Biomedical subjects

G Chinnadurai

Publications and source records attributed to G Chinnadurai.

At least 37 records · Page 2Linked to original sources

PubMed2001

The pro-apoptotic protein, Bik, exhibits potent antitumor activity that is dependent on its BH3 domain.

The Bcl-2 homology 3 (BH3) domain is present in most members of the Bcl-2 protein family and is required to confer the death-inducing properties of pro-apoptotic members, including Bax, Bak, Bad, and Bik, in cell-based assay systems. To determine whether the BH3 domain possesses a similar role in tumor tissues in vivo, we overexpressed the wild-type Bik protein and its BH3-deleted counterpart, using adenoviral technology, in chemoresistant human tumor prostate (PC-3) and colon (HT-29) cell lines growing in vitro and in vivo. Bik caused apoptosis in both PC-3 and HT-29 cells in vitro by inducing the release of cytochrome c from mitochondria to cytoplasm, resulting in the catalytic activation of caspases 9, 7, and 3 and cleavage of poly(ADP-ribose) polymerase and DNA fragmentation. When the BH3 domain was deleted from the Bik protein, no effect on mitochondrial activity or cell morphology could be observed. Furthermore, intratumoral injection of an adenovirus vector expressing the Bik gene, but not the deleted BH3 Bik gene, suppressed the growth of PC-3 and HT-29 xenografts established in nude mice. Histological examination of tumors from mice treated with the wild-type Bik adenoviral construct demonstrated cellular debris, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling positive staining, and morphological changes associated with apoptosis. In contrast, tissue sections obtained from tumors treated with the BH3-deleted Bik adenoviral construct showed no evidence of apoptosis. Thus, our results suggest that the BH3 domain is required for the antitumor activity of the Bik protein and provides a novel therapeutic approach for cancer therapy.

Adenoviridae↗

PubMed2000

Phosphorylation of the pro-apoptotic protein BIK: mapping of phosphorylation sites and effect on apoptosis.

BIK is a pro-apoptotic BCL-2 family member and is the founding member of a subfamily of pro-apoptotic proteins known as "BH3-alone" proteins. Ectopic expression of BIK induces apoptosis in variety of mammalian cells. BIK complexes with various anti-apoptotic BCL-2 family proteins such as adenovirus E1B-19K and BCL-2 via the BH3 domain. However, the heterodimerization activity of BIK alone is insufficient for its apoptotic activity. Previous studies have shown that phosphorylation regulates the functional activity of both anti-apoptotic and pro-apoptotic members of the BCL-2 family. Here, we have examined phosphorylation of BIK and its effect on the apoptotic activity of BIK. We show that BIK exists as a phosphoprotein and is phosphorylated at residues 33 (threonine) and 35 (serine). Mutation of the phosphorylation sites, in which the Thr and Ser residues were changed to alanine residues, reduced the apoptotic activity of BIK without significantly affecting its ability to heterodimerize with BCL-2. Our results suggest that phosphorylation of BIK is required for eliciting efficient apoptotic activity. Partial purification of the protein kinase from HeLa cell cytoplasmic extracts suggest that BIK may be phosphorylated by a casein kinase II-related enzyme.

Apoptosis↗

PubMed2000

Structural analysis of the human pro-apoptotic gene Bik: chromosomal localization, genomic organization and localization of promoter sequences.

The human Bik gene codes for a strong pro-apoptotic protein BIK. We have used fluorescent in-situ hybridization to establish the chromosomal localization of the Bik gene to 22q13.3. Genomic clones of the Bik gene were identified from a cosmid library of chromosome 22. Detailed analysis of the Bik gene revealed that it spans a region of about 19kb and comprises of five exons. Sequence analysis indicated that the 5' flanking region of Bik lacks canonical TATA and CAAT boxes but directs transcriptional initiation from a single site. A 1.9kb region containing the promoter elements of the Bik gene was identified and was found to direct expression of the reporter cat gene in transient transfection studies. By mutational analysis, the minimal Bik promoter was localized to a region between -211 to +153. Northern blot analysis showed a ubiquitous expression profile of the Bik mRNA with elevated levels of expression in heart and skeletal muscle.

