Trp53, or both are prone to lipid accumulation in the blood.

A The levels of ABCA1, SREBP1/2, MVD, MVK, and actin were measured in WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− mouse liver tissues. B–E The levels of blood cholesterol (B), triglycerides (C), LDL (D), and HDL (E) in 43-week-old WT, Fdxr^+/−, Trp53^+/−, and Fdxr^+/−;Trp53^+/− mice (n = 5). Values are mean ± SEM and analyzed by 2-tailed t-test.