Introduction To Ad5 E1A

Rationale for using E1A as a research tool

An understanding of the mechanisms of growth and development demands a detailed understanding of gene regulation and control as this comprises the basis from which these complex phenomena arise. Spontaneous aberrations in the central processes of growth and development can result in the formation of cells whose growth is unrestrained. Left unchecked it is from these cells that tumours are formed. The products of the Adenovirus 5 early region 1A (E1A) oncogene perform a large number of gene and growth regulatory activities, making E1A a useful tool with which to study important cellular processes.

The E1A transcript and proteins

Within one hour of infection, transcription from Adenovirus type 5 or the highly related Adenovirus type 2 becomes detectable. The first viral gene to be transcribed is early region 1A (E1A) (101). The primary E1A transcript is processed by differential splicing to yield five distinct messages with sedimentation coefficients of 13S, 12S 11S, 10S, and 9S (126,138)(Fig. 1A). The 13S and 12S mRNAs are the most abundant at early times during infection, while the 9S mRNA is the most abundant at late times. The 11S and 10S mRNA are minor species that become more abundant at late times after infection (126,138). The 13S, 12S, 11S, 10S and 9S E1A mRNA code for 289 residue (R), 243R, 217R, 171R and 55R proteins respectively (Fig.1A), all of which are detectable in vivo with the exception of the 9S product which has only been detected in vitro (126). The E1A proteins, particularly the major ones of 289R and 243R, regulate transcription of both viral and cellular genes in the infected cell.

Figure 1: Structure of the E1A mRNA transcripts and postions of the conserved regions in the E1A proteins

A: Coding regions are represented as boxes and are all read in the same frame except for the second exon of the 9S mRNA (blue box)

B: Regions of conserved amino acid sequence in the E1A proteins are represented as CR1, CR2 and CR3.

Physical and structural characteristics of the E1A proteins

The 289R and 243R proteins that are the major products encoded by the E1A region of Ad5 share identical amino and carboxyl sequences. The only difference between them is the presence of an additional 46 amino acids in the larger protein that arise as the result of differential splicing of the primary E1A transcript (Fig.1A). This extra internal stretch is often referred to as the "unique region".

A comparison of E1A sequences of various human and simian adenovirus serotypes has identified three regions of conserved amino acid homology (72,142). In Ad5, conserved region 1 (CR1) maps between amino acid 40-80, CR2 between amino acids 121-139, and CR3 between residues 140-188 which roughly coincides with the 13S unique region (Fig. 1B). Evolutionary conservation of these particular sequences suggests that they are critical for E1A function, but by no means limits the possibility that other regions are at least equally important.

The E1A proteins are proline rich, acidic, and localized in the nucleus. Rapid nuclear localization is mediated by a highly basic pentapeptide signal sequence (Lys-Arg-Pro-Arg-Pro) at the extreme carboxyl terminus of the polypeptides (84). The conformational constraints imposed by the high proline content likely limit the formation of substantial secondary structure in the E1A proteins. The extreme heat stability of bacterially produced E1A protein, which retains significant transcriptional activation activity even after boiling for five minutes (75), suggests that either E1A can readily refold to an active conformation, or that E1A can function as a random coil. The surprising ability of E1A protein to tolerate large deletions and insertions without total disruption of its biological activities, has led to the concept that E1A is a series of small modular domains that are relatively independent of surrounding sequences.

A potential metal binding domain with a consensus zinc finger motif (Cys-Xaa2-Cys-Xaa13-Cys-Xaa2-Cys) has been identified within the unique region of the E1A 289R protein. The larger E1A protein does bind a single zinc ion, and as expected the smaller E1A protein that lacks this region does not. This structure appears essential for transcriptional activation by E1A as substitution of glycine for any of the four cysteines within this motif not only abolishes the ability of E1A to bind Zn2+ but also results in a loss of transactivation activity (23). It is not known whether this metal binding structure mediates an interaction between E1A and DNA or with other proteins.

Post-translational modification of the E1A products is limited to phosphorylation that occurs at serine residues 89, 132, 219, and possibly 96 and 231 (30,31,134). The available evidence suggests that phosphorylation does little to regulate E1A activity (30,133).

Multiple activities of E1A

The E1A proteins exhibit a wide range of biological activities that include but are not limited to: transcriptional activation, transcriptional repression, induction of DNA synthesis, mitosis and apoptosis in quiescent rodent cells, suppression of differentiation, immortalization of primary rodent cells and transformation in cooperation with the viral E1B products or activated ras (6).