Apoptosis↗

PubMed2000

Functional identification of the apoptosis effector BH3 domain in cellular protein BNIP1.

BCL-2 family proteins play a central role in apoptosis regulation in mammals and in C. elegans. Mammalian cellular and viral anti-apoptosis proteins such as BCL-2 and E1B-19K interact with several cellular proteins. Some of these interacting proteins promote apoptosis and belong to the BCL-2 family. Certain BCL-2 family proapoptotic proteins such as BAX and BAK share extensive sequence homology with BCL-2. In contrast, certain pro-apoptotic proteins such as BIK and BID share a single death effector domain, BH3, with other BCL-2 family proteins. By mutational analysis, we show that one of the cellular proteins, BNIP1 (previously Nip-1), that interacts with BCL-2 family anti-apoptosis proteins is a 'BH3 alone' pro-apoptotic protein. Transient transfection of BNIP1 induces a moderate level of apoptosis. Deletions of the N-terminal 32 amino acid region and the C-terminal trans-membrane domain did not significantly affect pro-apoptotic activity. In contrast, deletions encompassing a region containing a motif similar to the BH3-domain abrogated the apoptotic activity. Substitution of BNIP1 BH3 domain for the corresponding sequence in BAX efficiently restored the apoptotic activity of BAX, establishing the functional identity of the BH3 domain of BNIP1. The N-terminal deletions of BNIP1 (that retain the BH3 domain) enhanced the level of interaction with BCL-XL. Mutants containing the BH3 deletions were still able to heterodimerize with BCL-XL while mutants lacking both the N-terminal region and the BH3 domain were unable to heterodimerize, suggesting that BNIP1 may bind to BCL-XL via two different binding motifs.

Amino Acid Motifs↗

PubMed1999

Modulation of mitochondrial Ca(2+) homeostasis by Bcl-2.

We have investigated the role of mitochondrial Ca(2+) (Ca(m)) homeostasis in cell survival. Disruption of Ca(m) homeostasis via depletion of the mitochondrial Ca(2+) store was the earliest event that occurred during staurosporine-induced apoptosis in neuroblastoma cells (SH-SY5Y). The decrease of Ca(m) preceded activation of the caspase cascade and DNA fragmentation. Overexpression of the anti-apoptosis protein Bcl-2 led to increased Ca(m) load, increased mitochondrial membrane potential (DeltaPsi(m)), and inhibition of staurosporine-induced apoptosis. On the other hand, ectopic expression of the pro-apoptotic protein Bik led to decreased Ca(m) load and decreased DeltaPsi(m). Inhibition of calcium uptake into mitochondria by ruthenium red induced a dose-dependent apoptosis as determined by nuclear staining and DNA ladder assay. Similarly, reducing the Ca(m) load by lowering the extracellular calcium concentration also led to apoptosis. We suggest that the anti-apoptotic effect of Bcl-2 is related to its ability to maintain a threshold level of Ca(m) and DeltaPsi(m) while the pro-apoptotic protein Bik has the opposite effect. Furthermore, both ER and mitochondrial Ca(2+) stores are important, and the depletion of either one will result in apoptosis. Thus, our results, for the first time, provide evidence that the maintenance of Ca(m) homeostasis is essential for cell survival.

Calcium↗

PubMed1999

Binding of CtIP to the BRCT repeats of BRCA1 involved in the transcription regulation of p21 is disrupted upon DNA damage.

