79 Molecular and Cellular Biochemistry 169: 79–83, 1997. © 1997 Kluwer Academic Publishers. Printed in the Netherlands. Receptor- ‘C k ’ dependent signalling regulates the LDL-receptor gene transcription Rajeev Goel, Jarnail Singh and D. Kaul Molecular Biology Unit, Department of Experimental Medicine and Biotechnology, Post Graduate Institute of Medical Education and Research, Chandigarh-160012, India Received 5 February 1996; accepted 20 August 1996 Abstract Evidence of a novel receptor, having affinity for cholesterol in lipoproteins and intrinsic tyrosine kinase activity (designated as Receptor ‘C k ’) prompted us to explore its role in LDL-dependent transcriptional regulation of apoprotein-B specific LDL receptor gene in human lymphocytes. The results of this study revealed that LDL-dependent activation of this Receptor ‘C k ’ was essen- tial for the regulation of apoprotein-B specific LDL receptor gene transcription. Further, by using various blockers of as well as simulating the Receptor ‘C k ’-dependent signalling in human lymphocytes, we were able to show that Receptor ‘C k ’-dependent signalling was involved in the LDL-dependent transcriptional regulation of 160 kDa apoprotein B specific LDL receptor gene. Based upon these results, we conclude that cholesterol derived from LDL catabolism within the cell does not have any role in the transcriptional regulation of apoprotein B specific LDL receptor gene. (Mol Cell Biochem 169: 79–83, 1997) Key words: LDL, signalling, Receptor ‘C k ’, transcription, LDL receptor gene Introduction Based upon our recent findings [1–8] we proposed that LDL has two types of cellular receptors responsible for: (a) Cho- lesterol/lipid endocytosis through conventional apoprotein- B specific 160 kDa receptor and b) transmembrane signalling through cholesterol specific 69 kDa receptor, having intrin- sic tyrosine kinase activity, designated by us as Receptor ‘C k ’. Further we were also able to show that this Receptor ‘C k ’- dependent signalling not only regulates mevalonate pathway (5 & 7) but this receptor also interacts with cholesterol moi- ety in LDL in calcium independent manner (4) which is in contrast with calcium dependent interaction of LDL with conventional 160 kDa apoprotein-B specific LDL receptor (9). Keeping in view that extracellular calcium level dictates whether or not LDL binds to either its conventional apopro- tein B specific LDL receptor or Receptor ‘C k ’, the present investigation was addressed to understand the role of Recep- tor ‘C k ’-dependent signalling in the regulation of the conven- tional apoprotein B specific LDL receptor gene at the transcriptional level. Address for offprints: D. Kaul, Molecular Biology Unit, Department of Experimental Medicine and Biotechnology, Post Graduate Institute of Medical Education and Research, Chandigarh-160012, India Materials and methods Materials Phosphatidic acid (PA), dibutyryl cAMP (db cAMP), dibutyryl cGMP (db cGMP), Histopaque and other chemicals were procured from Sigma Chemical Company (St. Louis, USA). Nucleic Acid Labelling and Detection Kit was procured from Boehringer Mannheim, Germany. All other reagents were of the highest quality available. Lymphocytes incubation experiments Lymphocytes were separated from healthy individuals hav- ing normal serum lipoprotein profile by making use of Ficoll- Hypaque density gradient and LDL fraction was isolated from the fresh fasting plasma of normal healthy donors [10]. Iso- lated lymphocytes were cultured for 24 h at 37°C in DMEM supplemented with 10% of lipoprotein deficient serum (LPDS) in 5% CO 2 atmosphere. At the end of this incuba-