Transcriptional Repression of Protein Kinase Cα via Sp1 by Wild Type p53 Is Involved in Inhibition of Multidrug Resistance 1 P-Glycoprotein Phosphorylation

Abstract

The protein kinase C (PKC) family consists of serine/ threonine protein kinases that play important roles in signal transduction, cell proliferation, and tumor formation. Recent studies found that PKCs are commonly overexpressed in human tumors, including soft tissue sarcoma (STS). Overexpression of PKCs contributes to invasion and migration of tumor cells and induction of angiogenesis. PKC can also phosphorylate the multidrug resistance (MDR) gene-encoded P-glycoprotein and induce MDR phenotype. Our previous studies showed that mutation of p53 enhanced STS metastasis and mediated the MDR phenotype. Restoring wild type (WT) p53 in STS cells containing mutant p53 sensitized the cells to chemotherapy. In the present study, we found that PKC␣ protein expression is inhibited by WT p53 partly due to reduced PKC␣ mRNA expression in STS cells, but p53 does not affect PKC␣ mRNA stability. Deletion and mutation analysis of the PKC␣ promoter fused to the luciferase reporter gene identified a Sp1 binding site (؊244/؊234) in the PKC␣ promoter that is required for p53-mediated inhibition of PKC␣ promoter activity. More importantly, PKC␣ phosphorylates and activates MDR1 P-glycoprotein, whereas inhibition of PKC␣ by p53 leads to decreased MDR1 phosphorylation in STS cells, which sensitizes STS cells to chemotherapeutic agents. These data indicate that WT p53 may resensitize STS to chemotherapeutic agents by reducing MDR1 phosphorylation via transcriptional repression of PKC␣ expression. Thus, molecular-based therapies targeting mutant p53 and PKC␣ may be an effective new strategy to improve chemotherapeutic efficacy in STS.The protein kinase C (PKC) 1 family consists of a large number of serine/threonine kinases that are activated by extracellular signals. In mammalian cells, the PKC family is further divided into three subfamilies, conventional PKCs (␣, ␤I, ␤II, ␥), novel PKCs (␦, ⑀, , ), and atypical PKCs ( , , ) (1). PKC is an additional PKC family member that was discovered recently and is known as protein kinase D (2). A distantly related PKC family includes three isoforms of the PKC-related kinases, known as PKN1, PKN2, and PKN3 (3). Although PKC family members share primary sequence similarities, the subfamily members have different enzymatic activation profiles. Specifically, conventional PKC isoforms are activated by 1,2-diacylglycerol and phosphatidylserine (PS) in a calcium-dependent manner, novel PKC members are activated only by 1,2-diacylglycerol and PS, and atypical PKC members are activated by PS alone as a co-factor (4). In resting cells, PKC predominantly resides in the cytosol in an inactive state and, upon stimulation, translocates as an active kinase to discrete subcellular locations, such as the plasma membrane, membranous vesicles, cytoskeleton, mitochondria, and nucleus (5).PKC functions have been the subject of intense study for many years after identification of PKC as the intracellular receptor for tumor-promoting phorbol esters such as tissuetype plasminogen activator. Studies have shown that PKC a...

Rad51 overexpression contributes to chemoresistance in human soft tissue sarcoma cells: a role for p53/activator protein 2 transcriptional regulation

“…We explained this observation as due to p53 induced down-regulation of MDR1/Pgp expression with concordantly reduced activity (3,4). However, it is likely that the wtp53-induced chemosensitivity is a multifactorial process.…”

“…Previously, we have studied MDR-1/P-glycoprotein expression and activity in STS chemoresistance (3,4). This transmembrane efflux transporter reduces intratumoral accumulation/retention of cytotoxic drugs and may induce the intracellular redistribution of these agents away from the putative site of action (e.g., outside of the nucleus).…”

Chemosensitizing multiple drug resistance of human carcinoma by bicyclol involves attenuated P-glycoprotein, GST-P and Bcl-2

“…27,[29][30][31] We have found at least two intracellular pathways in Vin R KB and Adr R MCF-7 cells that involve the increased level of GSH/GST activity and the overexpression of Bcl-2.…”

“…Previous studies notified that the mutation of Mdr-1 gene could upregulate the p-glycoprotein expression and PKC-mediated phosphorylation and other associated proteins could directly modify the p-glycoprotein function. [25][26][27][28] Studies are underway to investigate whether Bicyclol affects those correlated mechanisms for reducing p-glycoprotein expression and mediated MDR in both Vin R KB and Adr R MCF-7 cells.…”

Regulation of Class II β-Tubulin Expression by Tumor Suppressor p53 Protein in Mouse Melanoma Cells in Response toVincaAlkaloid

“…The development of broad-spectrum drug resistance is often characterized by overexpression of the transmembrane efflux pump, P-glycoprotein, which is encoded by the multidrug resistance-1 (MDR1) gene, or of the MDR-associated proteins (19)(20)(21). Additionally, alterations of microtubule proteins are thought to be associated with resistance to antimicrotubule drugs and altered expression of several h-tubulin isotypes in drugresistant cells has been reported in many cases (8,18,(22)(23)(24).…”