WO1999024605A2 - Adn a activite de promoteur pour gene a cycle cellulaire - Google Patents

Adn a activite de promoteur pour gene a cycle cellulaire Download PDF

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Publication number
WO1999024605A2
WO1999024605A2 PCT/DE1998/003397 DE9803397W WO9924605A2 WO 1999024605 A2 WO1999024605 A2 WO 1999024605A2 DE 9803397 W DE9803397 W DE 9803397W WO 9924605 A2 WO9924605 A2 WO 9924605A2
Authority
WO
WIPO (PCT)
Prior art keywords
dna
cell cycle
promoter activity
base pairs
base sequence
Prior art date
Application number
PCT/DE1998/003397
Other languages
German (de)
English (en)
Other versions
WO1999024605A3 (fr
Inventor
Michael Knehr
Monika Poppe
Original Assignee
Deutsches Krebsforschungszentrum Stiftung des öffentlichen Rechts
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Deutsches Krebsforschungszentrum Stiftung des öffentlichen Rechts filed Critical Deutsches Krebsforschungszentrum Stiftung des öffentlichen Rechts
Publication of WO1999024605A2 publication Critical patent/WO1999024605A2/fr
Publication of WO1999024605A3 publication Critical patent/WO1999024605A3/fr

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells

Definitions

  • the present invention relates to a DNA with promoter activity which can be linked to a cell cycle gene, a system containing such a DNA and the use of both.
  • Cell cycle is the term for the period and the processes between two cell divisions.
  • the cell cycle comprises four phases, namely phases G 1, S, G2 and M.
  • G 1 and G2 are the phases in which the cell is in the S phase for DNA synthesis and for mitosis in prepared for the M phase.
  • the cell cycle is subject to close control.
  • the transition from the G 1 to the S phase is seen as an important checkpoint.
  • the control of the cell cycle is not yet understood in detail. In particular, it is not known which factors are responsible for a disturbed control of the cell cycle, as is the case with tumors.
  • the present invention is therefore based on the object of providing a means by which the control of the cell cycle can be examined and, if necessary, interfered with.
  • the present invention thus relates to a DNA with promoter activity which can be linked to a cell cycle gene, the DNA comprising the base sequence of FIG. 1 or a sequence different therefrom by one or more base pairs.
  • the present invention is based on the knowledge of the applicant that the centromer protein C (CENP-C) known as a mitotic protein also at the transition the G 1 - is involved in the S phase.
  • CENP-C centromer protein C
  • the applicant has recognized that the maximum expression of CENP-C lies in the G 1 phase. He also identified, characterized and found that the DNA region with promoter activity of CENP-C contains important control motifs of the cell cycle in the form of DNA binding sites for transcription factors, such as the E2F family. Furthermore, the applicant has found that the DNA region is regulated by tumor suppressor proteins such as pRB and p107. This suggests that CENP-C has oncogenic properties at high expression.
  • these findings are used to provide a DNA with promoter activity which can be linked to a cell cycle gene, in particular the gene for CENP-C, the DNA being the base sequence of FIG. 1 or one of these by or comprises several base pairs of different sequences.
  • the DNA of FIG. 1 was deposited as pUC1 8/5386 with the DSMZ (German Collection of Microorganisms and Cell Cultures) on August 5, 997 under DSM 1 1 671.
  • a sequence different by one or more base pairs indicates that the DNA of FIG. 1 or a part thereof may have additions, deletions, substitutions and / or inversions of one or more base pairs.
  • Such DNA also has promoter activity. It can preferably be determined by hybridization with the DNA from FIG. 1 or a part thereof.
  • the term “hybridization” indicates hybridization under normal conditions, in particular at 20 ° C. below the melting point of the DNA used as the sample.
  • Conventional methods can be used to detect promoter activity.
  • a reporter gene e.g. a luciferase gene to which the 3 'end of the DNA to be tested for its promoter activity is ligated.
  • the DNA molecule obtained can be transfected into cells and the expression of the luciferase gene determined, whereby the promoter activity is detected.
  • a DNA according to the invention comprises the following the base pairs of the base sequence from FIG. 1:
  • a DNA according to the invention can exist as such or in a system with any other DNA.
  • the latter is preferably a DNA to be transcribed, such as a reporter gene, e.g. a luciferase gene, or a structural gene.
  • a reporter gene e.g. a luciferase gene
  • Suitable vectors for this are also known to him.
  • Maniatis, T. et al. Molecular Cloning, A Laboratory Manual (1 982), Cold Spring Harbor Laboratory.
  • a DNA according to the invention can be linked to a cell cycle gene, in particular the gene for CENP-C.
  • a DNA according to the invention thus represents a means of examining or better understanding the control of the cell cycle.
  • the substances mentioned give the possibility of intervening in a regulatory manner in the control of the cell cycle. Such offers especially when the control of the cell cycle is disturbed, as is the case with tumors.
  • the present invention represents a possibility of being able to intervene therapeutically for tumor diseases.
  • the present invention provides the possibility of expressing a structural gene in a targeted manner over time. This can be of great importance for gene therapy measures, especially in the area of tumor treatment.
  • 1 shows the base sequence of a DNA according to the invention. This comprises 772 bp promoter region, 1 8 bp exon 1, 1 140 bp intron 1, 48 bp exon 2 and 1 669 bp intron 2 of the CENP-C gene. Furthermore, DNA binding parts for the transcription factors listed in Table 1 below are given.
  • E2F Consensus E2F1 -E2F5 TTTCGCGC
  • SVSV4007 GGGCGG Spl 2 shows DNA constructs in which a DNA according to the invention is linked to a luciferase gene and the expression of the latter.
  • the DNA constructs are characterized by the indication (- / +) and the expression of the luciferase gene is indicated in%.
  • the present invention is illustrated by the example.
  • DNA constructs were produced in which a DNA according to the invention was linked to a luciferase gene.
  • the known vector pXP1 which contains the luciferase gene, was used as the vector.
  • the DNA constructs obtained were used for the transfection of NIH3T3 cells and the luciferase activity was determined in the usual way. Controls containing no DNA according to the invention showed no expression of the luciferase gene.
  • This international depository accepts the microorganism designated under I, which it received on 19 9 7 - 0 8 - 0 5 (date of first filing)
  • microorganism referred to under I was received by this international depository on (date of first deposit) and an application for conversion of this initial deposit into a deposit under the Budapest Treaty was received on (dam of receipt of the application for conversion)

