Download PDF
Thromb Haemost 2003; 90(06): 1029-1039
DOI: 10.1160/TH03-07-0443
Blood Coagulation, Fibrinolysis and Cellular Haemostasis
Schattauer GmbH

Characterization of the human protein S gene promoter: a possible role of transcription factors Sp1 and HNF3 in liver

Hideki Tatewaki
1   Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan
,
Hiroko Tsuda
1   Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan
,
Taisuke Kanaji
1   Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan
,
Kazunari Yokoyama
2   Tsukuba Life Science Center, The Institute of Physical and Chemical Research (RIKEN), Ibaraki, Japan
,
Naotaka Hamasaki
1   Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan
› Author AffiliationsFinancial support: This work was supported in part by grants from the Ministry of Education, Science, Sports, and Culture of Japan (to NH) and by grants from the Magnetic Health Science Foundation (to NH).
Further Information

Publication History

Received 07 July 2003

Accepted after revision 18 August 2003

Publication Date:
05 December 2017 (online)

    Permissions and Reprints

Summary

Protein S is a vitamin-K-dependent plasma glycoprotein that serves as a cofactor for activated protein C in the protein C anticoagulation pathway, which regulates blood coagulation by inactivating factors Va and VIIIa. Mechanisms regulating the expression of the protein S gene remain unknown to date. The aim of this study was to characterize the cis-acting DNA elements of the human protein S gene in liver.

We found that liver cell lines (HepG2 and PLC) transcribed the human protein S gene to mRNA, whereas non-liver cell lines (HEK293 and HeLa cells) either transcribed the gene weakly or not at all. Isolation and analysis of tissue-specific gene expression in HepG2 and HeLa cells of the 5’-flanking region from-6183 to +294 of the protein S gene indicated that the consensus binding motifs to HNF3 and Sp1 or MAZ transcription factors in the flanking region are essential for protein S gene expression. Exogenous expression of the Sp1 gene augmented the protein S-reporter gene expression in HepG2 or PLC cells but had no effect in HeLa cells. Taken together, we would conclude that transcription factors of HNF3, MAZ, and Sp1 are required for high-level expression of the protein S gene in hepatic cells, but in non-hepatic cells such as HeLa cells, an unknown factor(s) binds to the Sp1 region and disturbs the action of Sp1 and MAZ.

