1. Lipid Homeostasis and Transport CH353 February 12, 2008

2. Summary Major transported forms of lipids –Cholesterol esters –Triacylglycerols (triglycerides) Lipid transport particles (solubilize lipids & target cells) –Chylomicrons (dietary triglyceride transport) –VLDL (hepatic lipid transport) –LDL (cholesterol transport and regulation) –HDL (reverse cholesterol transport) Cholesterol Homeostasis –Intracellular transport –Regulation of enzyme levels –Regulation of enzyme activity

3. Cholesteryl Ester Synthesis in liver acyl-CoA-cholesterol acyl transferase (ACAT) outside cells (on HDLs) lecithin-cholesterol acyl transferase (LCAT) +

4. Triacylglycerol Biosynthesis Synthesis of triacylglycerol from phosphatidic acid Common precursor with membrane glycerolipid biosynthesis Mainly in liver and intestine (for transport) and adipose tissue (for storage)

5. Lipoproteins a)Structure of Low-Density Lipoprotein (LDL) b)Electron microscope images of lipoproteins

6. Chylomicrons Density: < 1.006 g/ml Mass: 50–1000 x 10 3 kDa Diameter: 75–1200 nm Composition: –2% protein (apoB-48, apoA, apoC, apoE) –9% phospholipids, 85% triglycerides, 4% cholesterol Origin: intestinal enterocytes (apoproteins and lipid) Function: Transports dietary triglycerides from intestine to tissues; returns remnants to liver

7. Very Low Density Lipoprotein (VLDL) Density: < 0.95–1.006 g/ml Mass: 10–80 x 10 3 kDa Diameter: 30–80 nm Composition: –10% protein (apoB-100, apoC, apoE) –20% phospholipids, 50% triglycerides, 20% cholesterol Origin: hepatocytes (apolipoproteins and lipid) Function: Transports hepatic triglycerides to tissues; is converted into LDL

8. Low Density Lipoprotein (LDL) Density: 1.006–1.063 g/ml Mass: 2.3 x 10 3 kDa Diameter: 18–25 nm Composition: –25% protein (apoB-100) –20% phospholipids, 10% triglycerides, 45% cholesterol Origin: derived from VLDL (by loss of triglycerides) Function: major carrier of cholesterol to liver and other tissues; regulates cholesterol biosynthesis by LDL receptor-mediated endocytosis

9. High Density Lipoprotein (HDL) Density: 1.063–1.210 g/ml Mass: 0.175–0.360 x 10 3 kDa Diameter: 5–12 nm Composition: –55% protein (apoA) –25% phospholipids, 5% triglycerides, 15% cholesterol Origin: pre-HDL secreted by hepatocytes Function: Loads cholesterol from tissues and delivers it to liver, steroidogenic tissues and other lipoproteins; apoA-1 binds to SR-B1; can release cholesterol without endocytosis

10. Systemic Transport of Lipid and Lipoprotein 1.Chylomicrons transport dietary lipid to tissues; endocytosis of remnants (apoB-48 or apoE) 2.VLDLs transport lipids from liver to tissues; remnants become LDLs or endocytosed by liver (apoB-100 or apoE) 3.LDLs transport cholesterol to liver and other tissues by endocytosis (apoB-100) 4.HDL precursors remove cholesterol from tissues; HDLs deliver cholesterol to the liver and other tissues (apoA) lipoprotein lipase regulated by apoC on lipoproteins

11. Receptor-Mediated Cholesterol Endocytosis Allows sensing of plasma cholesterol by producing cells LDL endocytosis requires LDL receptor (binds apoB-100) genetic deficiency of functional LDL receptors causes familial hypercholesterolemia LRP (lipoprotein receptor-related protein) binds apoE in remnants of chylomicrons and VLDLs Internalized cholesterol interacts with sensors on endoplasmic reticulum, regulating biosynthesis

12. Cell Membrane Transport Proteins ABC (ATP binding cassette) proteins transport cholesterol across cell membranes ABCA1 ubiquitous transport of cholesterol and phospholipid into HDLs –deficiency causes Tangier’s disease ABCG1 macrophage-specific transporter Hepatic and intestinal ABCG5/8 export cholesterol and other sterols into bile and lumen, respectively –deficiency causes β -sitosterolemia

13. Reverse Cholesterol Transport Secreted ApoA forms pre-HDL by acquiring transported lipids Cholesterol and phospholipid is transported across cell membrane by ABCA1 (and ABCG1) Cholesterol is esterified by plasma lecithin-cholesterol acyl transferase (LCAT) using lecithin from the cell Mature HDL binds to its receptor SRB1 on liver and steroidogenic cells, and delivers its cholesterol Cholesterol can be transferred to other lipoproteins by cholesteryl ester-transfer protein

14. Regulation of HMG-CoA Reductase Regulating amount of enzyme (200x) Transcription: 8x ↑ mRNA levels Translation: 5x ↑ protein synthesis Protein turnover: 5x ↓ protein degradation Regulating activity of enzyme by AMP-activated protein kinase –high [AMP] inhibits (phosphorylates) HMG-CoA reductase by hormone-dependent kinases / phophatases –glucagon inhibits (phosphorylates) HMG-CoA reductase –insulin activates (dephosphorylates) HMG-CoA reductase

15. Regulation of Sterol Biosynthetic Genes Steroid Regulatory Element Binding Protein (SREBP) is retained in ER by SCAP (SREBP cleavage-activating protein) Low cholesterol allows migration of SREBP from ER to golgi where it can be cleaved by 2 proteases Cleavage of SREBP allows N-terminal fragment to enter nucleus and activate steroid biosynthetic genes

16. Cholesterol Regulation of Transcription and Protein Turnover of HMG-CoA Abbreviations SRE: sterol response element (DNA sequence) SREBP: SRE binding protein (transcription factor) SCAP: SREBP cleavage-activating protein (SREBP chaperone) INSIG: Insulin-induced gene 1 protein (ER retention) SSD: sterol-sensing domain (5 membrane spanning helices) S1P: Site 1 protease S2P: Site 2 protease HMG-CoAR: HMG-CoA reductase LDLR: LDL receptor INSIG from Ikonen (2008) Nature Rev. Mol. Cell Biol. 9:125

17. Transcription of Lipid Biosynthetic Enzymes Regulated by SREBP from Horton, Goldstein, Brown (2002) J. Clin. Invest. 109:1125

18. Regulation of Cholesterol Biosynthesis Hormones regulate the phosphorylation of HMG-CoA reductase: –Phosphorylated – Inactive –Dephosphorylated – Active High intracellular cholesterol: –stimulates ACAT for cholesterol storage –lowers expression of LDL receptor gene