Genetic basis of sitosterolemia

Abstract

The molecular mechanisms regulating the amount of dietary cholesterol retained by the body, as well as the body's ability to exclude other dietary sterols selectively, are poorly understood. An average Western diet will contain approximately 250-500 mg of dietary cholesterol and approximately 200-400 mg of non-cholesterol sterols, of which plant sterols are the major constituents. Approximately 50-60% of dietary cholesterol is absorbed and retained by the normal human body, but less than 1% of the non-cholesterol sterols are retained. There thus exists a subtle mechanism that allows the body to distinguish between cholesterol and non-cholesterol sterols. In sitosterolemia, a rare autosomal recessive disorder, affected individuals hyperabsorb and retain not only cholesterol but also all other sterols, including plant and shellfish sterols from the intestine. Consequently, patients with this disease have very high levels of plant sterols in the plasma, and develop tendon and tuberous xanthomas, accelerated atherosclerosis, and premature coronary artery disease. The STSL locus has been mapped to human chromosome 2p21. Mutations in two tandem ABC genes, ABCG5 and ABCG8, encoding sterolin-1 and -2, respectively, are now known to be mutant in sitosterolemia. The identification of these genes should now lead to a better understanding of the molecular mechanism(s) governing the highly selective absorption and retention of cholesterol by the body. Indeed, it is the very existence of this disease that has given credence to the hypothesis that there is a molecular pathway that regulates dietary cholesterol absorption and sterol excretion by the body.

Figure 1. Plasma sitosterol and cholesterol levels in affected individuals, their parents, clinically unaffected siblings, and normal controls

The plasma sitosterol levels were determined using capillary gas liquid or high performance liquid chromatography. In general, most unaffected individuals had plasma sitosterol levels of less than 1 mg/dl. Of the three parents and three siblings who had values higher than 1 mg/dl, none was higher than 2 mg/dl. All of the affected individuals had plasma sitosterol values greater than 8 mg/dl, and many of these values were obtained while the patients were on treatment. Note that the plasma cholesterol levels are not helpful. Three children all have mutations.

Figure 2. Sitosterolemia pedigrees

The pedigrees analyzed in this study are shown above. All parents are shown as obligate carriers. The one known consanguineous marriage, pedigree 3400, is as indicated. Affected individuals are indicated by the filled symbols, the obligate carriers (parents) by the half-filled symbols. All unaffected siblings are shown by unfilled symbols.

Figure 3. Homozygosity detected in all of the pedigrees and haplotype sharing in some of the pedigrees

Genotypes for markers are shown for probands across the critical STSL area. The shaded regions indicate the minimal region of homozygosity present in all probands, bounded by markers D2S2294 and Afm210xe9. Note that only pedigree 3400 was known to be consanguineous. In addition to homozygosity, haplotype sharing, boxed areas, was detected in two Japanese probands, the Amish and Mennonite families and Norwegian and Finnish probands.

Figure 4. Genealogical analyses of the Amish and Mennonite families in this study

A genealogical analysis of the parents from pedigrees 2700 and 2200 was performed as described in the methods section. Five possible founders, who link all four parents, were identified. An approximate time index is as shown in the left margin for temporal orientation. As can be seen, individual 74 is the least related to all of the remaining three obligate carriers.

Figure 5. Gene structure for ABCG5 and ABCG8

The gene structures for ABCG5 and ABCG8 are as depicted. Each gene consists of 13 exons and the genes are arranged in a head-to-head configuration, with no more than 150 bases separating the start-transcription sites, with only 372 bases separating the two respective ‘ATG’. No TATA box is present in the 140 bases separating the two genes.

Figure 6. Possible models of action of sterolins

We propose two models that may explain how sterolins function to prevent non-cholesterol sterols (represented by sitosterol) from being retained by the body. In both models, both sterolin-1 and sterolin-2 function as a heterodimer. Model A predicts that the heterodimer is responsible for the exclusion of sterols by actively pumping them out of the cells (the enterocyte or the hepatocyte) at the apical border. However, this pump has a much higher affinity for non-cholesterol sterols compared with cholesterol. Note that this mode of action allows for the sterolins to pump cholesterol into bile, in the absence of non-cholesterol sterols in the liver. However, under model A, sterolins do not regulate the entry of sterols into the enterocyte or the hepatocyte. Model B predicts that the sterolin heterodimer acts in a bi-functional direction, again at the apical surfaces of the enterocyte or the hepatocyte. In this case, the heterodimer shows a higher affinity for the entry of cholesterol versus non-cholesterol sterols into the cells, and a higher affinity for pumping non-cholesterol sterols out of the cells.