Surprisingly, E1A can also function as an anti-oncogene to suppress transformation, metastasis and tumorigenicity. In addition, E1A can promote differentiation, partially reversing the transformed phenotype of cells, induce apoptosis and susceptibility to tumour necrosis factor and enhance susceptibility to the host cellular immune response (22,44,96).

E1A is also known to associate specifically with a number of cellular proteins. A large amount of work using E1A mutants has established which regions of E1A are necessary for these activities (see below)

The following sections provide brief introduction to a number of the E1A functions listed above. Numerous reviews cover each of these activities in far greater detail.

Association of E1A with cellular proteins

As the E1A proteins do not possess an intrinsic enzymatic or specific DNA binding activity, it is generally assumed that they exert their effects indirectly by affecting the activities of cellular regulatory proteins. Immunoprecipitation of E1A proteins from extracts of [35S]-methionine labelled infected cells shows that E1A complexes specifically with a number of cellular proteins. The major species of coprecipitating proteins migrate at rates corresponding to 300 kDa, 107 kDa, and 105 kDa, with minor species migrating at 130 kDa, 60 kDa and 33 kDa (54,159)(Fig. 2). It is now know that E1A interacts with the CREB binding protein (CBP) (2,83) and a related protein p300 (34), pRB (36,155) and related proteins p130 (53,78,87) and p107 (40), p60/cyclin A (105), p33 cdk2 (136), BS69 (55), CtBP (113), the TATA binding protein (48,56,121,122) and various other components of the TFIID complex (15,21,47,88).

Figure 2: Depiction of an idealized fluorograph of the proteins co-immunoprecipitated with E1A from lysates of radiolabelled Ad5 infected cells


The regions of E1A required for association with most of these proteins have been mapped in a variety of cells using a multitude of mutants (Fig. 3B).

It seems likely that E1A functions indirectly through its association with these and other as yet unidentified cellular proteins. Our current assumption is that E1A binds to these proteins and either sequesters them, preventing their action, or modifies their activities.

Figure 3: Map of the major E1A proteins and the regions required for selected E1A activities.

A) The 289 and 243 residue E1A proteins.
B) Consensus sites for binding to cellular proteins. The hatched regions are of secondary importance.

C-H: Regions required for:
C) Transformation with activated ras and induction of mitosis in BRK cells.
D) Repression of gene expression. Generally the hatched regions are necessary, but for repression of some genes either of the other two regions are required as well.
E) Activation of gene expression. The different shadings represent regions required for different pathways.
F) Suppression of differentiation. Depending on the cells, the minimum requirements are the hatched regions; either the hatched or the stippled region; or all three.
G) Induction of DNA synthesis, apoptosis and TNF sensitivity
H) Suppression of SWI/SNF function in Saccharomyces cerevisiae.

Immortalization of primary rodent cells and oncogenic transformation in combination with a second oncogene

Unlike infection of human cells, infection of rodent cells with human adenovirus does not result in efficient virus production and for this reason, rodent cells are referred to as semi-permissive for infection. Infections of semi-permissive primary rodent cells can lead to oncogenic transformation (135). Complete transformation requires only the gene products encoded by the E1A and E1B regions (89,141). E1A also can cooperate with activated ras or polyoma middle T antigen to morphologically transform primary baby rat kidney cells (112). Mutant adenoviruses that express only the 298R or the 243R E1A protein can cooperate with ras to transform cells, but both products appear necessary for complete transformation (92). Mutants retaining only the N-terminal 139 residues still transform with ras (144,156) and others have shown that a N-terminal region, and two regions that correspond to CR1 and CR2 are essential (67,76,80,81,94,114,127,144,156)(Fig. 3C). These regions are identical to those necessary for association with the cellular proteins p300 and pRb, suggesting roles for these two proteins in transformation (36,37,155,156).

The E1A proteins alone are sufficient to immortalize primary rodent epithelial cells allowing them to proliferate indefinitely (62,94). These cells will not grow to high saturation densities, and do not exhibit the morphological characteristics of fully transformed cells unless E1A is expressed at very high levels (115). Mutational analysis has shown that regions in the N-terminal half of E1A, similar to those required for transformation, are also necessary for immortalization of primary BRK cells (106,114,127). Immortalization also requires a region near the C-terminus of E1A (106,128). Since the C-terminal half of E1A is not required for transformation with ras (144,156) or E1B (5), this suggests that whatever function resides in the C-terminus is provided by the cooperating oncogene.

