Differential gene expression of Caenorhabditis elegans grown on unmethylated sterols or 4alpha-methylsterols.

Transcriptional profiles of Caenorhabditis elegans grown on unmethylated sterols (desMSs) or on 4alpha-methylsterols (4MSs) were compared using microarrays. Thirty-four genes were upregulated and 2 were downregulated>2-fold by growth on 4MSs, including 13 cuticle collagen (col) genes, 1 cuticulin gene (cut-1), 2 groundhog-like (grl) genes, and 1 groundhog gene (grd-4); col-36 and grl-20 were increased 12- and 19-fold, respectively. Fifteen of these 17 genes have been assigned to metabolic mountain 17, suggesting coordinate 4MS-mediated regulation of expression. Quantitative RT-PCR was performed on 27-51 h old animals grown on cholesterol (a desMS) or lophenol (a 4MS). col-36 and grl-20 showed similar cyclic peaks of expression in cholesterol and similar alterations in lophenol, suggesting coregulation. Of six additional grl genes, only grl-3 was upregulated on lophenol; the rest were downregulated. Cyclicity of expression was lost or altered in all six. Nuclear receptor genes nhr-23, nhr-25, nhr-41, and daf-12 all showed cyclic expression in cholesterol and significant downregulation in lophenol by RT-PCR. Expression of the insulin-like receptor daf-2 was lower in lophenol, whereas that of its major downstream target daf-16 was higher. Thus, major changes in gene expression accompany growth on 4MSs, but with surprisingly little effect on normal growth and development.

Long-term effects of sterol depletion in C. elegans: sterol content of synchronized wild-type and mutant populations.

Three major long-term effects of sterol deprivation in Caenorhabditis elegans are described. 1) The life expectancy of sterol-deprived wild-type animals is decreased by more than 40%. Similar decreases are found in animals carrying mutations in the daf-9, daf-12, daf-16, and clk-1 genes, suggesting that previously described aging pathways involving these genes are not involved in the life-extending effects of sterols. 2) There is a premature loss of motility, measured by response to mild touch. 3) There is a rapid postreproductive onset of sarcopenia (muscle wasting) as measured by total body fluorescence in a myo3::GFP-expressing strain. We also report that five sterols (the desmethylsterols cholesterol, 7-dehydrocholesterol, and lathosterol and the 4alpha-methyl sterols lophenol and 4alpha-methyl-cholesta-Delta8(14)-en-3beta-ol) are found in significant amounts at all stages of development and aging in cholesterol-fed animals. Supplying any one of these as the sole sterol confers similar protection from the long-term effects of sterol deprivation. These findings suggest that sterols are required continuously throughout the animal's life.

Sterol effects and sites of sterol accumulation in Caenorhabditis elegans: developmental requirement for 4alpha-methyl sterols.

Caenorhabditis elegans requires sterol, usually supplied as cholesterol, but this is enzymatically modified, and different sterols can substitute. Sterol deprivation decreased brood size and adult growth in the first generation, and completely, reversibly, arrested growth as larvae in the second. After one generation of sterol deprivation, 10 ng/ml cholesterol allowed delayed laying of a few eggs, but full growth required 300 ng/ml. C. elegans synthesizes two unusual 4alpha-methyl sterols (4MSs), but each 4MS supported only limited growth as the sole sterol. However, addition of only 10 ng of cholesterol to 1,000 ng of 4MS restored full growth and egg-laying, suggesting that both a 4MS and an unmethylated sterol are required for development. Filipin stained sterols in only a few specific cells: the excretory gland cell, two amphid socket cells, two phasmid socket cells and, in males, spicule socket cells. Sterols were also present in the pharynx and in the intestine of feeding animals in a proximal-to-distal gradient. This non-random sterol distribution, the low concentration requirements, and the effects of 4MSs argues that sterols are unlikely to be used for bulk structural modification of cell membranes, but may be required as hormone precursors and/or developmental effectors.