Physical and Functional Interactions between USF and Sp1 Proteins Regulate Human Deoxycytidine Kinase Promoter Activity

Deoxycytidine kinase (EC 2.7.1.74, dCK) is central to drug activity of anticancer and antiviral agents such as cytosine arabinoside (araC) and gemcitabine. HepG2 hepatocellular carcinoma cells were used to study the transcriptional regulation of dCK. 5′-Deletion and site-directed mutagenesis of the dCK upstream region (positions –464 to –27) confirmed the importance of two GC-boxes (positions –317 to –309 and –213 to –206) and two E-boxes (positions –302 to –297 and –278 to –273). In vitro electromobility shift assays with HepG2 nuclear extracts and in vivo chromatin immunoprecipitation assays with HepG2 chromatin extracts confirmed the presence of bound Sp1/Sp3 and USF1/2. Co-transfections in HepG2 cells showed that USF1 and USF2a stimulated and Sp1 repressed promoter activity from a dCK-luciferase reporter gene construct. In Sp- and USF-null Drosophila Mel-2 cells, both Sp1 and USF1 stimulated dCK promoter activity in a dose-dependent manner, however, both Sp3 and USF2a were effectively inert. Combined Sp1 and USF1 showed additive transactivation at lower concentrations of Sp1. Sp1 was inhibitory at higher levels. Stimulation by combined USF1/USF2a with Sp1 was similar to that for USF1 alone with Sp1, whereas transactivation by Sp1 and USF2a without USF1 was synergistic. Physical interactions between USF and Sp proteins were confirmed by immunoprecipitations with Sp- and USF-specific antibodies. Domain mapping of USF1 and USF2a localized the functional interactions between USF and Sp proteins to the DNA binding domain of USF. Identifying the physical and functional interactions between Sp and USF proteins may lead to a better understanding of the basis for differential expression of the dCK gene in tumor cells and may foster strategies for up-regulating dCK gene expression and improving chemotherapy with araC and gemcitabine. Deoxycytidine kinase (EC 2.7.1.74, dCK) is central to drug activity of anticancer and antiviral agents such as cytosine arabinoside (araC) and gemcitabine. HepG2 hepatocellular carcinoma cells were used to study the transcriptional regulation of dCK. 5′-Deletion and site-directed mutagenesis of the dCK upstream region (positions –464 to –27) confirmed the importance of two GC-boxes (positions –317 to –309 and –213 to –206) and two E-boxes (positions –302 to –297 and –278 to –273). In vitro electromobility shift assays with HepG2 nuclear extracts and in vivo chromatin immunoprecipitation assays with HepG2 chromatin extracts confirmed the presence of bound Sp1/Sp3 and USF1/2. Co-transfections in HepG2 cells showed that USF1 and USF2a stimulated and Sp1 repressed promoter activity from a dCK-luciferase reporter gene construct. In Sp- and USF-null Drosophila Mel-2 cells, both Sp1 and USF1 stimulated dCK promoter activity in a dose-dependent manner, however, both Sp3 and USF2a were effectively inert. Combined Sp1 and USF1 showed additive transactivation at lower concentrations of Sp1. Sp1 was inhibitory at higher levels. Stimulation by combined USF1/USF2a with Sp1 was similar to that for USF1 alone with Sp1, whereas transactivation by Sp1 and USF2a without USF1 was synergistic. Physical interactions between USF and Sp proteins were confirmed by immunoprecipitations with Sp- and USF-specific antibodies. Domain mapping of USF1 and USF2a localized the functional interactions between USF and Sp proteins to the DNA binding domain of USF. Identifying the physical and functional interactions between Sp and USF proteins may lead to a better understanding of the basis for differential expression of the dCK gene in tumor cells and may foster strategies for up-regulating dCK gene expression and improving chemotherapy with araC and gemcitabine. Deoxycytidine kinase (NTP:deoxycytidine 5′-phosphotransferase, EC 2.7.1.74; dCK) 1The abbreviations used are: dCKdeoxycytidine kinasearaCcytosine arabinosideCMVcytomegalovirusChIPchromatin immunoprecipitationDSDown syndrome.1The abbreviations used are: dCKdeoxycytidine kinasearaCcytosine arabinosideCMVcytomegalovirusChIPchromatin immunoprecipitationDSDown syndrome. is responsible for the phosphorylation of several deoxyribonucleosides such as 2′-deoxycytidine and purine nucleosides, including 2′-deoxyadenosine and 2′-deoxyguanosine. dCK also catalyzes the rate-limiting step in the phosphorylation of important anticancer and antiviral agents such as cytosine arabinoside (araC), difluorodeoxycytidine (gemcitabine), 2-fluoro-ara-AMP (fludarabine), 2′,3′-dideoxycytidine, and 2-chlorodeoxyadenosine (cladribine) (1.Kufe D.W. Spriggs D.R. Semin. Oncol. 