Student or educator? Sign up with your school email and get 6 months of Pro free.
CoChat
Sign Up
Cited by 14Cancer-related Molecular PathwaysProtein Structure and DynamicsHeat shock proteins research

The DOI matches its Crossref record; 82 of the 89 references it cites were found in a registry or an index; none of the 54 sources its claims cite is checked for support yet.

Evolutionary history of the p53 family DNA-binding domain: insights from an Alvinella pompejana homolog

Open access
Zhang, Qiang, Balourdas, Dimitrios-Ilias, Baron, Bruno, Senitzki, Alon, Haran, Tali E., Wiman, Klas G., Soussi, Thierry, Joerger, Andreas C.·2022·Cell Death & Disease
DOI 10.1038/s41419-022-04653-8PMC PMC8901663
Open this paper in CoChat — free accountOpen PDF

Abstract

The extremophile Alvinella pompejana, an annelid worm living on the edge of hydrothermal vents in the Pacific Ocean, is an excellent model system for studying factors that govern protein stability. Low intrinsic stability is a crucial factor for the susceptibility of the transcription factor p53 to inactivating mutations in human cancer. Understanding its molecular basis may facilitate the design of novel therapeutic strategies targeting mutant p53. By analyzing expressed sequence tag (EST) data, we discovered a p53 family gene in A. pompejana. Protein crystallography and biophysical studies showed that it has a p53/p63-like DNA-binding domain (DBD) that is more thermostable than all vertebrate p53 DBDs tested so far, but not as stable as that of human p63. We also identified features associated with its increased thermostability. In addition, the A. pompejana homolog shares DNA-binding properties with human p53 family DBDs, despite its evolutionary distance, consistent with a potential role in maintaining genome integrity. Through extensive structural and phylogenetic analyses, we could further trace key evolutionary events that shaped the structure, stability, and function of the p53 family DBD over time, leading to a potent but vulnerable tumor suppressor in humans.

Overview

CoChat Summary

TL;DR

The authors identified and characterized a p53 family DNA-binding domain (DBD) from the extremophile annelid *Alvinella pompejana*, showing it is more thermostable than all vertebrate p53 DBDs tested but less stable than human p63. Structural and phylogenetic analyses trace key evolutionary changes in p53 family DBD stability, hydrophobic core packing, and DNA-binding cooperativity.

Key findings

  • A p53 family gene was found in *A. pompejana* EST data; its DBD shares 47% identity with human p53, 58% with human p63, 56% with human p73, and >80% with the *Capitella teleta* homolog.
  • The crystal structure of the *A. pompejana* DBD was determined at 1.92 Å; it has an immunoglobulin-like β-sandwich, conserved zinc-binding residues (Cys176, His179, Cys238, Cys242), and conserved DNA-contact residues (Arg248, Arg273, Arg280), with a more recessed L1 loop than human p53.
  • By DSC, the *A. pompejana* DBD had an apparent Tm of 56.2 °C, higher than human p53 (43.6 °C), *C. teleta* p53 homolog (39.1 °C), *Xenopus* p53 (38.2 °C), chicken p53 (54.0 °C), and rainbow trout p53 (37.5 °C), but lower than human p63 (60.4 °C).
  • EMSA with the Con1 site showed the *A. pompejana* DBD binds with the same band-shift pattern as human p53, likely as a tetramer but with lower affinity; p63 DBDs migrated as dimers and formed the most temperature-stable complexes.
  • Evolutionary analysis identified p63/p73-like stabilizing features, including a 255/270 hydrophobic-core packing swap (F270V compensated by I255F), an aromatic residue at the human N239Y position, absence of the human p53 cysteine cluster (Cys124 replaced by tryptophan), and absence of the vertebrate p53 Glu180–Arg181 double salt bridge.

Methods

  • Design: combined phylogenetic, structural, biophysical, and DNA-binding analyses of p53 family DBDs from vertebrates and invertebrates.
  • Data/sample: DBDs from human, *Xenopus*, chicken, and rainbow trout p53; human and *Xenopus* p63; and *A. pompejana* and *C. teleta* p53 homologs.
  • Main analysis: BLAST/EST and genome searches; MUSCLE/Jalview alignments; recombinant protein expression in *E. coli*; X-ray crystallography at 1.92 Å; differential scanning calorimetry (DSC), differential scanning fluorimetry (DSF), circular dichroism (CD); and electrophoretic mobility shift assays (EMSA) with Con1 DNA.