Mutations in BRCA1 are responsible for nearly all of the hereditary ovarian and breast cancers, and about half of those in breast cancer-only kindreds. The ability of BRCA1 to transactivate the p21 promoter can be inactivated by mutation of the conserved BRCA1 C-terminal (BRCT) repeats. To explore the mechanisms of this BRCA1 function, the BRCT repeats were used as bait in a yeast two-hybrid screen. A known protein, CtIP, a co-repressor with CtBP, was found. CtIP interacts specifically with the BRCT repeats of BRCA1, both in vitro and in vivo, and tumor-derived mutations in this region abolished these interactions. The association of BRCA1 with CtIP was also abrogated in cells treated with DNA-damaging agents including UV, gamma-irradiation, and adriamycin, a response correlated with BRCA1 phosphorylation. The transactivation of the p21 promoter by BRCA1 was diminished by expression of exogenous CtIP and CtBP. These results suggest that the binding of the BRCT repeats of BRCA1 to CtIP/CtBP is critical in mediating transcriptional regulation of p21 in response to DNA damage.

BRCA1 Protein↗

PubMed1999

BNIP3alpha: a human homolog of mitochondrial proapoptotic protein BNIP3.

Apoptosis is regulated by interaction of viral and cellular BCL-2 family antiapoptotic proteins with various pro-apoptotic proteins, several of which are also members of the BCL-2 family. Cellular protein BNIP3 is a BCL-2 family proapoptotic protein that interacts with viral antiapoptosis proteins such as adenoviruses E1B-19K and EBV-BHRF1 and cellular antiapoptosis proteins such as BCL-2 and BCL-xL. Database searches indicate that the human genome encodes an open reading frame for a protein, BNIP3alpha, that shares substantial homology with BNIP3. The BNIP3alpha open reading frame encodes a protein of 219 amino acids that contains a conserved BH3 domain and a COOH-terminal trans-membrane domain, characteristic of several BCL-2 family proapoptotic proteins. BNIP3alpha interacts with viral antiapoptosis protein E1B-19K and cellular antiapoptosis proteins BCL-2 and BCL-xL. Overexpression of BNIP3alpha in transfected cells results in apoptosis and suppresses the antiapoptosis activity of E1B-19K and BCL-xL. Like BNIP3, BNIP3alpha seems to be predominantly localized in mitochondria. These results suggest that BNIP3alpha is a structural and functional homologue of BNIP3. BNIP3 and BNIP3alpha seem to be the first examples of homologues among the various human proapoptotic proteins. Northern blot analysis reveals that BNIP3alpha is expressed ubiquitously in most human tissues. In contrast, BNIP3 is expressed well in several human tissues and less abundantly in certain tissues such as placenta and lung. These results suggest that although BNIP3 and BNIP3alpha may promote apoptosis simultaneously in most human tissues, BNIP3alpha may play a more universal role.

Adenovirus E1B Proteins↗

PubMed1998

Regulation of apoptosis by a Caenorhabditis elegans BNIP3 homolog.

We have identified a C. elegans protein, ceBNIP3, homologous to the human BCL-2/EIB-19K interacting BCL-2 family pro-apoptotic protein BNIP3. In transiently transfected mammalian cells, ceBNIP3 complexes with CED-9, the worm homolog of BCL-2. CeBNIP3 also efficiently heterodimerizes with the cell death protease proCED-3 by direct binding via the prodomain. Transfection of ceBNIP3 and CED-3 results in enhanced proteolytic processing of the CED-3 zymogen and in cooperative induction of apoptosis. Coexpression of CED-9 suppresses the cooperative cell death induced by ceBNIP3 and CED-3. In cells coexpressing CED-9, ceBNIP3 and CED-3, all three proteins exist as a ternary complex suggesting that CED-9 may suppress cooperative apoptosis induced by CED-3 and ceBNIP3 by simultaneous complex formation with CED-3 and ceBNIP3. Our results suggest that ceBNIP3 may be a novel component of the C. elegans apoptosis paradigm and may initiate apoptosis by recruiting CED-3 to mitochondria and other cytoplasmic membranes.

Amino Acid Sequence↗

PubMed1998

Structural determinants present in the C-terminal binding protein binding site of adenovirus early region 1A proteins.