1985; 12: 34-48PubMed Google Scholar, 2.Piro L.D. Blood. 1992; 79: 843-845Crossref PubMed Google Scholar). dCK levels are critical determinants of araC and gemcitabine antitumor activities, and resistance to nucleoside analogs is frequently associated with decreased dCK (3.Grant S. Adv. Cancer Res. 1998; 72: 197-233Crossref PubMed Google Scholar, 4.Galmarini C.M. Mackey J.R. Dumontet C. Leukemia. 2001; 15: 875-890Crossref PubMed Scopus (400) Google Scholar). Furthermore, transfection of the dCK cDNA into dCK-deficient tumor cell lines restores in vitro sensitivities to araC (5.Manome Y. Wen P.Y. Dong Y. Tanaka T. Mitchell B.S. Kufe D.W. Fine H.A. Nat. Med. 1996; 2: 567-573Crossref PubMed Scopus (109) Google Scholar, 6.Hapke D.M. Stegmann A.P. Cancer Res. 1996; 56: 2343-2347PubMed Google Scholar). deoxycytidine kinase cytosine arabinoside cytomegalovirus chromatin immunoprecipitation Down syndrome. deoxycytidine kinase cytosine arabinoside cytomegalovirus chromatin immunoprecipitation Down syndrome. The product of dCK gene is a 30.5-kDa polypeptide that is present at low levels in most tissues (7.Durham J.P. Ives D.H. Mol. Pharmacol. 1969; 5: 358-375PubMed Google Scholar, 8.Arner E.S. Flygar M. Bohman C. Wallstrom B. Eriksson S. Exp. Cell Res. 1988; 178: 335-342Crossref PubMed Scopus (56) Google Scholar, 9.Kawasaki H. Carrera C.J. Carson D.A. Anal. Biochem. 1992; 207: 193-196Crossref PubMed Scopus (21) Google Scholar, 10.Spasokoukotskaja T. Arner E.S. Brosjo O. Gunven P. Juliusson G. Liliemark J. Eriksson S. Eur. J. Cancer. 1995; 31: 202-208Abstract Full Text PDF Scopus (82) Google Scholar). The human dCK gene spans over 34 kb on chromosome 4 (4q13.3-q21.1) and includes a coding region consisting of seven exons, ranging in size from 90 to 1544 bp (11.Song J.J. Walker S. Chen E. Johnson 2nd, E.E. Spychala J. Gribbin T. Mitchell B.S. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 431-434Crossref PubMed Scopus (33) Google Scholar). Promoter activity was localized to a 697-bp upstream fragment, including 386 bp of 5′ upstream region, 250 bp of exon 1, and 61 bp of intronic sequence (11.Song J.J. Walker S. Chen E. Johnson 2nd, E.E. Spychala J. Gribbin T. Mitchell B.S. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 431-434Crossref PubMed Scopus (33) Google Scholar). The dCK promoter is highly GC-rich and lacks a TATA-box but contains a transcription initiator region located adjacent to the major transcription start site at position –146 relative to the start of translation. The initiation region also contains an imperfect E2F binding site (12.Chen E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google Scholar). In vitro DNase I footprint and electromobility shift assays demonstrated binding of Sp1 to two GC-boxes and upstream stimulatory factor (USF) to a critical E-box. Although site-directed mutagenesis of these cis-elements implied their transcriptional importance, the detailed mechanisms by which these binding associations regulate dCK gene expression have not been established. The Sp and USF proteins are ubiquitously expressed, yet both are implicated in the regulation of genes characterized by tissue-specific or developmental patterns of expression (13.Suske G. Gene (Amst.). 1999; 238: 291-300Crossref PubMed Scopus (982) Google Scholar, 14.Henrion A.A. Martinez A. Mattei M.G. Kahn A. Raymondjean M. Genomics. 1995; 25: 36-43Crossref PubMed Scopus (38) Google Scholar). Sp1 and the related Sp3 proteins can exert activating or inhibiting effects on transcription, depending on the cell or promoter context, through binding to GC- or GT-box elements (13.Suske G. Gene (Amst.). 