Tankyrase homolog protein (THP), nucleic acids, and methods...

Details Tankyrase homolog protein (THP), nucleic acids, and methods... Tankyrase homolog protein (THP), nucleic acids, and methods...

1999-07-09

2002-09-24

Prouty, Rebecca E. (Department: 1652)

Chemistry: molecular biology and microbiology

Enzyme , proenzyme; compositions thereof; process for...

Transferase other than ribonuclease

C435S183000, C435S193000, C435S252300, C435S252330, C435S254110, C435S254200, C435S255100, C435S255200, C435S320100, C435S325000, C435S348000, C435S254800, C435S357000, C435S358000, C435S364000, C435S367000, C435S369000, C435S021000, C530S350000, C530S358000, C536S023100, C536S023200, C536S023500

Reexamination Certificate

active

06455290

ABSTRACT:

FIELD OF THE INVENTION
The present invention is directed, in part, to nucleic acid molecules encoding tankyrase homolog protein (THP), novel THP polypeptides, and assays for screening compounds which bind to THP and/or modulate the activity of THP.
BACKGROUND OF THE INVENTION
Telomeres, the physical ends of eukaryotic chromosomes, consist of short repeated nucleotide sequences that are characteristic for all telomeric DNA, from yeast to human (see, e.g., Harley et al.,
Curr. Opin. Genet. Dev
., 1995, 5, 249). Telomeres are essential for the integrity of chromosomes (see, e.g., Lundblad et al.,
Cell
, 1996, 87, 369), since DNA polymerases are incapable of replicating the very end of DNA. This replication defect affects the telomeres which are progressively shortened with each cell cycle, and as they reach a critical short length, cells will die (see, e.g. Goldstein,
Science
, 1990, 249, 1129). In some cells, telomere length is preserved by a ribonucleoprotein reverse transcriptase, referred to as “telomerase,” which adds the characteristic telomere repeats (Greider et al.,
Cell
, 1985, 43, 405). Telomerase is not expressed in most somatic cells and, as a consequence, these cells have a finite life-span. Extensive studies in a broad range of tumors indicate that 85-95% of all tumors cells express telomerase (see, e.g., Shay et al.,
Eur. J Cancer
, 1997, 33, 787). Telomerase expression is an early event in tumor progression and is often present at the premalignant stage. Telomerase expression alone is not sufficient for tumorogenesis, but it is likely to be an essential permissive event to allow the tumor to continue to grow.
In addition to telomerase, another protein has been identified as capable of regulating telomere length, referred to as telomere repeat binding factor 1 (TRF1), that binds as a dimer to double stranded telomeric repeats (Chong et al.,
Science
, 1995, 270, 1663). Overexpression of TRF1 in a telomerase-expressing cell line leads to progressive telomere shortening, whereas inhibition of TRF1 increases telomere length (van Steensell et al.,
Nature
, 1997, 385, 740). TRF1 does not control the expression of telomerase itself but, rather, is thought to act by inhibiting telomerase action at the telomere termini (Smith et al.,
Science
, 1998, 282, 1484).
The ability of TRF1 to bind to telomeric DNA can be inhibited by its modification through the action of an enzyme called tankyrase, which covalently modifies TRF1 by ADP-ribosylation (Smith et al.,
Science
, 1998, 282, 1484). Tankyrase is a multidomain protein characterized by the presence of four distinct domains and are listed here in order from the polypeptide NH
2
-terminus to the carboxy-terminus; HPS, ANK, SAM, and PARP. The HPS domain is a region containing homopolymeric runs of His, Pro, and Ser and has unknown function. The ANK domain contains 24 repeat units of a 33-amino acid ANK motif, as found in ankyrins. This domain is responsible for the binding of tankyrase to TRF1. The SAM domain (sterile alpha module) has unknown function but is believed to contribute to protein-protein interactions. The C-terminal catalytic active region is homologous to the active region of human Poly(ADP-Ribose) polymerase (PARP) enzymes. PARP catalyzes the formation of poly(ADP-ribose) onto a suitable protein receptor using NAD
+
as substrate. The present invention involves the surprising discovery of a novel polypeptide, herein designated THP, that exhibits at least one functional homology to human tankyrase, and its role as a key component, for example, in regulating telomerase activity and thereby telomere length. THP is, thus, useful in the search for novel anti-tumor agents that can modify and/or control telomere length and/or telomere maintenance. These and other aspects of the invention are described below.