The C-terminal binding protein (CtBP) has previously been shown to bind to a highly conserved six-amino acid motif very close to the C terminus of adenovirus early region 1A (Ad E1A) proteins. We have developed an enzyme-linked immunosorbent assay that has facilitated the screening of synthetic peptides identical or similar to the binding site on Ad E1A for their ability to bind CtBP and thus inhibit its interaction with Ad12 E1A. It has been shown that amino acids both C-terminal and N-terminal to the original proposed binding site contribute to the interaction of peptides with CtBP. Single amino acid substitutions across the binding site appreciably alter the Kd of the peptide for CtBP, indicative of a marked reduction in the affinity of the peptide for CtBP. The solution structures of synthetic peptides equivalent to the C termini of both Ad5 and Ad12 E1A and two substituted forms of these have been determined by proton NMR spectroscopy. Both the Ad12 and Ad5 peptides dissolved in trifluoroethanol/water mixtures were found to adopt regular secondary structural conformations seen as a series of beta-turns. An Ad12 peptide bearing a substitution that resulted in only very weak binding to CtBP (Ad12 L258G) was found to be random coil in solution. However, a second mutant (Ad12 V256K), which bound to CtBP rather more strongly (although not as well as the wild type), adopted a conformation similar to that of the wild type. We conclude that secondary structure (beta-turns) and an appropriate series of amino acid side chains are necessary for recognition by CtBP.

Adenovirus E1A Proteins↗

PubMed1998

A potent cell death activity associated with transient high level expression of BCL-2.

The BCL-2 proto-oncogene contains unusually long untranslated 5' and 3' sequences. Deletion of the sequences flanking the BCL-2 open reading frame dramatically increases the level of protein expression. Transient high level BCL-2 protein expression mediated by plasmid transfection or by infection with recombinant adenovirus results in potent apoptosis of several cell lines. Detailed mutational (deletion and add-back) analysis reveals that both 5'- and 3'-flanking sequences contribute to the negative modulation of protein expression from the BCL-2 open reading frame. It appears that these sequences exert the negative regulatory effect in an orientation-dependent manner. Analysis of BCL-2 RNA levels indicate that elevated levels of mRNA may be the primary cause of elevated levels of protein expression. Apoptosis induced by adenovirus vectors expressing elevated levels of BCL-2 can be readily inhibited by the caspase inhibitor z-VAD-fmk, suggesting that high levels of BCL-2 expression induce apoptosis via the caspase cascade. Mutational analysis of BCL-2 indicates that its pro-apoptotic activity is separable from its anti-apoptosis activity. Our results raise the possibility that oncogenic conversion of BCL-2 may require somatic mutations in the pro-apoptotic activity, in addition to other activating mutations that result in enhanced expression. Consistent with this hypothesis, a somatic mutation of BCL-2 observed in multiple human tumors results in reduced apoptosis activity.

Adenoviridae↗

PubMed1998

Functional dissection of Bfl-1, a Bcl-2 homolog: anti-apoptosis, oncogene-cooperation and cell proliferation activities.