Limitations

  • Stated: the *A. pompejana* genome is not yet available, the full-length protein sequence is unknown, no MDM2-like transcript was detected in the EST data, and the exact temperatures experienced by the worm are uncertain.
  • Stated: sequencing the genome is needed to clarify whether an MDM2-like degradation pathway exists and to enable more in-depth functional studies.
  • Evident: DNA-binding conclusions rely on qualitative EMSA rather than quantitative affinities or in vivo function, and structural conclusions are based on the isolated DBD rather than the full-length protein.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1038/s41419-022-04653-8

Cites

82 of 82 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Figures

Fig. 1 Simplified evolutionary tree of p53 family proteins. Vertebrates typically have three p53 family members, p53, p63, and p73, as a result of two gene duplications at the beginning of vertebrate evolution. Multiple copies as a result of independent gene duplications are also found in several invertebrate lineages. The invertebrate p53 family genes in this tree are classified solely based on the presence or absence of a SAM domain in the predicted proteins (Ensembl Metazoa release 52 and Ensembl Protists release 52), which is a characteristic feature of vertebrate p63 and p73 proteins, but absent in vertebrate p53. The genome of the Pompeii worm (Alvinella pompejana), a member of the annelid worms, has not yet been published. Comparison with the published genome of another polychaete annelid, Capitella teleta, suggests the presence of a single p53 family gene in this organism that contains an extended p63/p73-like tetramerization domain with a second helix [77] and a SAM domain. From the analysis of the chicken genome, only p63 and p73 proteins are predicted, but a p53 transcript has been verified experimentally [78]. Branch lengths in this tree do not reflect evolutionary distance. The figure has been adapted from Joerger and Fersht [1].
Fig. 1 Simplified evolutionary tree of p53 family proteins. Vertebrates typically have three p53 family members, p53, p63, and p73, as a result of two gene duplications at the beginning of vertebrate evolution. Multiple copies as a result of independent gene duplications are also found in several invertebrate lineages. The invertebrate p53 family genes in this tree are classified solely based on the presence or absence of a SAM domain in the predicted proteins (Ensembl Metazoa release 52 and Ensembl Protists release 52), which is a characteristic feature of vertebrate p63 and p73 proteins, but absent in vertebrate p53. The genome of the Pompeii worm (Alvinella pompejana), a member of the annelid worms, has not yet been published. Comparison with the published genome of another polychaete annelid, Capitella teleta, suggests the presence of a single p53 family gene in this organism that contains an extended p63/p73-like tetramerization domain with a second helix [77] and a SAM domain. From the analysis of the chicken genome, only p63 and p73 proteins are predicted, but a p53 transcript has been verified experimentally [78]. Branch lengths in this tree do not reflect evolutionary distance. The figure has been adapted from Joerger and Fersht [1].CC BY · PubMed Central
Fig. 2 Sequence alignment of p53 family DNA-binding domains. The partial sequence of the Alvinella pompejana p53 homolog, including the full DBD, was aligned with the sequences of human p53, p63, and p73, and the p53 homolog from Capitella teleta using MUSCLE [68] and visualized with Jalview [69]. Amino acid residues are colored according to the Clustal X color scheme based on sequence conservation and similarity. The secondary-structure assignment above the alignment refers to the structure of the human p53 DBD (2XWR) [79]. Strictly conserved DNA-contact and zinc-binding residues as well as p53 cancer mutation hotspot sites are highlighted. UniProt accession codes: human p53, P04637; human p63, Q9H3D4; human p73, O15350; chicken p53 (Gallus gallus), P10360; rainbow trout p53 (Oncorhynchus mykiss), P25035; Xenopus laevis p53, Q7T1D0; C. teleta p53 homolog, R7UHV7. The numbering of the A. pompejana p53 homolog is based on EST N72937.
Fig. 2 Sequence alignment of p53 family DNA-binding domains. The partial sequence of the Alvinella pompejana p53 homolog, including the full DBD, was aligned with the sequences of human p53, p63, and p73, and the p53 homolog from Capitella teleta using MUSCLE [68] and visualized with Jalview [69]. Amino acid residues are colored according to the Clustal X color scheme based on sequence conservation and similarity. The secondary-structure assignment above the alignment refers to the structure of the human p53 DBD (2XWR) [79]. Strictly conserved DNA-contact and zinc-binding residues as well as p53 cancer mutation hotspot sites are highlighted. UniProt accession codes: human p53, P04637; human p63, Q9H3D4; human p73, O15350; chicken p53 (Gallus gallus), P10360; rainbow trout p53 (Oncorhynchus mykiss), P25035; Xenopus laevis p53, Q7T1D0; C. teleta p53 homolog, R7UHV7. The numbering of the A. pompejana p53 homolog is based on EST N72937.CC BY · PubMed Central
Fig. 3 Crystal structure of the Alvinella pompejana p53 DBD. A Cartoon representation of the overall structure of the A. pompejana DBD (green; chain A) superimposed onto the structure of human p53 DBD in its unbound state (gray; PDB code 2XWR) [79]. Residue numbers given refer to the human protein. B Conservation of zinc coordination and the Arg249-mediated polar interaction network in the L3 loop region of the A. pompejana DBD. Stabilization of the hairpin conformation of the L3 loop is key for positioning the DNA contact residue Arg248 for docking to the minor groove of DNA response elements. The oncogenic R249S mutation destabilizes the human protein and impairs DNA binding [80]. The structure of the A. pompejana DBD is shown in green superimposed onto the structure of human p53 DBD in gray.
