Abnormal sterols in cholesterol-deficiency diseases cause secretory granule malformation and decreased membrane curvature

Marjorie C Gondré-Lewis,Christopher A Wassif,Horia I Petrache,Forbes D Porter,Adrian Parsegian,Y Peng Loh,Daniel Harries

doi:10.1242/jcs.02906

Abstract

Cholesterol is an abundant lipid in eukaryotic membranes, implicated in numerous structural and functional capacities. Here, we have investigated the mechanism by which cholesterol affects secretory granule biogenesis in vivo using Dhcr7(-/-) and Sc5d(-/-) mouse models of the human diseases, Smith-Lemli-Opitz syndrome (SLOS) and lathosterolosis. These homozygous-recessive multiple-malformation disorders are characterized by the functional absence of one of the last two enzymes in the cholesterol biosynthetic pathway, resulting in the accumulation of precursors. Cholesterol-deficient mice exhibit a significant decrease in the numbers of secretory granules in the pancreas, pituitary and adrenal glands. Moreover, there was an increase in morphologically aberrant granules in the exocrine pancreas of Dhcr7(-/-) acinar cells. Regulated secretory pathway function was also severely diminished in these cells, but could be restored with exogenous cholesterol. Sterol precursors incorporated in artificial membranes resulted in decreased bending rigidity and intrinsic curvature compared with cholesterol, thus providing a cholesterol-mediated mechanism for normal granule budding, and an explanation for granule malformation in SLOS and lathosterolosis.

Highlights

Cholesterol is a crucial player at the subcellular level in defining functional membrane microdomains for cellular activity (Schrader, 2004; van Meer, 1993)
Substitution of other sterols, i.e. 7-DHC and lathosterol, for cholesterol in Dhcr7–/– and Sc5d–/– mice, respectively, was not sufficient to rescue the regulated secretory pathway (RSP) and caused abnormal membrane characteristics attributed to the reduced rigidity of these sterols as determined by small-angle X-ray scattering methods. These findings clearly demonstrate that cholesterol is essential for conferring the rigidity necessary for membrane curvature during secretory granule biogenesis in vivo
Examination of the chemical structure for cholesterol, 7-DHC and lathosterol reveals differences between cholesterol and lathosterol only in the location of the double bond, and between cholesterol and 7-DHC, in the number of double bonds in the sterol ring, possibly altering contribution to membrane structure

Summary

Introduction

Cholesterol is a crucial player at the subcellular level in defining functional membrane microdomains for cellular activity (Schrader, 2004; van Meer, 1993). Studies in endocrine and other cells have suggested that cholesterol may be necessary for vesicle biogenesis at the trans-Golgi network (TGN), which contains cholesterol-sphingolipid-rich microdomains from which dense core granules (DCGs) bud (Keller and Simons, 1998; Loh et al, 2004; Tooze, 1998; Wang et al, 2000). Inborn errors of cholesterol synthesis lead to developmental abnormalities, including mental retardation, and the most common of these genetic diseases is Smith-Lemli-Opitz Syndrome (SLOS). SLOS is an autosomal-recessive, multiple malformation and/or mental retardation syndrome in which the function of the enzyme, 7-dehydrocholesterol reductase (DHCR7), necessary for the final step of cholesterol biosynthesis, is impaired (Porter, 2000; Tint et al, 1994). SLOS reportedly affects 1:10,000 to 1:40,000 caucasian Americans and is caused by mutations in the gene encoding

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