Keywords

Gene expression - protein C/S pathway - transcription factors
 

  • References

  • 1 Allaart CF, Briet E. Familial venous thrombophilia. In: Haemostasis and Thrombosis. Bloom AL, Forbes CB, eds. Edinburgh: Churchill Livingstone 1994; 1349-60.
    PubMedGoogle Scholar
  • 2 Dahlbäck B, Carlsson M, Svensson PJ. Familial thrombophilia due to a previously unrecognized mechanism characterized by poor anticoagulant response to activated protein C: prediction of a cofactor to activated protein C. Proc Natl Acad Sci U S A 1993; 90: 1004-8.
    CrossrefPubMedGoogle Scholar
  • 3 Bertina RM, Koeleman BP, Koster T. et al. Mutation in blood coagulation factor V associated with resistance to activated protein C. Nature 1994; 369: 64-7.
    CrossrefPubMedGoogle Scholar
  • 4 Rosendaal FR. Risk factors for venous thrombosis: prevalence, risk, and interaction. Semin Hematol 1997; 34: 171-87.
    PubMedGoogle Scholar
  • 5 Tsuda H, Hattori S, Tanabe S. et al. Screening for aetiology of thrombophilia: a high prevalence of protein S abnormality. Ann Clin Biochem 1999; 36: 423-32.
    CrossrefPubMedGoogle Scholar
  • 6 Watkins PC, Eddy R, Fukushima Y. et al. The gene for protein S maps near the centromere of human chromosome 3. Blood 1988; 71: 238-41.
    PubMedGoogle Scholar
  • 7 Edenbrandt CM, Lundwall A, Wydro R. et al. Molecular analysis of the gene for vitamin K dependent protein S and its pseudogene cloning and partial gene organization. Biochemistry 1990; 29: 7861-8.
    CrossrefPubMedGoogle Scholar
  • 8 Schmidel DK, Tatro AV, Phelps LG. et al. Organization of the human protein S gene. Biochemistry 1990; 29: 7845-52.
    CrossrefPubMedGoogle Scholar
  • 9 Pools van Amstel HK, Reitsma PH, Van der Logt PE. et al. Intron-exon organization of the active human protein S gene PSα and its pseudogene PSβ 1: duplication and silencing during primate evolution. Biochemistry 1990; 29: 7853-61.
    CrossrefPubMedGoogle Scholar
  • 10 Fair DS, Marlar RA. Biosynthesis and secretion of factor VII, protein C, protein S, and the protein C inhibitor from a human hepatoma cell line. Blood 1986; 67: 64-70.
    PubMedGoogle Scholar
  • 11 Fair DS, Marlar RA. Human endothelial cells synthesize protein S. Blood 1986; 67: 1168-71.
    PubMedGoogle Scholar
  • 12 Stern DM, Brett J, Harris K. et al. Participation of endothelial cells in the protein C-protein S anticoagulant pathway: the synthesis and release of protein S. J Cell Biol 1986; 102: 1971-8.
    CrossrefPubMedGoogle Scholar
  • 13 Schwarz HP, Heeb MJ, Wencel-Drake JD. et al. Identification and quantitation of protein S in human platelets. Blood 1985; 66: 1452-5.
    PubMedGoogle Scholar
  • 14 Ogura M, Tanabe N, Nishioka J. et al. Biosynthesis and secretion of functional protein S by a human megakaryoblastic cell line (MEG-01). Blood 1987; 70: 301-6.
    PubMedGoogle Scholar
  • 15 Malm J, He X, Bjartell A. et al. Vitamin K-dependent protein S in leydig cells of human testis. Biochem J 1994; 302: 845-50.
    CrossrefPubMedGoogle Scholar
  • 16 He X, Shen L, Bjartell A. et al. The gene encoding vitamin K-dependent anticoagulant protein S is expressed in multiple rabbit organs as demonstrated by northern blotting, in situ hybridization, and immunohistochemistry. J Histochem Cytochem 1995; 43: 85-96.
    CrossrefPubMedGoogle Scholar
  • 17 Maillard C, Berruyer M, Serre CM. et al. Protein S, a vitamin K-dependent protein, is a bone matrix component synthesized and secreted by osteoblasts. Endocrinology 1992; 130: 1599-1604.
    PubMedGoogle Scholar
  • 18 Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidium thiocyanate-phenol-chloroform extraction. Anal Biochem 1987; 162: 156-9.
    PubMedGoogle Scholar
  • 19 Dignam JD, Lebovitz RM, Roeder RG. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res 1983; 11: 1475-89.
    CrossrefPubMedGoogle Scholar
  • 20 He XD, Dahlbäck B. Molecular cloning, expression and functional characterization of rabbit anticoagulant vitamin-K-dependent protein S. Eur J Biochem 1993; 217: 857-65.
    CrossrefPubMedGoogle Scholar
  • 21 Azizkhan JC, Jensen DE, Pierce AJ. et al. Transcription from TATA-less promoters: dihydrofolate reductase as a model. Crit Rev Euk Gene Expr 1993; 3: 229-54.
    PubMedGoogle Scholar
  • 22 Gill G, Pascal E, Tseng ZH. et al. A gluta-mine-rich hydrophobic patch in transcription factor Sp1 contacts the dTAFII110 component of the drosophilia TFIID complex and mediates transcriptional activation. Proc Natl Acad Sci U S A 1994; 91: 192-6.
    CrossrefPubMedGoogle Scholar
  • 23 Goodrich JA, Tjian R. TBP-TAF complexes: selectivity factors for eukaryotic transcription. Curr Opin Cell Biol 1994; 13: 5710-24.
    PubMedGoogle Scholar
  • 24 Pugh BF, Tjian R. Mechanism of transcriptional activation by Sp 1 evidence of coactiva-tors. Cell 1990; 61: 1187-97.
    CrossrefPubMedGoogle Scholar
  • 25 Parks CL, Shenk T. The serotonin 1a receptor gene contains a TATA-less promoter that responds to MAZ and Sp1. J Biol Chem 1996; 271: 4417-30.
    CrossrefPubMedGoogle Scholar
  • 26 Dahlbäck B. Inhibition of protein Ca cofactor function of human and bovine protein S by C4b-binding protein. J Biol Chem 1986; 261: 12022-7.
    PubMedGoogle Scholar
  • 27 Arenzena N, Rodriguez de Cordoba S. Promoter region of the human gene coding for β-chain of C4b binding protein. J. Immunol 1996; 156: 168-75.
    PubMedGoogle Scholar
  • 28 Tsay W, Lee YM, Shen MC. et al. Synergistic transactivation of HNF-1α, HNF-3, and NF-1 contributes to the activation of the liver-specific protein C gene. DNA Cell Biol 1997; 16: 569-77.
    CrossrefPubMedGoogle Scholar
  • 29 Hayashi T, Usui M, Nishioka J. et al. Regulation of the human protein C inhibitor gene expression in HepG2 cells: role of Sp1 and AP2. Biochem J 1998; 332: 573-82.
    CrossrefPubMedGoogle Scholar
  • 30 Rance JB, Follows GA, Cockerill PN. et al. Regulation of the human endothelial cell protein C receptor gene promoter by multiple Sp1 binding sites. Blood 2003; 101: 4393-401.
    CrossrefPubMedGoogle Scholar