Low Concentration N-methyl-N'-nitro-N-nitrosoguanidine Activates DNA Polymerase-Beta Expression via cyclic-AMP-protein Kinase A-cAMP Response Element Binding Protein Pathway

Mutat Res. 2001 Jul 1;478(1-2):177-84. doi: 10.1016/s0027-5107(01)00146-4.

Abstract

Ultraviolet light (UV), ionizing-radiation or alkylating agents are known as carcinogens, mostly because of their ability to damage DNA directly. However, they may also play a diverse role in activating the signal pathways and altering the gene expression. We have shown previously that N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) of 0.2 microM can increase the transcription of DNA polymerase-beta gene, which has a cyclic AMP response element (CRE) motif in its promoter. Using the CRE report vector, we show here, such treatment can stimulate the CRE-driven gene expression by approximately 1.5-fold compared with control. Consistent with it, the proportion of ser-133 phosphorylated CRE binding protein (CREB), the related transcription factor was 2.08-fold higher versus control in vero cells after 60 min of MNNG treatment. Although CREB has many putative kinases for its phosphorylation, such as p38 mitogen-activated protein kinase (p38 MAPK), Ca(2+)/calmodulin-dependent protein kinase Pi (CaMK Pi) and protein kinase C (PKC), we found the protein kinase A (PKA) was activated and its activation peaked when cells were treated for 60 min (with arbitrary activity unit of 11.03+/-2.80 and 0.86+/-0.43 in treatment and control, respectively), this phasic character was similar to that of the CREB phosphorylation. We also determined the intracellular cyclic AMP (cAMP) levels and it was found that the cAMP concentration was elevated after 60 min treatment (1.53-fold higher). However, to our surprise, we did not find any accompanying cAMP elevation in cells treated by MNNG for 30 min, in which PKA was activated significantly. These findings, together with other observations, suggest that cAMP-PKA-CREB signaling pathway mediates the low concentration MNNG induced pol-beta expression. In addition to elevated cAMP, there might exist a cAMP-independent PKA activation manner in this course.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chlorocebus aethiops
  • Cyclic AMP / metabolism
  • Cyclic AMP / physiology
  • Cyclic AMP Response Element-Binding Protein / genetics
  • Cyclic AMP Response Element-Binding Protein / metabolism*
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • DNA Polymerase beta / genetics
  • DNA Polymerase beta / metabolism*
  • Dose-Response Relationship, Drug
  • Enzyme Activation / drug effects
  • Gene Expression Regulation / drug effects
  • Genetic Vectors / genetics
  • Methylnitronitrosoguanidine / pharmacology*
  • Phosphorylation / drug effects
  • Response Elements / genetics
  • Signal Transduction / drug effects
  • Time Factors
  • Transcription, Genetic / drug effects
  • Vero Cells

Substances

  • Cyclic AMP Response Element-Binding Protein
  • Methylnitronitrosoguanidine
  • Cyclic AMP
  • Cyclic AMP-Dependent Protein Kinases
  • DNA Polymerase beta