Transactivation of viral and cellular genes

E1A can activate transcription from a wide variety of cellular and viral genes in trans. During a lytic infection, E1A increases transcription of the other viral early genes about 50-100 fold (9,68). As the E1A proteins do not bind to DNA with any sequence specificity (17,41), they must trans-activate by binding to factors that regulate transcription. The gene promoters that respond to E1A have little in common, suggesting that the E1A proteins must either affect wide range of transcription factors or they effect several different trans-activating pathways. Surprisingly, both of these strategies appear to be employed by E1A. The results of extensive investigation have shown that E1A contains a minimum of four independent transactivation functions (Fig. 3E). The unique region and regions in exons 1 and 2 can each independently induce gene expression.

A: Induction of transcription by the unique region (CR3)

Of the trans-activation pathways through which E1A acts, that involving CR3 is the most potent: most of the activation of viral early genes and of exogenous cellular genes by E1A is brought about by this region (8,116). This region alone is sufficient to activate transcription as microinjection of a synthetic 49 residue peptide representing CR3 into HeLa cells activates expression (81).

CR3 consists of two domains (49,150). At its C-terminal end, residues 183-188 locate the E1A protein on a promoter by interacting with a promoter-bound transcription factor, in this way recruiting E1A to a promoter (103). Once recruited to a promoter, the activation domain in the N-terminal part of CR3, residues 141-178, containing a zinc finger (23,85,150), binds to TBP and activates transcription by stimulating the formation of a multiprotein transcription initiation complex with RNA polymerase II (15,48,61). Evidence suggests that the region of CR3 that binds TBP binds another cellular factor as well (15,48). Furthermore, trans-activation of some promoters requires one of two auxiliary elements in exon 2 besides CR3 (11).

B: Induction of transcription by exon 1

It seems likely that alterations in gene expression induced by E1A would play an important role in the changing cell growth. Surprisingly, the unique region is not necessary for transformation or for most of the other cellular effects of E1A, whereas exon 1 is (Fig. 3).

A number of pathways have now been identified by which E1A exon 1 can induce gene expression (52,69,74,97,140,147). Of these, the most studied is the action of E1A in effectively freeing the E2F family of transcription factors from association with pRb, p107 and p130 (see Fig. 4). By freeing E2F, E1A activates transcription from a variety of genes containing E2F-binding sites in their promoters, including the adenovirus E2 early promoter (4,19,73,109), c-myc (60,131) and dihydrofolate reductase (59). Similar increases have been reported for the transforming oncogenes of other DNA tumour viruses that bind pRb, including HPV 16 E7 protein (104,108), and possibly the immediate early protein of human cytomegalovirus (146). In addition, HPV E7 protein and SV40 large T antigen disrupt complexes of E2F with pRb and cyclin A in vitro (18,102). Interestingly, E2F is not required for transcription of viral genes in SV40 and HPV, as it is in adenovirus. Instead, the release of E2F by these viral proteins appears to be related to the presence of E2F sites in the promoters of several S-phase specific cellular genes, activation of which probably facilitates viral growth (65).

Figure 4: Effect of E1A on pRb/E2F function

In the G0/G1 phase of the cell cycle, hypo-phosphorylated pRb complexes with transcription factors of the E2F family (indicated as E2F and DP) preventing their ability to activate transcription. Cell cycle dependent phosphorylation of pRb by cyclin/cdk complexes releases E2F to activate transcription of target genes required for S-phase of the cell cycle. Expression of E1a overrides the normal cellular control of the pRb-E2F interaction by binding hypo-phosphorylated pRb and freeing E2F.

In primary BRK cells, which are regularly used for studies of transformation by E1A, the 243R protein activates expression of a range of viral early and late genes that lacked E2F binding sites in their promoters (97). Expression of viral late genes requires E1A to bind to p300. On the other hand, viral early genes and the cellular gene for PCNA are activated through at least two independent, redundant pathways, requiring binding to p300 and to pRb, respectively, as only E1A mutants that fail to bind to p300 and pRb simultaneously are defective (71,97). These results show that E1A can activate expression by binding to pRb through some mechanism that does not involve E2F. Besides activating gene expression directly, the p300/CBP pathway may also be able to activate the pRb/E2F pathway indirectly. E1A induces synthesis of p34cdc2 kinase in BRK cells through redundant p300/CBP and pRb pathways (27,147), indicating that binding to pRb is not essential for induction. Increased synthesis of p34cdc2 leads to increased phosphorylation of pRb (147). As only hypophosphorylated pRb complexes with and inactivates E2F (19), hyperphosphorylation likely results in the release of E2F and activation of E2F dependent transcription.