The human Bfl-1 gene codes for a 175-amino acid BCL-2 family protein that has an anti-apoptosis activity and is overexpressed in certain human epithelial and hematopoietic malignancies. Bfl-1 efficiently suppresses apoptosis induced by the p53 tumor suppressor protein and cooperates with a dominant nuclear oncogene, E1A, in transformation of primary epithelial cells in vitro. Unlike other BCL-2 family proteins, expression of BFL-1 permits limited cell proliferation over an extended period of time when cells are induced to undergo apoptosis. We have carried out mutational analysis to dissect the various activities encoded by Bfl-1 and to determine the sequence requirements for these activities. BFL-1 shares four conserved domains, BH1, BH2, BH3 and BH4 with other BCL-2 family proteins. Mutations within BH1, BH2 and BH4 domains abolish or greatly attenuate the anti-apoptotic, oncogene cooperation and proliferation facilitating activities of BFL-1. In contrast, a mutation within the BH3 domain (which is essential for the activity of pro-apoptotic members of the BCL-2 family) does not significantly affect the BFL-1 functions. Although BFL-1 does not contain a well-defined C-terminal transmembrane domain, deletion of the C-terminal 24 amino acid region (corresponding to the transmembrane domain of other BCL-2 family proteins) partially reduces the various activities of BFL-1. All the mutants defective in the anti-apoptosis activity are also defective in the oncogene cooperation activity suggesting that these two activities may be linked. A unique feature of BFL-1 is the presence of a Gln-rich N-terminal region that overlaps with the BH4 domain. The Gln residues appear to be essential for the proliferation permitting activity of BFL-1. Since mutations of the Gln residues located within the BH4 domain appear to confer an extended cell survival activity in the absence of cell proliferation, our results suggest that BFL-1 communicates with both cell proliferation and apoptosis machineries and suggest a link between these two activities.

Amino Acid Sequence↗

PubMed1998

Adenovirus E1B-19K/BCL-2 interacting protein BNIP3 contains a BH3 domain and a mitochondrial targeting sequence.

Adenovirus E1B-19K and BCL-2 anti-apoptosis proteins interact with certain BCL-2 family pro-apoptotic proteins. A conserved domain, BH3, present in these proteins is essential for their pro-apoptotic activity and for heterodimerization with anti-apoptosis proteins. Cellular protein BNIP3 (previously NIP3) interacts with E1B-19K, BCL-2, BCL-xL, and EBV-BHRF1. BNIP3 contains a motif similar to the BH3 domain. Deletion of the BH3-like motif in BNIP3 abrogates its ability to heterodimerize with E1B-19K and BCL-xL. Substitution of the BH3 domain of BNIP3 for the corresponding sequences of BAX functionally restores the pro-apoptotic and protein heterodimerization activities of BAX. BNIP3 exhibits a delayed cell death activity that is partially relieved by deletion of the BH3 domain. BNIP3 suppresses the anti-apoptosis activity of BCL-xL in a BH3-dependent manner. BNIP3 contains a C-terminal trans-membrane (TM) domain similar to other BCL-2 family proteins and BNIP1 (previously NIP1). The TM domains of BNIP3 and BNIP1 can functionally substitute for the TM domain of a BCL-2 family member EBV-BHRF1. The BNIP3 TM domain exclusively targets the heterologous green fluorescent protein (GFP) to mitochondria. These results suggest that BNIP3 is a member of the BH3-contaning BCL-2 family of pro-apoptotic proteins and functions in mitochondria.

Adenovirus E1B Proteins↗

PubMed1998

Interaction between a cellular protein that binds to the C-terminal region of adenovirus E1A (CtBP) and a novel cellular protein is disrupted by E1A through a conserved PLDLS motif.

Adenovirus E1A proteins immortalize primary animal cells and cooperate with several other oncogenes in oncogenic transformation. These activities are primarily determined by the N-terminal half (exon 1) of E1A. Although the C-terminal half (exon 2) is also essential for some of these activities, it is dispensable for cooperative transformation with the activated T24 ras oncogene. Exon 2 negatively modulates in vitro cooperative transformation with T24 ras as well as the tumorigenic and metastatic potentials of transformed cells. A short C-terminal sequence of E1A governs the oncogenesis-restraining activity of exon 2. This region of E1A binds with a cellular phosphoprotein, CtBP, through a 5-amino acid motif, PLDLS, conserved among the E1A proteins of human adenoviruses. To understand the mechanism by which interaction between E1A and CtBP results in tumorigenesis-restraining activity, we searched for cellular proteins that complex with CtBP. Here, we report the cloning and characterization of a 125-kDa protein, CtIP, that binds with CtBP through the PLDLS motif. E1A exon 2 peptides that contain the PLDLS motif disrupt the CtBP-CtIP complex. Our results suggest that the tumorigenesis-restraining activity of E1A exon 2 may be related to the disruption of the CtBP-CtIP complex through the PLDLS motif.

