Flux analysis of cholesterol biosynthesis in vivo reveals multiple tissue and cell-type specific pathways

Abstract

Two parallel pathways produce cholesterol: the Bloch and Kandutsch-Russell pathways. Here we used stable isotope labeling and isotopomer analysis to trace sterol flux through the two pathways in mice. Surprisingly, no tissue used the canonical K–R pathway. Rather, a hybrid pathway was identified that we call the modified K–R (MK–R) pathway. Proportional flux through the Bloch pathway varied from 8% in preputial gland to 97% in testes, and the tissue-specificity observed in vivo was retained in cultured cells. The distribution of sterol isotopomers in plasma mirrored that of liver. Sterol depletion in cultured cells increased flux through the Bloch pathway, whereas overexpression of 24-dehydrocholesterol reductase (DHCR24) enhanced usage of the MK–R pathway. Thus, relative use of the Bloch and MK–R pathways is highly variable, tissue-specific, flux dependent, and epigenetically fixed. Maintenance of two interdigitated pathways permits production of diverse bioactive sterols that can be regulated independently of cholesterol.DOI: http://dx.doi.org/10.7554/eLife.07999.001

Cholesterol-mediated Degradation of 7-Dehydrocholesterol Reductase Switches the Balance from Cholesterol to Vitamin D Synthesis

“…8A). The skin predominantly utilizes a modified Kandutsch-Russell pathway (2), indicating that the pathway can provide a constant source of 7DHC for the production of vitamin D (58). In addition, the highly labile nature of DHCR7, destabilized further by cholesterol, also serves to switch flux toward vitamin D synthesis (Fig.…”

“…7-Dehydrocholesterol can be converted to cholesterol by acting as a substrate for DHCR7 or to vitamin D through exposure to ultraviolet B (UVB) light. A simplified version of the modified Kandutsch-Russell pathway is shown as described in detail previously (2). B, CHO-DHCR24-V5 cells were transfected with empty vector (EV) or DHCR7-myc plasmid and treated with or without 10 g/ml cycloheximide (CHX) for 0 -8 h. Protein levels were analyzed using Western blotting with myc, V5, and GAPDH antibodies.…”

“…The alternative terminal enzyme in the Bloch pathway of cholesterol synthesis is 24-dehydrocholesterol reductase (DHCR24) (3). Despite a certain level of redundancy, both the Kandutsch-Russell and Bloch pathways are utilized in different cellular contexts, and are both essential for cell survival (2). DHCR7 acts by reducing the C(7-8) double bond of 7-dehydrocholesterol (7DHC) to form cholesterol.…”

“…Its synthesis requires a complex series of reactions that involves Ͼ20 steps, which must be tightly controlled to balance cholesterol levels. Here, we focus on the post-translational regulation of 7-dehydrocholesterol reductase (DHCR7), 2 the terminal enzyme in the Kandutsch-Russell pathway of cholesterol synthesis (1, 2) (Fig. 1A).…”

Transcription Factor Hepatocyte Nuclear Factor–1β Regulates Renal Cholesterol Metabolism

“…50,51 Cholesterol synthesis rate was estimated by fitting the relationship between fraction of newly synthesized and time to a firstorder kinetic models. 23 For sterols and oxysterols measurements, cells were seeded in six-well cell culture plates, induced for 48 hours, and subsequently processed for HPLC/MS analysis as previously described. 22 …”