E1A also activates gene expression through the AP-1 family of transcription factors. These proteins and those of the similar ATF/CREB family contain a basic DNA-binding domain, a so-called leucine zipper, and an activation domain. Through their leucine zippers, the proteins form dimers either with themselves or with other members of the AP-1 and ATF families, and by binding to specific promoter sites, activate transcription. The sites to which the dimers bind are promoter elements that confer response to the phorbol ester TPA or to cyclic AMP (cAMP). The promoter of the gene c-jun binds a heterodimer of its own product, c-Jun, with ATF-2 (139), and E1A is able to stimulate transcription of this gene (139,140), apparently by causing hyperphosphorylation of the trans-activating domain of c-Jun (52). This action of E1A depends on CR1 but not on CR2 (140), which suggests that binding to p300/CBP is involved (52). In cooperation with cAMP, E1A also induces expression of c-fos (95), which is another member of the AP-1 family. This induction requires E1A to bind to p300 (46). In in vitro experiments, E1A, acting either on its own (24) or in cooperation with cAMP (46), increases binding of AP-1 to a TPA-responsive element; in the latter case, the increase requires E1A bind to p300/CBP (46). The influence of E1A on AP-1 activity provides a mechanism by which E1A may interact with signal transduction pathways in the cell.

C: Induction of transcription by exon 2

A mutant virus that produces only exon 2 of E1A activates expression of viral genes in both rat and human cells, suggesting that yet another transactivation function resides within exon 2 (97). So far the only protein identified as associating with exon 2 is CtBP (113) (Fig. 3B).

Transcriptional repression of viral and cellular genes

In contrast to transcriptional activation, E1A proteins can also repress transcription from the Ad2 E2 late promoter (111), the SV40 and polyoma virus early promoters (12,143), and the promoters of a number of differentiation specific genes (38,58,120,125,149). Repression by E1A can be titrated out by introduction of excess copies of the affected enhancer, suggesting that repression results from the presence of a negatively acting repressor product that is activated by E1A (12).

Mutational analysis of E1A has mapped the domains of E1A required for enhancer repression primarily to a N-terminal region including CR1 (5,67,110,124,144)(Fig. 3D). This region correlates well with the region of E1A required for binding to p300/CBP (67,124), suggesting that transrepression is mediated through the association of E1A with p300/CBP. Several studies have also suggested a role for CR2 in transrepression (80,81,114), but this is not usually observed. As p300 and CBP act as transcriptional co-activators for a number of transcription factors (66), binding by E1A may sequester them making them unavailable for transcriptional activation. p300/CBP associates with another cellular protein referred to as p/CAF that is a histone acetylase and E1A can disrupt this interaction (158). Recruitment of p/CAF to a promoter by p300/CBP could result in promoter specific histone acetylation that helps overcome the repressive effects of assembly of genes as chromatin. Interestingly, in Saccharomyces cerevisiae , an N-terminal region of E1A suppresses SWI/SNF activation of gene expression (90), suggesting that E1A can repress transcription by blocking chromatin remodelling.(Fig. 3H)

E1A mutants that fail to repress also fail to transform (67,124), suggesting a requirement for transcriptional repression in oncogenic transformation. Although others have reported mutants that are defective for transrepression that still transform (5,76,144),at least some of these mutants were later found to repress transcription from a different enhancer (5,124), suggesting a dependence on the promoter.

Effects on cellular differentiation

As differentiation and proliferation are mutually exclusive processes, it is not surprising that an immortalizing oncogene such as E1A can block differentiation. Indeed, E1A can suppress the neuron-like differentiation of rat PC12 pheo-chromocytoma cells, differentiation in the PC Cl 3 rat epithelial thyroid cell line, muscle differentiation in rat and mouse myoblasts and reverse retinoic acid/cAMP induced differentiation of mouse F9 teratocarcinoma cells. In each of these cases, E1A suppresses morphological changes and differentiation specific gene expression (10,14,57,70,86,98,149,151,152,160). Indeed, the regions of E1A required for suppression of differentiation are the same as those required for repression of gene expression (Fig. 3F).