Adenovirus E1A Proteins↗

PubMed1997

Functional dissection of the pro-apoptotic protein Bik. Heterodimerization with anti-apoptosis proteins is insufficient for induction of cell death.

Bik is a potent pro-apoptotic protein, which complexes with various anti-apoptotic proteins such as Bcl-2, Bcl-xL, 19-kDa adenovirus E1B, and EBV-BHRF1. The mechanism by which Bik promotes cell death is not known. It shares a conserved domain, BH3, with other pro-apoptotic proteins, Bax, Bak, Bid, and Hrk, and certain anti-apoptosis proteins such as Bcl-2 and Bcl-xL. Mutations within the BH3 domain of Bik abrogate its ability to induce cell death and to complex with anti-apoptosis proteins. This result is consistent with the hypothesis that Bik may promote cell death by complexing with and antagonizing the activity of endogenous cellular anti-apoptosis proteins such as Bcl-2 and Bcl-xL. To elucidate the relationship between protein complex formation and induction of cell death, we have identified the minimal sequences of Bik, from a library of N-terminal and C-terminal deletion mutants, required for interaction with Bcl-2 and Bcl-xL and for inducing efficient cell death. Two-hybrid analysis in yeast and immunoprecipitation analysis of proteins expressed in mammalian cells indicate that a 52-amino acid region (amino acids 43-94) of Bik, encompassing the BH3 domain, is sufficient for efficient heterodimerization with Bcl-2 and Bcl-xL. Protein interaction studies further reveal that an 18-amino acid region, encompassing the BH3 domain (residues 57-74), constitutes the core heterodimerization domain. Functional analysis indicates that a Bik deletion mutant expressing residues 43-120, which efficiently heterodimerizes with the anti-apoptosis proteins Bcl-2 and Bcl-xL, is defective in eliciting cell death. In contrast, a mutant expressing additional C-terminal sequences (amino acids 43-134) interacts with the survival proteins and elicits efficient cell death. Our results suggest that for Bik-mediated cell death, the heterodimerization activity encoded by the BH3 domain alone is insufficient and raise the possibility that Bik may induce cell death autonomous of heterodimerization with survival proteins such as Bcl-2 and Bcl-xL.

Apoptosis↗

PubMed1997

Mapping of adenovirus 5 E1A domains responsible for suppression of neu-mediated transformation via transcriptional repression of neu.

Overexpression of neu (also known as c-erbB-2 or HER-2) commonly occurs in human cancer and is also known to enchance tumor metastasis and chemoresistance. Our earlier reports showed that the adenovirus 5 E1A can suppresss the neu-mediated transformation by repression of neu. Thus, E1A has the potential to be used as a therapeutic agent against the neu-overexpressing human cancers. However, a serious concern to this approach is that E1A is also capable of immortalizing primary culture cells and can co-operate with ras or E1B oncogenes to transform them. The E1A CR2 domain (amino acid residues 120 to 140) necessary for binding to RB is believed to be required for this oncogenic function. Here, we report that deletion of CR2 region did not affect E1A's capability to repress neu. Interestingly, deletion of the amino acid residues 4 to 25 or 40 to 80 completely disrupted E1A-mediated neu repression. By deleting the amino acid residues from 81 to 185, we have successfully generated a mini-E1A mutant that was sufficient to inhibit neu promoter activity and suppress neu-mediated transformation. The mini-E1A mutant does not contain the CR2 domain that is crucial for RB binding and immortalization, and hence, may serve as a more selective tumor suppressor, and a safer therapeutic agent. It may also be a useful tool to further investigate the molecular mechanism(s) of neu overexpression and E1A-mediated transcriptional repression in cancer cells.

