Cell biology and disorders of cholesterol homeostasis
Heiko Runz and Rainer Pepperkok
Our research interest aims to identify and functionally characterize novel factors regulating cellular cholesterol metabolism. Our long-term goals are a better molecular understanding of elevated blood cholesterol levels (hypercholesterolemia) and Niemann-Pick Type C (NPC) disease with the perspectives to develop novel diagnostic and therapeutic tools. The experimental approaches we use are (i) identification of candidate genes by disease related gene expression analyses, (ii) cell biological characterization of candidate genes with respect to cholesterol metabolism and (iii) analysis of possible gene defects in patient samples.
Cholesterol levels in animal cells need to be tightly balanced. Within recent years, a network of proteins has been identified that keep cellular cholesterol levels to narrow limits. The concerted interplay of these proteins confers that reduced levels of membrane-bound cholesterol may activate the expression of genes necessary to increase cholesterol levels in cells, e.g. by uptake of low-density lipoprotein (LDL)-bound cholesterol or cholesterol biosynthesis. It is, however, assumed that only a limited number of proteins involved in cellular cholesterol regulation have been identified to date.
- To better understand the basic molecular and cell biological mechanisms of cellular cholesterol regulation.
- To understand the role of the candidate genes in patient samples and cell models of human disease.
- To combine the cell biological and genetic studies for improving diagnostic tools and maybe opening novel therapeutic strategies.
Research Focus 1
Identification of genes predisposing hypercholesterolemia
One project that we are focussing on is the identification of genes pre-disposing for elevated blood cholesterol levels (hypercholesterolemia), a major risk factor for atherosclerosis and coronary heart disease. Candidate genes with a putative role in hypercholesterolemia are first identified by microarray-based gene expression analyses in cells with different cholesterol load. A putative cell biological function of these candidates is then further characterized by using systematic RNAi-based gene knockdown experiments and automated microscopy. To this end we use the EMBL microscopy based RNAi-screening platform that allows cell-based loss-of-function studies at a high throughput. We have developed fluorescence-based assays by which cellular uptake of fluorescence-labelled low-density lipoprotein (LDL) and its dynamics can be analysed under high-throughput conditions.
Research Focus 2
Identification of novel regulators of Niemann-Pick Type C disease
In a second project we aim at the identification of novel regulators of Niemann-Pick Type C (NPC) disease. NPC disease is a rare autosomal-recessive lysosomal storage disease caused by mutations in the genes NPC1 or NPC2 that is typically accompanied by progressive impairment of nervous system and liver function. On a cellular level, the disorder presents with an inhibited transport of cholesterol and glycosphingolipids out of endosomal and lysosomal compartments, which we can visualize in cultured cells. Presently, we analyze if the knock-down of candidate genes leads to a cellular cholesterol distribution that resembles, aggravates or ameliorates the cellular NPC phenotype.
Characterization of drug-induced transcriptional modules: towards drug repositioning and functional understanding
Iskar M, Zeller G, Blattmann P, Campillos M, Kuhn M, Kaminska KH, Runz H, Gavin AC, Pepperkok R, van Noort V, Bork P.
Mol Syst Biol. 2013 Apr 30;9:662. doi: 10.1038/msb.2013.20
RNAi-based functional profiling of loci from blood lipid genome-wide association studies identifies genes with cholesterol-regulatory functions.
Blattmann P, Schuberth C, Pepperkok R, Runz H.
PLOS Genetics 2013, Feb;9(2):e1003338
Identification of cholesterol-regulating genes by targeted RNAi-screening
Bartz F, Kern L, Erz D, Zhu M, Gilbert D, Meinhof T, Wirkner U, Erfle H, Muckenthaler M, Pepperkok R, Runz H
Cell Metabolism 8 July 2009, 10, 63-75
DetecTiff: A novel image analysis routine for high content screening microscopy
Gilbert D, Meinhof T, Pepperkok R, Runz H.
J Biomol Screen. 2009 Sep;14(8):944-55. Epub 2009 Jul 29
Sterols regulate ER-export dynamics of secretory cargo protein ts-O45-G
Runz H, Miura K, Weiss M and Pepperkok R.
EMBO J. 2006, 25(13), 2953-65.
Inhibition of intracellular cholesterol transport alters presenilin localization and APP processing in neuronal cells
Runz H, Rietdorf J, Tomic I, de Bernard M, Beyreuther K, Pepperkok R and Hartmann T.
J. Neuroscience 2002, 22(5), 1679-89