Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Recombinant dna technique included in method of making a...
Reexamination Certificate
2000-05-25
2002-04-02
Park, Hankyel T. (Department: 1648)
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Recombinant dna technique included in method of making a...
C435S320100, C536S023100, C530S350000, C530S389600
Reexamination Certificate
active
06365372
ABSTRACT:
BACKGROUND OF THE INVENTION
(1) Field of the Invention
The present invention generally relates to regulation of transcription factors. More specifically, the invention relates to a novel SNF2/SWI2 protein family member, SRCAP, that is capable of activating transcription directly as well as interacting with CREB binding protein (CBP) to enhance the ability of CBP to activate transcription.
(2) Description of the Related Art
Transcription factors are well known as proteins that bind to regulatory regions of genes and other proteins to modulate transcription of the genes. Examples of transcription factors relevant to this invention include CREB, c-jun, c-myb, MyoD, E2F1, YY1, TBP, TFIIB, and RNAP II. The action of these transcription factors is affected by co-activators, notably CREB binding protein (CBP).
CBP is a histone acetyltransferase capable of acetylating not only histones but also several transcription factors such as GATA-1 and p53 (Boyes et al., 1998
, Nature
396, 594-598; Hung et al., 1999
, Cell Biol
. 19, 3496-3505; Webster et al., 1999
, Mol. Cell. Biol
. 19, 3485-3495). CBP also binds to several proteins that also finction as histone acetyltransferases (P/CAF, p/CIP and the p160 co-activators such as SRC-1).
Precisely how CBP interacts with these co-activators and other cellular factors to activate transcription has not been completely elucidated. The notion that CBP interacts with a specific subset of factors at different promoters was first suggested by the work of Korzus et al., 1998
, Science
279, 703-707. These authors showed that CBP in conjunction with P/CIP, SRC-1 and P/CAF was required for activation of transcription of a RARE reporter gene by the retinoic acid receptor, whereas only CBP, P/CAF and p/CIP were needed for activation of a CRE reporter gene by CREB and only CBP and P/CAF are required for transcription of a GAS-reporter gene by STAT-1. In addition the HAT activity of each of these co-activators was not needed at each promoter. For example, with the RARE- reporter gene, despite the fact that the pCIP-P/CAF-SRC-1-CBP complex was needed for activation of transcription, only the HAT activity of P/CAF was needed, whereas transcription of the CRE-reporter by the CBP-P/CAF-pCIP complex required the HAT activity of CBP not p/CAF. Thus, the specific transcription factors which CBP binds determine not only the requirement for specific co-activators but whether their HAT activity is also needed. The requirement for a specific HAT function may also be altered depending on what signaling pathways activate transcription. Xu et al. (1998
, Nature
395, 301-306) have reported that forskolin activation of Pitl mediated transcription requires the HAT finction of CBP whereas insulin activation of Pitl mediated transcription does not.
Why different HAT activities are needed is unclear but CBP, P/CAF and p/CIP have been shown to have different substrate specificities (Perissi et al., 1999
, Proc. Natl. Acad. Sci. U.S.A
. 96, 3652-3657) suggesting they acetylate a different subset of proteins. The possibility that substrate specificity of these proteins may be regulated is illustrated by the work of Perissi et al. (Id.) who reported that binding of p/CIP to CBP changes the substrate specificity of CBP. Repression of the HAT activity of CBP by the adenoviral 12S E1A protein has been reported by several laboratories and occurs through E1A contacts with the HAT domain of CBP (Charkravati et al., 1999
, Cell
96, 393-403, reviewed in Goldman et al., 1997
, Recent Prob. Horm. Res
. 52, 103-120). In contrast to these findings, recent work by Ait-Si Ali et al. (1998
, Nature
396, 184-186) indicates that in some circumstances E
1
A can stimulate the HAT activity of CBP.