3T3 Cells↗

PubMed1996

bfl-1, a bcl-2 homologue, suppresses p53-induced apoptosis and exhibits potent cooperative transforming activity.

The bcl-2 family of genes code for proteins that contain anti-apoptotic or pro-apoptotic activity. The human bfl-1 gene contains an open reading frame for a 175-amino acid Bcl-2 family protein. Among the various Bcl-2 family members, the Bfl-1 protein shares the highest homology with the mouse A1 protein. These two proteins share three conserved domains, Bcl homology (BH)1, BH2, and BH3, with other Bcl-2 family proteins. Unlike other Bcl-2 family members, Bfl-1 contains a GIn-rich NH2-terminal region and lacks an NH (19K homology) domain 1. We demonstrate that the Bfl-1 protein suppresses apoptosis induced by the p53 tumor suppressor protein in a manner similar to other Bcl-2 family members such as Bcl-2, Bcl-xL and EBV-BHRF1. In addition, the bfl-I gene cooperates efficiently with the Ela oncogene in transformation of primary rodent epithelial cells. Our results suggest that the human bfl-1 gene may play an important role in carcinogenesis.

Adenovirus E1B Proteins↗

PubMed1996

A hydrophobic region within the adenovirus E1B 19 kDa protein is necessary for the transient inhibition of NF-kappaB activated by different stimuli.

The early transcribed adenovirus proteins E1A and E1B display a variety of functions in the transformation of primary rodent cells and the regulation of apoptosis and transcription. We have recently shown recently that the E1B 19 kDa protein from Adenovirus 5 (Ad 5) can functionally antagonize the stimulatory effect of E1A 13S on the human transcription factor NF-kappaB. Here we show that expression of E1B 19 kDa negatively interfered with the activation of NF-kappaB by different stimuli, such as the E1A 13S protein, and treatment with phorbol ester and tumor necrosis factor alpha. This suggests that E1B 19 kDa acts on a common upstream signaling event. Band shift experiments showed that expression of E1B 19 kDa impaired the generation of the nuclear, DNA-binding form of NF-kappaB. Domain mapping experiments employing various E1B 19 kDa mutants revealed the necessity of a hydrophobic Bcl-2 homology region between amino acids 90 and 96 for NF-kappaB inhibition. Co-transfection experiments showed that the inhibitory effect of E1B 19 kDa on E1A 13S-activated NF-kappaB transcription was gradually lost in the course of time. Thus the continuous stimulatory action of E1A 13S can finally override the antagonistic effects of E1B 19 kDa on NF-kappaB activity. In contrast to E1B 19 kDa, expression of the E1B 55 kDa protein did not result in a de novo activation of NF-kappaB, but co-stimulated the transcriptional potential of activated NF-kappaB.

Adenovirus E1B Proteins↗

PubMed1996

The CtBP binding domain in the adenovirus E1A protein controls CR1-dependent transactivation.

The adenovirus E1A-243R protein has the ability to force a resting cell into uncontrolled proliferation by modulating the activity of key targets in cell cycle control. Most of these regulatory mechanisms are dependent on activities mapping to conserved region 1 (CR1) and the non-conserved N-terminal region of E1A. We have previously shown that CR1 functions as a very patent transactivator when it is tethered to a promoter through a heterologous DNA binding domain. However, artificial DNA binding was not sufficient to convert full-length E1A-243R to a transactivator. Thus, an additional function(s) of the E1A-243R protein modulates the effect of CR1 in transcription regulation. Here we demonstrate that a 44 amino acid region at the extreme C-terminus of ElA inhibited transactivation by a Gal4-CR1 fusion protein. Inhibition correlated with binding of the nuclear 48 kDa C-terminal binding protein (CtBP), which has been implicated in E1A-mediated suppression of the metastazing potential of tumour cells. This might suggest that CtBP binding can regulate E1A-mediated transformation by modulating CR1-dependent control of transcription.

Adenovirus E1A Proteins↗