Critical Role of Diacylglycerol- and Phospholipid-Regulated Protein Kinase Cε in Induction of Low-Density Lipoprotein Receptor Transcription in Response to Depletion of Cholesterol

Abstract

Protein kinase C (PKC) represents a family of serine/threonine protein kinases that plays a key role in signal transduction and regulation of gene expression (11,39,40,43). Molecular cloning and biochemical studies have revealed at least 12 PKC subspecies, which can be classified into three groupsclassic (cPKC), novel (nPKC), and atypical (aPKC)-on the basis of their structures. The PKC family comprises a regulatory domain in the amino terminus and a catalytic domain in the carboxyl terminus. The cPKCs, including PKC␣, -␤I, -␤II, and -␥, have two common regions, C1 and C2, in the regulatory domain. The C1 region has two cysteine-rich loops that are the binding site for diacylglycerol (DAG) and phorbol ester (tetradecanoyl phorbol acetate [TPA]). The C2 region binds to calcium. The nPKCs, including PKC␦, -ε, -, and -, lack the C2 region. The aPKCs, including PKC and -/, lack the C2 region and have only one cysteine-rich loop in the C1 region. Key lipid cofactors of PKCs are DAG, phosphatidylserine (PS), and calcium. All of these are required to activate cPKCs, whereas calcium is not required to activate nPKCs. aPKCs are insensitive to both DAG and calcium. Other lipid species, such as cholesterol sulfate, fatty acids, lysophospholipids, and phosphatidylinositols, also modulate the activity of certain PKC isoforms (3,12,24,41). Several PKC family members displaying responsiveness to calcium, DAG, and fatty acids are expressed in hepatic cells (16,26), but their precise functional roles in cholesterol uptake and biosynthesis are not yet clearly defined.Animal cells regulate their cholesterol content by fine-tuning of the supply of exogenous and endogenous cholesterol (5). Both cholesterol and fatty acids have been demonstrated to be important regulators of plasma low-density lipoprotein (LDL) cholesterol levels due to alterations in LDL receptor activity (6,7,20). The mechanism by which sterols modulate LDL receptor gene transcription has been examined in great detail. Involved in this pathway are a family of proteins, designated sterol regulatory element binding proteins (SREBPs), that play an integral role in the feedback pathway by which cholesterol suppresses transcription of LDL receptor gene. The SREBPs are transcription factors that are bound to the endoplasmic reticulum and nuclear envelope by virtue of two membranespanning regions. When these membranes are depleted of sterols, a two-step proteolytic process releases the amino terminal of SREBPs, which then travel to the nucleus and activate LDL receptor transcription via interaction with multiple nuclear factors and coactivators (7,42). When cells are overloaded with sterols, cleavage of SREBPs is inhibited, resulting in suppression of the LDL receptor gene. A similar mechanism has been shown for the regulation of other sterol-responsive genes

Sphingomyelinase D, a novel probe for cellular sphingomyelin

“…However, it should be noted that the regeneration of SM is accompanied by the formation of equimolar amounts of diacylglycerol, a potent activator of protein kinases (31,39). Furthermore, diacylglycerol and other PKC activators stimulate the translocation of intracellular cholesterol to the plasma membrane, whereas PKC inhibitors reduce it (40).…”

Phorbol ester promotes histone H3-Ser10 phosphorylation at the LDL receptor promoter in a protein kinase C-dependent manner

“…Whole cell extracts were prepared by and probed with the required antibody. Immunoreactive peptides on the filters were detected by ECL as described earlier (23)(24)(25). Quantitative analysis of protein levels was performed by densitometric scanning of the autoradiograms and represented three or more independent experiments.…”

“…TPA is a well-known activator of PKC and p42/44 MAPK and dramatically induces LDL receptor expression at the transcriptional level in human hepatoma HepG2 cells (23). We have earlier established these signaling pathways as critical mediators of LDL receptor induction in HepG2 cells by a variety of transcriptional modulators, including TPA (23)(24)(25). Based on the suggested role of p42/44 MAPK cascade in regulating histone H3-Ser10 phosphorylation, the involvement of this kinase raised an interesting possibility that this histone modification may play an important role in LDL receptor induction by TPA.…”

Protein kinase C activation stabilizes LDL receptor mRNA via the JNK pathway in HepG2 cells

“…In this cell line, LDL receptor levels are suppressed in response to cholesterol and lipoprotein loading (14). However, expression of LDL receptors is also regulated through several signal transduction pathways in HepG2 cells, including the cyclic AMP, diacylglycerol-protein kinase C (PKC), and mitogen-activated protein kinase (MAPK) pathways (15)(16)(17). Recent studies have probed the relationships between these signaling cascades in the control of receptor expression.…”