In contrast to the cell lines described above, in undifferentiated mouse F9 teratocarcinoma cells E1A induces changes in morphology and gene expression characteristic of differentiation (91,100,145). Similar effects have been observed upon introduction of E1A into P19 embryonal carcinoma cells (118). Thus, in embryonal and teratocarcinoma cells, E1A can act as a tumour-suppressor to induce differentiation characteristics. Mapping studies indicate that induction of differentiation of P19 cells by E1A requires the N-terminal 25 residues and CR1 region and this correlates with binding to p300 (118). Whether this same region is required for induction of differentiation in F9 cells has not been determined.

It has been noted that introduction of E1A into a number of human tumour cell lines induces a conversion to a less refractile "flat" morphology, a reorganization of the cytoskeleton, a restoration of contact inhibition and a reduction in anchorage independent growth indicative of a less transformed, hence more highly differentiated state (42). It was later shown that expression of the 243R E1A protein induces the expression of epithelial characteristics in a diverse array of human tumour cells regardless of parental cell type (43). This surprising result has led to speculation that the epithelial cell phenotype may be a "default" that requires only "ubiquitous" transcription factors. Thus, the mechanism by which E1A stimulates conversion to the epithelial phenotype may result from its ability to inhibit the expression or activity of cell type specific transcription factors (43). If this were indeed the case, the regions of E1A involved in suppression of differentiation would likely correlate with those involved in induction of the epithelial phenotype, but this has yet to be determined.

Induction of DNA synthesis and mitosis in rodent cells

Infection with human adenovirus induces growth arrested rodent cells to enter the cell cycle and begin cellular DNA replication. Microinjection of a plasmid containing the E1A gene induces cellular DNA synthesis in quiescent mouse 3T3 cells, demonstrating that E1A alone is necessary and sufficient for this process (123). Although small deletions in E1A have no effect on the ability of E1A to induce DNA synthesis, more extensive deletions of at least 25 residues in the region of CR1 eliminate induction (93,119). To induce quiescent BRK cells to move from G0/G1 into S phase of the cell cycle and to replicate their DNA, the 243R E1A protein requires either residues 2-25 and 36-69 or residues 121-127 but not necessarily all three (Fig. 3G) (64). As only E1A mutants that fail to bind p300/CBP and pRb together are defective, this can be interpreted to mean that it is sufficient for the E1A proteins to bind either p300/CBP or pRb to induce DNA synthesis (64,147,148). Besides inducing entry into S phase, E1A can also force quiescent embryonic or baby rat cell cultures to traverse the cell cycle (7,93,161). For E1A to induce cells to complete the cell cycle and undergo mitosis, all three regions of the E1A protein, residues 2-25, 36-69 and 121-127, are required simultaneously, as they are for transformation (Fig. 3C), suggesting that here too, the E1A protein must bind to p300/CBP, p107 and pRb (63).

Induction of apoptosis and sensitivity to TNF

Besides playing important roles in regulating viral gene expression and inducing cell growth, E1A also displays several cytotoxic activities. In human cells, E1A stimulates the degradation of both cellular and viral DNA in the infected cell (deg phenotype) and results in enhanced cytopathic effect (cyt phenotype) resembling programmed cell death or apoptosis (107,154). Apoptosis is an active process by which the cell directs its own destruction. Apoptosis provides an important means of regulating cell number and suppression of this process likely contributes to oncogenesis (79,157). In the absence of the viral E1B products, either of the two major E1A proteins can induce apoptosis in a wide variety of cell types (6,153). The 243R E1A protein can also sensitize cells to "anoikis", a form of apoptosis induced by disruption of epithelial cell-matrix interactions (45). Induction of apoptosis in rodent cells by the 243R E1A protein requires wt p53 (25,82). The 289R E1A protein can induce apoptosis in a p53 independent process, but this requires other as yet unidentified viral proteins that are likely expressed in response to transactivation by the unique region (129,130). In this case, the 289R protein induces the processing and activation of CPP32, a Ced-3/ICE protease that cleaves and inactivates poly-ADP ribose polymerase, leading to apoptosis (13). Other than the unique region, it is not known which regions of the 289R protein are necessary to induce apoptosis in a p53 independent fashion (130).

The regions of the 243R protein required for induction of apoptosis are the same as those required for inducing quiescent cells to enter and traverse the cell cycle (99) (Fig. 3G). Surprisingly, the 243R protein induces apoptosis only in cells prevented from proliferating by cell-cell contact or by serum deprivation (99,107), suggesting that induction of apoptosis is an unintended consequence of a proliferation block opposing induction of cell division by E1A (99). A similar conclusion has been made for the induction of apoptosis by inappropriate expression of c-myc protein (39). To overcome the tendency of E1A to induce apoptosis, the viral E1B encoded 19 kDa and 55 kDa products suppress apoptosis (107) and this may explain the co-operativity between E1A and E1B in cell transformation.

