FC0083
Adenovirus early region 1A (E1A) oncoprotein  -  CBP TAZ2 domain, Retinoblastoma (Rb) protein

Biological function
E1A deregulates the cell cycle through interactions with key cellular proteins, including the general transcriptional coactivators cyclic-AMP response element binding (CREB) binding protein (CBP) and p300 and the retinoblastoma protein (pRb), which lead to epigenetic modifications and reprogramming of host cell transcriptional processes. Binding of E1A to pRb displaces the E2F transcription factors, thereby relieving E2F repression and resulting in premature S-phase entry and transcriptional activation of E2F-regulated genes. Interactions between E1A and CBP/p300 are central to its ability to transform cells and induce mitosis. E1A represses CBP/ p300-dependent transcriptional activity by binding to promoter-bound CBP/p300. E1A inhibits p53-mediated activation of p21 and other stress response genes and thereby suppresses cell cycle arrest and apoptosis. E1A has been variously reported to inhibit and to activate the histone acetyltransferase (HAT) activity of CBP/p300.

Domain organization/sequence features
E1A is a multifunctional protein composed of 4 conserved regions (CR1–CR4). E1A uses both CR1 and CR2 for interaction with pRb. CR2 contains a characteristic pRb recognition motif, LXCXE, which binds with high affinity to the B cyclin fold domain of the pRb pocket region.

Structural evidence
A model of the pRb:E1A:TAZ2 ternary complex was constructed from structures of the pRb pocket domain in complex with residues 37–49 from the E1A CR1 region and with the LXCXE motif from human papilloma virus (HPV) E7, together with the NMR structure. Residues 37–49 of E1A bind in a deep groove at the interface of the A and B cyclin fold domains of pRb, while residues 53–91 in the C-terminal half of CR1 interact with the TAZ2 domain of CBP/p300. Residues 92–120 of E1A are invisible between the TAZ2 binding site in CR1 and the LXCXE motif (residues 122–126) in CR2. The CR1–CR2 linker must be long enough to allow simultaneous binding of CR1 and CR2 in their cognate sites on pRb and to accommodate binding of CBP/p300 without creating steric clashes. CR1 and CR2 are connected by a long (~30 residues) intrinsically disordered linker in all sequenced E1A serotypes.

Biochemical evidence
The relative independence of the two E1A binding sites is represented by several lines of data. Either site alone is able to induce G₁ gene expression and DNA synthesis in primary rodent cells. Nevertheless, unless both E1A active sites are present, these cell cycle effects are incomplete and the stimulated cells do not go on to divide successfully. For example E1A transforming activity is about 10-fold more efficient when the two active sites are present on the same molecule. Conservation of a spacer region within E1A correlates with the requirement for simultaneous occupation of both binding sites. A non-conserved stretch of about 35 residues that is not required for efficient binding at either of the two protein binding sites in the E1A products. The spacer region is indeed required for stable simultaneous binding of pRB- related proteins and p300. E1A lacking the 86-120 region binds to both target proteins, but not readily in the same complexes, such that the pRB- related proteins are not brought together stably with p300. Furthermore, immortalization activity of E1A depends not on passive sequestering of these cellular targets but on actively promoting an association between them. smFRET shows that E1A–CBP–pRb interactions have either positive or negative cooperativity, depending on the available E1A interaction sites.

Structure/Mechanism
Conservation of a spacer region between the two binding sites that is required for simultaneous binding and efficient induction of proliferation supports the concept that the E1A protein structure has evolved to facilitate simultaneous binding. These results indicate that the E1A proteins are designed not merely to sequester these cellular products, but also to bring them into proximal association with each other in biologically significant complexes.

Mechanism category
tethering

Significance
The fuzzy linker enables allosteric coupling between the two binding regions thus permitting a context-specific tuning of the associated downstream signaling outputs.

Medical relevance
Association of E1A with CBP/p300 results in global hypoacetylation of K18 of histone H3 and may be linked to the ability of E1A to induce oncogenic transformation.

Further reading
23783631, 7494304

Submitted by
Ashok Deniz   deniz@scripps.edu