p53 Y220C: Effects on Structure and Cell Growth

Mutations in the TP53 gene are among the most common genetic alterations in human cancers, often leading to loss of tumor suppressor function. One such mutation, Y220C, occurs within the DNA-binding domain of p53 and destabilizes its structure, impairing normal activity. Understanding this mutation provides insight into how altered p53 function contributes to cancer progression.

Investigating Y220C’s effects on protein structure and cell growth regulation highlights its role in disrupting cellular processes.

The Y220C Mutation In The DNA-Binding Domain

The Y220C mutation in TP53 results from a nucleotide substitution that replaces tyrosine with cysteine at codon 220 within p53’s DNA-binding domain. This domain enables p53 to regulate gene expression in response to cellular stress. Y220C weakens protein stability, making it prone to unfolding and aggregation. Unlike mutations that completely abolish DNA binding, Y220C retains partial function but has reduced affinity for target sequences, impairing tumor suppression.

This mutation introduces a surface cavity near the DNA-binding interface, exposing hydrophobic residues that are normally buried, increasing susceptibility to misfolding. X-ray crystallography and molecular dynamics simulations show local unfolding, particularly in loop regions critical for DNA interaction. The mutant protein tends to adopt an inactive conformation, reducing its ability to regulate cell cycle arrest and apoptosis.

Unlike mutations that entirely disrupt function, Y220C presents a unique therapeutic opportunity. Small molecules designed to stabilize the mutant protein by filling the surface cavity have shown promise in restoring p53 activity. Compounds like PK7088 bind to the destabilized region, enhancing structural integrity and allowing partial recovery of tumor-suppressive properties. These findings highlight the potential for targeted therapies aimed at rescuing structurally compromised p53 variants.

Structural Consequences For Protein Conformation

Y220C alters p53’s structure by introducing a surface cavity in its DNA-binding domain. Tyrosine’s bulky aromatic side chain helps maintain stability, whereas cysteine’s smaller, flexible thiol-containing side chain fails to provide the same support. As a result, the surrounding region destabilizes, creating an energetically unfavorable pocket that disrupts the native fold.

X-ray crystallography and molecular dynamics simulations reveal that Y220C increases local flexibility, weakening the domain’s ability to maintain proper conformation. Exposed hydrophobic residues exacerbate instability, promoting aberrant interactions and aggregation. Unlike mutations that directly disrupt DNA contact, Y220C primarily affects thermodynamic stability, making the protein more prone to degradation and misfolding.

This destabilization extends beyond the Y220 residue, affecting other functionally important regions. Nuclear magnetic resonance (NMR) spectroscopy shows subtle conformational shifts in adjacent loop structures essential for high-affinity DNA binding. Increased fluctuations in the L2 and L3 loops reduce the mutant protein’s ability to adopt the precise geometry needed for transcriptional activation. These structural disturbances significantly impair function, as the protein struggles to maintain an active conformation.

Changes In Transactivation Functions

The Y220C mutation weakens p53’s ability to regulate gene expression, reducing its effectiveness in controlling cell cycle progression and apoptosis. Although the mutant retains partial DNA-binding capacity, its instability lowers transcriptional activation efficiency. Growth suppression genes like CDKN1A (p21) exhibit significantly reduced expression in Y220C-expressing cells, weakening p53’s ability to induce cell cycle arrest in response to stress.

This impairment is not uniform across all p53-regulated genes. Chromatin immunoprecipitation sequencing (ChIP-seq) studies show that Y220C selectively dysregulates transcription, with genes like BAX and PUMA, critical for apoptosis, showing diminished activation. This imbalance favors survival pathways, allowing tumor progression. Even when DNA damage is detected, cells evade apoptosis, increasing genomic instability.

Y220C also interferes with coactivator recruitment, further diminishing transactivation efficiency. Normally, p53 interacts with transcriptional coactivators like p300 and CBP, which facilitate chromatin remodeling and enhance gene expression. The structural instability introduced by Y220C weakens these interactions, leading to inefficient transcriptional initiation. Under stress, cells with Y220C often fail to mount an adequate response, reducing their ability to halt proliferation or trigger apoptosis.

Influence On Cell Growth Regulation

The Y220C mutation disrupts p53’s role in controlling cell proliferation, leading to unchecked growth that contributes to tumor development. Normally, p53 induces growth arrest or apoptosis in response to stress. The instability introduced by Y220C weakens this regulatory control, allowing cells to bypass critical cell cycle checkpoints. This defect is particularly evident in the G1 phase, where impaired activation of inhibitors like p21 allows unchecked transition to S phase, facilitating mutation accumulation.

Beyond cell cycle control, Y220C-expressing cells show elevated activity in mitogenic pathways like PI3K/AKT and MAPK, which promote survival and proliferation despite genomic instability. This increased signaling makes Y220C-expressing tumors particularly aggressive, as they evade growth suppression while gaining a proliferative advantage. The inability of p53 to restrain these pathways enhances metabolic activity, angiogenesis, and resistance to senescence, fostering tumor expansion.

Interactions With Other Regulatory Proteins

Y220C not only destabilizes p53 but also alters interactions with key regulatory proteins that influence its function. These disrupted interactions contribute to tumor progression by weakening p53’s ability to regulate cell fate.

One major consequence is altered binding with MDM2, p53’s principal negative regulator. Normally, MDM2 binds p53 and promotes its degradation via the ubiquitin-proteasome pathway. The Y220C mutation enhances MDM2-mediated degradation, as the destabilized protein is more susceptible to ubiquitination. This lowers p53 levels, further impairing its transcriptional function. Additionally, mutant p53 proteins can accumulate in cancer cells due to disrupted degradation, leading to dominant-negative effects that interfere with any remaining wild-type p53. This accumulation can also sequester essential co-regulators, altering the cellular transcriptional landscape.

Beyond MDM2, Y220C disrupts interactions with transcriptional co-factors like p300/CBP, which are necessary for chromatin remodeling and gene activation. Normally, these coactivators acetylate p53, enhancing DNA-binding affinity and promoting expression of growth suppression and apoptosis genes. The structural instability of Y220C-mutant p53 weakens its ability to recruit these coactivators, leading to reduced transcriptional responses. Additionally, mutant p53 proteins can acquire novel interactions that contribute to oncogenic signaling, further exacerbating the loss of tumor-suppressive functions.