1999; 238: 291-300Crossref PubMed Scopus (982) Google Scholar). The USF proteins belong to the class of b-HLH-ZIP (basic helix-loop-helix leucine zipper) transcription factors, including the nuclear proteins Myc, Max, Mad, Mxi1, AP4, TFEB, TFE3, MiTF, and ADD1 (15.Viollet B. Lefrancois-Martinez A.M. Henrion A. Kahn A. Raymondjean M. Martinez A. J. Biol. Chem. 1996; 271: 1405-1415Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar, 27.Gregor P.D. Sawadogo M. Roeder PubMed Scopus Google Scholar). USF was by to transcription from the promoter and was from cells as two USF1 and USF2a M. P.D. Roeder J. Biol. Chem. 1988; Full Text PDF PubMed Google Scholar, M. J. Biol. Chem. 1988; Full Text PDF PubMed Google Scholar). a was (15.Viollet B. Lefrancois-Martinez A.M. Henrion A. Kahn A. Raymondjean M. Martinez A. J. Biol. Chem. 1996; 271: 1405-1415Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). USF1 and proteins are by two and USF2a and of (15.Viollet B. Lefrancois-Martinez A.M. Henrion A. Kahn A. Raymondjean M. Martinez A. J. Biol. Chem. 1996; 271: 1405-1415Abstract Full Text Full Text PDF PubMed Scopus (181) Google Scholar). USF1 and proteins have similar DNA binding but in their transactivation yet to the as and M. T. Sawadogo M. Res. PubMed Scopus Google Scholar, M. Walker S. Sawadogo M. Gene 1992; 2: Google Scholar). of USF and Sp proteins is their to a of transcriptional interactions with of transcription for interactions between USF and Sp proteins been binding to and B. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, Y. Biochem. J. 2001; PubMed Scopus Google Scholar). the of the GC- and elements in the dCK interactions between USF and Sp proteins to regulate expression of In of dCK promoter and in cells (12.Chen E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google Scholar). the major cis-elements and transcription that regulate dCK in HepG2 human cells and the of physical interactions between the USF and Sp proteins that can regulate dCK promoter activity over a and was from were from and reporter gene and were from The and were by of and the and were by The and the expression of USF2a were by Cell human HepG2 hepatocellular carcinoma cell was from the and was as Y. Biochem. J. 2001; PubMed Scopus Google Scholar). Drosophila Mel-2 cells were from and in with and and at of dCK and dCK promoter was from human DNA from cells and The were into of the dCK promoter was and by and and the dCK promoter was into at the and to the and the was by and and and and and and were with DNA Promoter in transcription factor binding elements were an J. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). were for and mutagenesis with or as The two were and a was the and The were with DNA and into the site of the and site-directed mutagenesis were to the or of for and in a of USF1 and Drosophila and USF1 coding cDNA was by from HepG2 with and and were into the for the were with and into the Drosophila expression of the Sp1 coding sequence from 1988; Full Text PDF PubMed Scopus Google Scholar). the of the as a were with a The were as upstream and The was used as the for The were with and into as the expression the USF1 cDNA from HepG2 cells was with and and the into at the and and were by the expression as and as the site is and the site in is and The was by with the and and as a The were with and and into at the was used to the USF2a were with the and and and and and and The were and a was the and The were into the and in were confirmed by DNA and of dCK-luciferase reporter gene into HepG2 cells and reporter gene assays were as Y. Biochem. J. 2001; PubMed Scopus Google Scholar). were to Mel-2 cells were with of the dCK-luciferase reporter gene and Sp1 of or Sp3 of and for the and USF1 of or USF2a of were for assays the were to cell by the The expression levels of Sp1, and USF2a in Mel-2 cells were by of and extracts from HepG2 cells or Mel-2 were by Res. PubMed Scopus Google Scholar). shift assays with HepG2 and Mel-2 nuclear extracts were as Y. Biochem. J. 2001; PubMed Scopus Google Scholar). were for with of and and assays were in HepG2 cells as Y. Blood. PubMed Scopus Google with Sp1 or for the dCK promoter region was with and –464 to human gene coding exon was also with and to the of the and extracts from Mel-2 cells, with and or and were to with immunoprecipitation and and were with for at 4 at 4 with of USF1 or or by with of for 90 at 4 The were by and with immunoprecipitation and the were by for in of The were through and a The were with Sp1 USF1 or and and to of the in the dCK Promoter by 5′-Deletion and demonstrated that two GC-boxes (positions –317 to –309 and –213 to and and (positions –302 to are important for dCK promoter activity in human cell lines (12.Chen E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google Scholar). the between these elements and their bound transcription factors, a upstream of the dCK gene was from DNA bp of sequence and bp of the coding and into to of dCK promoter sequence to the sequence (12.Chen E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google at was between dCK promoter sequence and the human sequence In to and and the a (positions –278 to of into HepG2 cells in levels of promoter activity of bp from the of decreased activity of 61 bp including the and the in a of promoter to of the for the construct. decreased to of an bp of 5′ sequence including and of the E2F site not in in promoter to with cells, site-directed mutagenesis of the and elements in was inhibitory in reporter gene assays of and relative to the the of both the and elements was the as for the of in of dCK promoter of in activity over the for cells (12.Chen E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google Scholar). The and were also of the two E-boxes in additive of dCK promoter activity of the The and were and these a bp upstream of the start and including the and elements as important to dCK promoter Furthermore, that the is and that can the inhibitory effects from of the or the The of the to activity to an similar to the may the presence of between the and the effects of on In and in of to the and in the dCK were used to the major transcription that bound to the dCK a to both the and the elements was with nuclear extracts from HepG2 cells, were which were by with and a between USF binding to and Sp proteins binding to the Sp1 1, and whereas a USF1 1, and 4 1, and were by Sp1 Sp3 and whereas USF1 or 1, and 4 and decreased and and with the of 1, and and of including a 1, and and decreased the of a the to was with HepG2 nuclear major was which was by or a USF1 was with the including a of USF1 or the and that contains both USF1 and were also HepG2 nuclear extracts and a the to and were and were effectively by or a Sp1 and was a was used in which the was of Sp1 or Sp3 both and the and that both Sp1 and Sp3 were bound to the In vivo binding of the transcription to the dCK promoter in HepG2 chromatin extracts was confirmed by the assays binding of the Sp and USF of proteins to the critical and elements in the dCK Co-transfections of Sp1 and USF1/USF2a in HepG2 the functional of and binding to the dCK were in HepG2 cells the reporter with Sp1, or USF2a expression Sp1 alone a of dCK promoter activity of USF1 or in the of Sp1, in a of activity and USF1 or USF2a was with Sp1, the transactivation with USF1 or was decreased by an for Sp1 with the of mutagenesis with the a functional between Sp and USF with the of shift of Sp1/Sp3 and USF1/USF2a in Mel-2 the functional between Sp and USF proteins in the dCK were in Drosophila Mel-2 cells, which a for these transcription 1988; Full Text PDF PubMed Scopus Google Scholar, S. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, M. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). The reporter gene was with expression for Sp1 or Sp3 and and USF1 or USF2a were with with the expression The transactivation by Sp1 in Mel-2 cells showed a from to of Sp1, a of promoter activity was over USF1 alone showed a dose-dependent transactivation with a of by of Sp1 and USF1 was additive at levels of Sp1 to however, at of Sp1, promoter activity with USF1 at higher levels of Sp1 these of promoter activity at of USF1 In to USF2a to was of the reporter gene Sp1 was with a transactivation of promoter activity was at levels of Sp1 and USF2a were with combined USF1 and between of the USF1 and USF2a and USF1/USF2a In of combined USF1/USF2a without Sp1, was to of with Sp1, combined USF1/USF2a was USF2a and to the effects of USF1 not Although Sp3 binding to the and elements was on with HepG2 nuclear extracts and the the of Sp3 dCK promoter activity into Mel-2 a of promoter activity was Sp3 or was combined with was of combined with USF2a on promoter activity not the binding of these transcription in these nuclear extracts were from Mel-2 cells with USF and Sp1, and used for with the were with extracts from cells the binding was and The of the