SUMMARY OF THE INVENTION
The present invention is directed to, in part, isolated nucleic acid molecules comprising SEQ ID NO:3, or a fragment thereof; SEQ ID NO:4, or a fragment thereof; a nucleotide sequence complementary to at least a portion of SEQ ID NO:3 or SEQ ID NO:4; a nucleotide sequence homologous to SEQ ID NO:3 or SEQ ID NO:4, or a fragment thereof; a nucleotide sequence that encodes a polypeptide comprising SEQ ID NO:5, or a fragment thereof; or a nucleotide sequence that encodes a polypeptide comprising an amino acid sequence homologous to SEQ ID NO:5, or a fragment thereof.
The present invention is also directed to recombinant expression vectors comprising any of the nucleic acid molecules described above.
The present invention is also directed to host cells transformed with a recombinant expression vector comprising any of the nucleic acid molecules described above.
The present invention is also directed to methods of producing a polypeptide comprising SEQ ID NO:5, or a homolog or fragment thereof, by introducing a recombinant expression vector comprising any of the nucleic acid molecules described above into a compatible host cell, growing the host cell under conditions for expression of the polypeptide, and recovering the polypeptide from the host cell.
The present invention is also directed to compositions comprising any of the nucleic acid molecules described above and an acceptable carrier or diluent.
The present invention is also directed to isolated polypeptides encoded by any of the nucleic acid molecules described above.
The present invention is also directed to compositions comprising a polypeptide encoded by any of the nucleic acid molecules described above and an acceptable carrier or diluent.
The present invention is also directed to isolated antibodies which bind to an epitope on a polypeptide encoded by any of the nucleic acid molecules described above.
The present invention is also directed to kits comprising antibodies which bind to a polypeptide encoded by any of the nucleic acid molecules described above and a negative control antibody.
The present invention is also directed to methods of inducing an immune response in a mammal against a polypeptide encoded by any of the nucleic acid molecules described above by administering to the mammal an amount of the polypeptide sufficient to induce the immune response.
The present invention is also directed to methods of identifying a compound which binds to tankyrase homolog protein (THP) by contacting THP with a compound, and determining whether the compound binds THP.
The present invention is also directed to methods of identifying a compound which binds a nucleic acid molecule encoding tankyrase homolog protein (THP) by contacting THP with a compound, and determining whether the compound binds the nucleic acid molecule.
The present invention is also directed to methods of identifying a compound which modulates the activity of tankyrase homolog protein (THP) by contacting THP with a compound, and determining whether THP activity is modified.
The present invention is also directed to compounds which modulate THP activity identified by contacting THP with the compound, and determining whether the compound modifies activity of THP, binds to THP, or binds to a nucleic acid molecule encoding THP.
These and other aspects of the invention are described in greater detail below.
DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS
The present invention provides, inter alia, isolated and purified polynucleotides that encode THP or a portion thereof, vectors containing these polynucleotides, host cells transformed with these vectors, processes of making THP, methods of using the above polynucleotides and vectors, isolated and purified THP, and methods of screening compounds which modulate THP activity.
Various definitions are made throughout this document. Most words have the meaning that would be attributed to those words by one skilled in the art. Words specifically defined either below or elsewhere in this document have the meaning provided in the context of the present invention as a whole and as are typically understood by those skilled in the art.
As used herein, the term “activity” refers to a variety of measurable indicia suggesting o

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