Abstract

L'invention concerne un ADN à activité de promoteur, qui peut être en liaison avec un gène à cycle cellulaire, un système contenant un ADN de ce type et l'utilisation des deux.
PCT/DE1998/003397 1997-11-12 1998-11-11 Adn a activite de promoteur pour gene a cycle cellulaire WO1999024605A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DE19750172.9 1997-11-12
DE1997150172 DE19750172C1 (de) 1997-11-12 1997-11-12 DNA mit Promotor-Aktivität für Zellzyklus-Gen

Publications (2)

Publication Number Publication Date
WO1999024605A2 true WO1999024605A2 (fr) 1999-05-20
WO1999024605A3 WO1999024605A3 (fr) 1999-07-29

Family

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Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/DE1998/003397 WO1999024605A2 (fr) 1997-11-12 1998-11-11 Adn a activite de promoteur pour gene a cycle cellulaire

Country Status (2)

Country Link
DE (1) DE19750172C1 (fr)
WO (1) WO1999024605A2 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001077377A2 (fr) * 2000-04-06 2001-10-18 Epigenomics Ag Diagnostic de maladies associees a la replication de l'adn

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1993005151A1 (fr) * 1991-09-12 1993-03-18 The Johns Hopkins University Antigene cenp-c humain clone
WO1994028127A1 (fr) * 1993-05-24 1994-12-08 Imperial Cancer Research Technology Limited Traitement du cancer

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1993005151A1 (fr) * 1991-09-12 1993-03-18 The Johns Hopkins University Antigene cenp-c humain clone
WO1994028127A1 (fr) * 1993-05-24 1994-12-08 Imperial Cancer Research Technology Limited Traitement du cancer

Non-Patent Citations (6)

* Cited by examiner, † Cited by third party
Title
"CLONTECH 96/97" 1996 , CLONTECH XP002100371 PromoterFinder(TM) DNA Walking Kits siehe Seite 56 *
ALAM J ET AL: "REPORTER GENES: APPLICATION TO THE STUDY OF MAMMALIAN GENE TRANSCRIPTION" ANALYTICAL BIOCHEMISTRY, Bd. 188, Nr. 2, 1. August 1990, Seiten 245-254, XP000563914 *
KALITSIS ET AL.: "GENE STRUCTURE AND SEQUENCE ANALYSIS OF MOUSE CENTROMERE PROTEINS A AND C" GENOMICS, Bd. 47, Januar 1998, Seiten 108-114, XP002100370 *
KNEHR ET AL.: "CELLULAR EXPRESSION OF HUMAN CENTROMERE PROTEIN C DEMONSTRATES A CYCLIC BEHAVIOR WITH HIGHEST ABUNDANCE IN THE G1 PHASE" PNAS, Bd. 93, September 1996, Seiten 10234-10239, XP002100369 *
SAITOH ET AL.: "CENP-C, AN AUTOANTIGEN IN SCLERODERMA, IS A COMPONENT OF THE HUMAN INNER KINETOCHORE PLATE" CELL, Bd. 70, 1992, Seiten 115-125, XP002100368 *
WILLIAMS T M ET AL: "ADVANTAGES OF FIREFLY LUCIFERASE AS A REPORTER GENE: APPLICATION TO THE INTERLEUKIN-2 GENE PROMOTER" ANALYTICAL BIOCHEMISTRY, Bd. 176, Nr. 1, Januar 1989, Seiten 28-32, XP000601607 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001077377A2 (fr) * 2000-04-06 2001-10-18 Epigenomics Ag Diagnostic de maladies associees a la replication de l'adn
WO2001077377A3 (fr) * 2000-04-06 2002-07-11 Epigenomics Ag Diagnostic de maladies associees a la replication de l'adn

Also Published As

Publication number Publication date
DE19750172C1 (de) 1998-10-01
WO1999024605A3 (fr) 1999-07-29

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