bound proteins were confirmed as Sp1, and USF2a by with Sp1 and USF and in with Sp1 and USF and assays for USF1 and USF2a binding not the as but also decreased the binding of Sp1 and the that functional or between Sp and USF proteins with the dCK promoter and that these effects are in physical interactions between these proteins on The transcriptional to to Sp1, Sp3 was and in both and with the Sp proteins and with the of the USF of Sp1 and USF for the of physical associations between Sp1 and USF1 were from nuclear extracts from Mel-2 cells with expression for these transcription in for both and USF1 and USF2a were with to these both USF1 and also Sp1, that Sp1 with the USF of the USF1 and USF2a for dCK or with functional with HepG2 and Mel-2 cells a for ranging regulation of the dCK promoter by the Sp and USF of whereas USF1 and Sp1 were both USF2a and Sp3 were effectively inert. promoter activity was repressed at levels of Sp1, and Sp1 combined with USF2a was highly synergistic. binding associations between Sp1 and USF1/USF2a proteins were implied by and were confirmed by with USF-specific antibodies. the USF responsible for dCK transactivation and the or functional interactions between Sp1 and USF a of USF were for for and by of Mel-2 cells with with or without Sp1. USF1 and USF2a to DNA as and the leucine and helix-loop-helix M. T. Sawadogo M. Res. PubMed Scopus Google Scholar, M. Walker S. Sawadogo M. Gene 1992; 2: Google Scholar). for the of of USF were to the region and the helix-loop-helix two and for and for for and the leucine the with USF1 levels of USF1 and Sp1 proteins were on and of the from the of USF1 USF1 with and without Sp1, a for region both promoter activity was of an to position in a in the of USF1 transactivation in the of of was by a of to and the additive of USF1 with Sp1. that are in a functional between USF1 and Sp1. The USF1 and the and leucine were inert. effects on Sp1 that the between Sp1 and USF1 also the DNA binding the with USF2a USF2a and Sp1 levels on were and the of of the by the dCK promoter The effects of USF2a with Sp1 on dCK promoter activity were and was by of with and activity decreased and at of the for of exon 4 not the with Sp1 but also decreased promoter activity that with Sp1, alone higher for and the of exon in decreased activity with the and the with the construct. of 1, and 4 and in a similar activity to the of exon not whereas of the region the transactivation with Sp1, that DNA binding of USF2a is for functional with Sp1. as with for the DNA binding are to with Sp1. in the of the effects of the DNA binding a transactivation the The exon is an whereas exon 4 as a in the presence of exon exon 4 and the region are critical for the with Sp1. of a of nucleoside analogs araC and that are important to chemotherapy by dCK is a rate-limiting step in drug (1.Kufe D.W. Spriggs D.R. Semin. Oncol. 1985; 12: 34-48PubMed Google Scholar, 2.Piro L.D. Blood. 1992; 79: 843-845Crossref PubMed Google Scholar). dCK levels in cells with to araC T. T. Tanaka A. M. 1998; 31: PubMed Scopus Google Scholar). study of determinants of araC in with a dCK expression in araC sensitivities Y. Blood. 1999; Google Scholar). dCK promoter activity was localized to a 697-bp upstream fragment, including 386 bp of 5′ sequence 250 bp of exon 1, and 61 bp of intronic sequence (11.Song J.J. Walker S. Chen E. Johnson 2nd, E.E. Spychala J. Gribbin T. Mitchell B.S. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 431-434Crossref PubMed Scopus (33) Google Scholar, E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google Scholar). The dCK promoter is highly GC-rich and lacks a of Sp1 to two GC-boxes and USF to a to critical for promoter activity in (12.Chen E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google Scholar). in on the of levels of dCK in HepG2 cells by used cell to the transcriptional regulation of the dCK and confirmed that the two GC-boxes and and the (12.Chen E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google important and that and were and that was functional was also which to an important of USF and Sp proteins their to a of transcriptional interactions with of transcription including H. U. T. J. 1998; PubMed Scopus Google Scholar, P. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Scopus (21) Google P. M. Sawadogo M. J. 2001; PubMed Scopus Google nuclear A. J. P. