Organelle dysfunction modulates cholesterol biosynthesis pathway

Mitochondria are known as the powerhouse of the cells. However, in recent years, it has become increasingly evident that mitochondria are involved in several signaling pathways. Lysosomes are membrane-bound organelles that receive and degrade macromolecules from the secretory, endocytic and autophagy pathways. Our lab has recently reported that mitochondrial respiratory chain deficiency inhibits lysosomal hydrolysis and that lysosomal lipid storage diseases are characterized by the repression of mitochondrial biogenesis; thereby showing interdependence between these organelles. However, little is known about the connection between the dysfunction of these organelles and lipid metabolism. Here, we aimed to elucidate if the dysfunction in mitochondria or lysosomes yields any consequences on lipid metabolism, more specifically cholesterol biosynthesis, and to describe the underlying mechanism. We have employed cell lines with defects in each of these organelles and measured the effects on gene expression by RNA sequencing (RNA-Seq), followed by comprehensive multi-dimensional transcriptomics analysis, as well as by quantitative real-time PCR. Interestingly, changes in the expression of genes of the cholesterol synthesis pathway were among the significant consequences of both the mitochondrial and the lysosomal perturbations, but with opposite trends. While mitochondrial dysfunction results in the downregulation of the cholesterol biosynthesis, lysosomal dysfunction leads to its activation. Analysis of the cholesterol content showed a significant decrease of unesterified (free) cholesterol level in cells with mitochondrial dysfunction. On the other hand, free cholesterol level was increased in the cells with lysosomal dysfunction. Mitochondrial cholesterol content was increased only in the cells with mitochondrial dysfunction. To further investigate the origin of the opposite trends of cholesterol biosynthesis resulting from organelle dysfunction, we measured the activity of SREBP1, an established regulator of the cholesterol biosynthesis. We found a decrease in the SREBP1 activity in cells with mitochondrial dysfunction and its increase in the case of lysosomal dysfunction. The crucial role of AMPK in the regulation of the cholesterol biosynthesis pathway is evidenced by the changes in AMPK activity in the cells with organelle dysfunctions: while in the case of mitochondrial dysfunction AMPK showed increased activity, the trend was opposite in the cells with lysosomal dysfunction. Importantly, based on our results, the effect of organellar dysfunction on cholesterol biosynthesis depends on AMPK activity. This work contributes to the understanding of the underlying mechanisms of pathologies associated with organelle dysfunction, such as lysosomal storage diseases or mitochondrial diseases, and the impact they may have on lipid homeostasis, such as the cholesterol biosynthesis.2020-09-2