Characterisation of the potentially diverse transcriptional regulatory mechanisms of human coagulation PZ gene.

Protein Z (PZ) is a vitamin k-dependent plasma glycoprotein produced mainly in the liver, which circulates as a complex with PZ-dependent protease inhibitor. Human PZ has been reported as having both procoagulant and anticoagulant activity, but its anticoagulant role appears to be more physiologically relevant. PZ levels vary widely among healthy individuals (normal range of 0.6 to 5.7 mug/mL) with the average being higher in men than women. The aim of this study was to investigate the mechanisms by which PZ expression is controlled at the level of transcription, including whether hormonal and/or inflammatory signals modify expression and, therefore, possibly account, at least in part, for the wide normal range.Bioinformatic analysis enabled the identification of two age stable elements (ASEs) in the PZ gene promoter showing close homology to those found in two evolutionarily related proteins, human factor IX and protein C. These regions were found to be bound specifically by potentially the same or a similar protein; but the identity of the protein could not be confirmed as either PEA3 or Ets1, although transactivation data from HepG2 cells suggested PEA3 is important in controlling expression of endogenous PZ.Two further regions, proximal and distal to the PZ transcription start site, were identified by bioinformatic analysis for binding of hepatocyte nuclear factor 4 alpha (HNF4alpha), with the proximal site also reported by Sugawara and colleagues. Although both sites showed specific binding of protein only the protein binding the proximal site was confirmed as being HNF4alpha. The importance of HNF4alpha was further demonstrated by overexpression and siRNA knockdown producing a respective increase and decrease in PZ mRNA levels. Furthermore, the role of steroid hormones and inflammatory signals as potential modifiers of PZ transcription were also assessed in the HepG2 model system. This study has demonstrated, to varying degrees, the importance of tissue-specific factors (HNF4alpha), more ubiquitous factors (PEA3, Ets1), steroid hormones (oestrogen and glucocorticoid) and inflammatory pathways (bacterial LPS induction) on the regulation of PZ gene transcription.