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google and the transcription C.M. Roeder 1995; PubMed Scopus Google Scholar, M. M. Roeder Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google and M. P. Roeder PubMed Scopus Google Scholar, H. P.D. Martinez E. Roeder J. PubMed Scopus Google Scholar). In the dCK the elements and the elements are functional interactions between Sp1/Sp3 proteins bound to the and bound to the elements can to regulate gene expression over a with the dCK including both the and elements and nuclear extracts from HepG2 In to the major and as Sp and USF proteins (12.Chen E.H. Johnson 2nd, E.E. Vetter S.M. Mitchell B.S. J. Clin. Invest. 1995; 95: 1660-1668Crossref PubMed Scopus (37) Google two and were that by and assays were to both the Sp and and USF and of physical interactions between these two of binding to the and Sp1/Sp3 binding to the were demonstrated on with the and In vivo binding of Sp1, and to the dCK promoter between –464 and was confirmed by of HepG2 cells with a dCK promoter reporter gene and Sp1, and USF2a expression that functional interactions between Sp1 and the physical and functional interactions between Sp proteins and were to Mel-2 Sp1 and USF1 the dCK however, Sp3 and USF2a were inert. of Sp1 and USF1 were additive at low levels of Sp1, but promoter activity decreased at higher Sp1 levels. In the effects of Sp1 and USF2a on dCK promoter activity were highly synergistic. The of USF1 USF1/USF2a decreased the between Sp1 and USF2a to a from that for USF1 and Sp1. USF and Sp proteins regulate dCK promoter activity over a relative levels and of Sp1 and as as of USF1 and Sp and USF expression were into Mel-2 cells, and nuclear extracts were for shift and Physical interactions between Sp1 and USF1 or USF2a were by the decreased levels of Sp1 binding to on in the presence of and by with USF1 and USF2a antibodies. for an between Sp1 and USF1/USF2a was by of Sp1 with USF1 and these that both Sp and USF proteins are for transactivation of the dCK promoter and that physical interactions between these proteins to interactions between Sp1/Sp3 and USF proteins were to for the of human promoter B. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). of the human promoter that interactions between USF1 binding to the and Sp1/Sp3 binding to the were in the transcriptional regulation of important gene Y. Biochem. J. 2001; PubMed Scopus Google Scholar). on the human promoter have also implicated interactions as critical to J. J. S. G. M. Y. J. and H. for interactions between Sp1/Sp3 and USF proteins can to a for tissue-specific gene by that Sp1 and USF proteins can from Mel-2 the that a physical between these proteins may to a functional The of USF1 and USF2a responsible for dCK transactivation and functional interactions with Sp1 were by mutagenesis and with in and USF-null Mel-2 with USF1 both and and that also were critical to functional interactions between USF1 and Sp1. the USF1 DNA binding the region, and leucine repressed the activating effects of Sp1 on the dCK a functional between region and Sp1. two 1, and and two and were yet the DNA binding by were to a transactivation in with Sp1. from USF1 was in of the between the USF2a and USF1 DNA binding yet functional between these related transcription a from of exon 4 not the with Sp1 but also decreased promoter activity that with Sp1, that may into for tissues in which is present at a by on the of dCK transactivation by Sp1, and USF1 and been to transcriptional activity of the major class I gene in C. Sawadogo M. Mol. Biol. 1999; PubMed Google Scholar). In of transcriptional regulation of dCK in important for Sp and USF1/USF2a binding to GC- and and the for a physical between these proteins that in promoter interactions between Sp and USF including Sp1, and can to effectively regulate dCK expression over a and to tissue-specific patterns of expression of critical understanding of the major determinants of dCK gene expression may lead to for by dCK activity at the transcriptional in with antiviral and antitumor in the basis for the of Down to including araC Y. Blood. 1999; Google that at the of dCK transcription in a for Y. Blood. 1999; Google as and for the and are to for the and the expression of