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1 Extracellular adenosine 5'-triphosphate (ATP) and uridine 5'-triphosphate (UTP) have been shown to activate a nucleotide receptor (P 2U receptor) in rat mesangial cells that mediates phosphoinositide and phosphatidylcholine hydrolysis by phospholipases C and D, respectively. This is followed by an increased activity of the mitogen-activated protein kinase cascade and cell proliferation. Here we show that ATP and UTP potently stimulate the stress-activated protein kinase pathway and phosphorylation of the transcription factor c-Jun. 2 Both nucleotides stimulated a rapid (within 5 min) and concentration-dependent activation of stressactivated protein kinases as measured by the phosphorylation of c-Jun in a solid phase kinase assay. 3 When added at 100 mM the rank order of potency of a series of nucleotide analogues for stimulation of c-Jun phosphorylation was UTP4ATP=UDP=ATPgS42-methylthio-ATP4bg-imido-ATP= ADP4AMP=UMP=adenosine=uridine. Activation of stress-activated protein kinase activity by ATP and UTP was dose-dependently attenuated by suramin. 4 Down-regulation of protein kinase C-a, -d and -e isoenzymes by 24 h treatment of the cells with 12-O-tetradecanoylphorbol 13-acetate did not inhibit ATP-and UTP-induced activation of c-Jun phosphorylation. Furthermore, the speci®c protein kinase C inhibitors, CGP 41251 and Ro 31-8220, did not inhibit nucleotide-stimulated c-Jun phosphorylation, suggesting that protein kinase C is not involved in ATP-and UTP-triggered stress-activated protein kinase activation. 5 Pretreatment of the cells with pertussis toxin or the tyrosine kinase inhibitor, genistein, strongly attenuated ATP-and UTP-induced c-Jun phosphorylation. Furthermore, N-acetyl-cysteine completely blocked the activation of stress-activated protein kinase in response to extracellular nucleotide stimulation. 6 In summary, these results suggest that ATP and UTP trigger the activation of the stress-activated protein kinase module in mesangial cells by a pathway independent of protein kinase C but requiring a pertussis toxin ± sensitive G-protein and tyrosine kinase activation.

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The presence of sequences related to the agr of Staphylococcus aureus was demonstrated in Staphylococcus epidermidis by agr-specific PCR, and Southern blot. The agr-like locus of S. epidermidis A086 was cloned and sequenced. An overall homology of 68% was found between the agr locus from S. epidermidis and S. aureus. The agr locus from S. epidermidis was organized similar to those from S. aureus and S. lugdunensis. The putative RNAII molecule contains four open reading frames, agr A, B, C and D. AgrA was a response regulator. AgrB showed homology with transducer and translocase molecules. AgrC is expected to act as a histidine protein kinase in which a leucine zipper is present. AgrD is presumably processed into an autoinducer peptide. The putative RNAIII molecule contained an open reading frame encoding a putative 26 amino acid (aa) polypeptide, which differed in 3 aa from the RNAIII encoded delta-toxin of S. aureus. Kinetic studies showed that the production of this RNAIII was elevated during the post-exponential phase. delta-Toxin activity was demonstrated for 21 of 23 tested S. epidermidis strains. Kinetic studies of the production of delta-toxin showed that the toxin was produced during the post-exponential phase. Sequencing of S. epidermidis A097, which showed a delayed agr-response, revealed a truncated AgrC lacking the histidine kinase domain. These data indicate that an agr-like locus is active in S. epidermidis during the post-exponential phase.

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