In addition to inducing apoptosis, E1A can also sensitize cells to killing by tumour necrosis factor (TNF) (20,28). TNF was originally identified through its anti-tumour activity (16) and can induce both necrotic and apoptotic forms of cell lysis in transformed cells (77). Detailed analysis of the regions of E1A responsible for induction of TNF sensitivity shows an exact correlation with those regions required for induction of DNA synthesis (117) (Fig. 3G). These results are largely consistent with earlier less comprehensive studies (1,29,137). In infected cells, the viral E3 encoded 10.4 kDa, 14.5 kDa and 14.7 kDa proteins suppress TNF mediated cytotoxicity induced by E1A (50,51).

Conclusions: Functions of E1A in infection

E1A could not have evolved to transform cells, and so transformation must result from infection gone awry. What purpose then does E1A serve by forcing quiescent cells to enter and traverse the cell cycle? It has been suggested that although the virus encodes its own replication machinery, maximal virus production depends upon obtaining appropriate substrates from the host cell (132). The cells normally infected by adenovirus in the animal host are likely to be Go arrested or terminally differentiated, and as such contain low levels of these substrates. Thus it would likely be advantageous for the virus if the host cell entered the cell cycle, as the levels of these substrates would increase allowing more efficient virus replication. E1A appears to fulfil this function, apparently by interacting with important regulators of cell growth such as p300/CBP and pRb (63,64). We now know that this strategy of acting through cellular regulators of growth is not unique to E1A but is shared by the products of other viral oncogenes such as SV40 T antigen or the human papilloma virus E7 protein, which can also bind pRb (26,33), p130 (53), p107 (32) and p300/CBP (3,35). Normally, in a permissive infection of human cells the remainder of the viral life cycle would ensue, leading to host cell death by the passive process of attrition, and the release of viral progeny.

In a semi-permissive infection of rodent cells with human adenovirus, E1A appears to perform its proper functions in preparing the host cell for maximal viral replication. The E1A proteins bind to cellular regulatory proteins such as pRb and p300, alter gene expression and force the infected cell to enter the cell cycle. In many of these cells the life cycle of human adenovirus is not properly completed, allowing some cells to survive. As the result of programming the infected cell to commence division, E1A apparently eliminates the option for these cells to undergo differentiation, the opposite process to cell division. The extended proliferative capacity induced by E1A in these cells probably allows the accumulation of other genetic aberrations, leading to the transformed phenotype. This argument is firmly supported by the requirement for identical regions of E1A for both transformation, and entry and passage through the cell cycle.

Interestingly, in the absence of E1B, the presence of E1A induces programmed cell death or apoptosis in otherwise healthy cells. Induction of apoptosis appears to require exactly the same regions of E1A necessary for induction of DNA synthesis, suggesting that the same signals produced by E1A that induce cell division can trigger suicide (99). This undesirable side effect would hamper efficient virus production. The virus appears to have compensated by evolving the E1B 19 kDa protein, which can inhibit apoptosis by an as yet unknown mechanism. It has been speculated that this ability to block apoptosis may be one reason why E1B 19 kDa can cooperate with E1A to transform cells (107). This is a definite possibility as the product of the proto-oncogene Bcl-2, which is known to block apoptosis, but is otherwise distinct from the E1B 19 kDa protein in structure and location, can also cooperate with E1A to transform primary BRK cells (107). This suggests that other oncogenes, such as activated ras, that can cooperate with E1A to transform cells might function to block apoptosis as well.

Thus, E1A affects each of the three processes, division, differentiation and death, that a hypothetical cell sitting in the Go phase of the cell cycle could choose (or be forced) into. Significantly, the ability of E1A to do this depends on two key domains that are required for binding to pRb and p300/CBP. These studies demonstrate that the interaction and likely inactivation of pRb and p300/CBP by E1A also appears central to the ability of E1A to reprogram gene expression (6), strengthening the argument that it is the changes in gene expression induced by E1A that ultimately result in the effects E1A has on cell growth and development.

Without question, far more must be learned about how the interactions of E1A with cellular factors alter gene expression before we understand in detail how E1A alters cell growth and leads to transformation. Nevertheless the recognition that E1A acts to regulate gene expression through multiple distinct mechanisms and the apparent connection of some of these with transformation is significant.


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Page Contributors:
  • Joe Mymryk Ph.D.
  • Stan Bayley Ph.D.