The activity of several kinases has been shown to regulate CBP function. The NGF stimulated kinase, p42/44
MAPK
, activates CBP mediated transcription by a phosphorylation event which is blocked by the MAPK inhibitor PD 98059. p42/44 MAPK directly associates with CBP and can phosphorylate CBP in vitro (Liu et al., 1999
, Neuroreport
10, 1239-1243). Studies have also demonstrated the cyclin E-cyclin dependent kinase 2 (cdk2) complex binds CBP and mediates hyperphosphorylation of CBP at the G1/S boundary (Ait-Si Ali et al., supra). Inhibition of cyclin E-Cdk2 by the cyclin dependent kinase inhibitor p21, results in the activation of NF-kB mediated transcription (Perlins et al., 1997
, Science
275, 523-527). The binding of the kinase pp90
RSK
to CBP has different effects depending on the signaling pathway. pp90
RSK
-CBP interaction is needed for insulin stimulated transcription, and blocks the ability of CBP to function as a co-activator for cAMP-mediated transcription. The activity of pp90
RSK
can be mimicked by a kinase defective mutant, indicating association of pp90
RSK
with CBP is enough to alter its finction (Nakajima et al., 1996
, Cell
86,465-474). Other studies have also demonstrated that protein kinase A and CAM kinase II and IV positively modulate the ability of specific domains within CBP to activate transcription. However, the precise mechanism by which this occurs has not been delineated (Swope et al., 1996
, J. Biol. Chem
271, 28138-28145, Liu et al., 1998
, J. Biol. Chem
. 273, 25541-25544).
CBP interacts with several general transcription factors. Swope et al. (Id.) demonstrated that TBP binds in vitro to the N-terminal end of CBP and Abrahams et al. (1993
, Oncogene
8, 1639-1647) and Dallis et al. (1997
, J. Virol
. 71, 1726-1731) have shown that TBP binds CBP in vivo. TFIIB binds to the C-terminal end of CBP and this binding is blocked by the adenoviral protein E1A (Felzien et al., 1999
, Mol. Cell. Biol
. 19,4261-4266). Deletion of the TBP binding domain from CBP prevents it from acting as a co-activator for CREB whereas deletion of the TFIIB binding site has no effect (Swope et al., Id.). CBP also binds RNA helicase A and this association appears to be required for the interaction of CBP with RNA polymerase II Nakajima et al., 1997
, Cell
90, 1107-1112).
Phosphorylation of CREB in the KID domain enhances its ability to activate transcription and is mediated by a number of kinases (CAM kinase H and IV and protein kinase A) in response to biological stimuli such as neuronal signals or increased levels of intracellular cAMP (Gonzalez et al., 1991
, Mol. Cell. Biol
. 11, 1306-1312; Fiol et al., 1994
, J. Biol. Chem
. 269, 32187-32193; Sun et al., 1994
, Genes Develop
8, 2527-2539). Specific iphosphorylation of serine 133 within CREB promotes association with CBP but studies by Sun and Maurer (1995
, J. Biol. Chem
. 270, 7041-7044) indicate this phosphorylation is not sufficient for activation of transcription. Other studies have found that phosphorylation of CREB on serines 129 and 142 modulate the ability of CREB to activate transcription (Fiol et al., supra; Sun and Maurer, supra) suggesting they change the way in which CREB and CBP interact perhaps alters the function of CBP (e.g. the HAT activity or binding of other proteins). The ability of CREB to activate transcription is also blocked by the MAPK inhibitor PD 98059 (Seternes et al., 1999
, Mol. Endocrinol
. 13, 1071-1083). Although the mechanism by which this occurs has not been completely elucidated other studies indicate that PD 98059 completely inhibits the ability of CBP to activate transcription (Liu et al., 1998
, J. Biol. Chem
. 273, 25541-25544).
Structure-finction studies by Swope et al. (1996
, J. Biol. Chem
271, 28138-28145) indicated that deletion of amino acids 272-460 from the amino terminal end of CBP prevent it from acting as a co-activator for CREB. A CBP
227-460
peptide containing this deleted region also functions as a strong transcriptional activator when expressed as a Gal chimera (Gal-CBP
227-460
). In addition, overexpression of a CBP
1-460
peptide squelches the ability of full length CBP to function as a CREB co-activator. Collectively, these studies suggest that a factor binding to the amino terminal end of CBP is essential for it to function as a co-activator for CREB. These studies also indica
Chrivia John
Yaciuk Peter
Coburn LLP Thompson
Park Hankyel T.
Saint Louis University
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