Fig. 3 Crystal structure of the Alvinella pompejana p53 DBD. A Cartoon representation of the overall structure of the A. pompejana DBD (green; chain A) superimposed onto the structure of human p53 DBD in its unbound state (gray; PDB code 2XWR) [79]. Residue numbers given refer to the human protein. B Conservation of zinc coordination and the Arg249-mediated polar interaction network in the L3 loop region of the A. pompejana DBD. Stabilization of the hairpin conformation of the L3 loop is key for positioning the DNA contact residue Arg248 for docking to the minor groove of DNA response elements. The oncogenic R249S mutation destabilizes the human protein and impairs DNA binding [80]. The structure of the A. pompejana DBD is shown in green superimposed onto the structure of human p53 DBD in gray.CC BY · PubMed Central
Fig. 4 Thermostability of different p53 family DBDs. Raw data of the differential scanning calorimetry (DSC) analysis for the DBDs of A. pompejana p53 homolog, C. teleta p53 homolog, human p63, Xenopus laevis p63, human p53, and Xenopus laevis p53, recorded at a heating rate of 300 °C/h. Tm values measured at a much lower heating rate of 60 °C/h were on average about 2–3 °C lower with both DSC and DSF assays. It has been shown previously that p53 family proteins denature irreversibly and that the measured apparent Tm depends on the heating rate but approximates the true Tm if the heating rate is fast enough [29, 81].
Fig. 4 Thermostability of different p53 family DBDs. Raw data of the differential scanning calorimetry (DSC) analysis for the DBDs of A. pompejana p53 homolog, C. teleta p53 homolog, human p63, Xenopus laevis p63, human p53, and Xenopus laevis p53, recorded at a heating rate of 300 °C/h. Tm values measured at a much lower heating rate of 60 °C/h were on average about 2–3 °C lower with both DSC and DSF assays. It has been shown previously that p53 family proteins denature irreversibly and that the measured apparent Tm depends on the heating rate but approximates the true Tm if the heating rate is fast enough [29, 81].CC BY · PubMed Central
Fig. 5 Stabilizing interactions in the DBD of the Alvinella pompejana p53 homolog. A, B Hydrophobic core of the A. pompejana DBD. Superposition of the A. pompejana DBD (green) onto the structure of human p53 DBD (gray; PDB code 2XWR) [79] in (A) and the structure of human p63 DBD (PDB code 3QYN; yellow) [82] in (B) shows key differences in the hydrophobic core of the A. pompejana protein. Most notably, a phenylalanine and a neighboring smaller hydrophobic amino acid have swapped places when comparing the A. pompejana and the human DBDs (residues 255 and 270 in human p53). A non-saturated hydrogen bond found in the hydrophobic core of the human protein (orange dotted line) is replaced by equivalent hydrophobic residues seen in the human p63/p73 structure. C Superposition of the A. pompejana p53 DBD onto the structure of the superstable quadruple mutant M133L/V203A/N239Y/N268D of human p53 (PDB code 1UOL; gray) [55] shows a phenylalanine in the A. pompejana structure at the position of the stabilizing N239Y substitution in human p53 next to the zinc-binding site. D The same superposition as in panel C but focusing on the location of the cysteine cluster in the human protein shows that this cysteine cluster is absent in the A. pompejana DBD. Cys124 in the human p53 DBD is substituted by tryptophan in the A. pompejana DBD.
Fig. 5 Stabilizing interactions in the DBD of the Alvinella pompejana p53 homolog. A, B Hydrophobic core of the A. pompejana DBD. Superposition of the A. pompejana DBD (green) onto the structure of human p53 DBD (gray; PDB code 2XWR) [79] in (A) and the structure of human p63 DBD (PDB code 3QYN; yellow) [82] in (B) shows key differences in the hydrophobic core of the A. pompejana protein. Most notably, a phenylalanine and a neighboring smaller hydrophobic amino acid have swapped places when comparing the A. pompejana and the human DBDs (residues 255 and 270 in human p53). A non-saturated hydrogen bond found in the hydrophobic core of the human protein (orange dotted line) is replaced by equivalent hydrophobic residues seen in the human p63/p73 structure. C Superposition of the A. pompejana p53 DBD onto the structure of the superstable quadruple mutant M133L/V203A/N239Y/N268D of human p53 (PDB code 1UOL; gray) [55] shows a phenylalanine in the A. pompejana structure at the position of the stabilizing N239Y substitution in human p53 next to the zinc-binding site. D The same superposition as in panel C but focusing on the location of the cysteine cluster in the human protein shows that this cysteine cluster is absent in the A. pompejana DBD. Cys124 in the human p53 DBD is substituted by tryptophan in the A. pompejana DBD.CC BY · PubMed Central
Fig. 6 Reshaping of the hydrophobic core of p53 family proteins during evolution. A sequence alignment of vertebrate and invertebrate p53 family DBDs shows a switch of the packing pattern of the DBD hydrophobic core via residues 255 and 270. The aromatic swap illustrated in Fig. 5A appears to have occurred at the transition from invertebrates to vertebrates. Also highlighted is an unsaturated hydrogen-bond pair in the hydrophobic core of vertebrate p53 DBDs (residues 236 and 253, human p53 numbering). UniProt accession numbers are given after the name of each species. The numbering of the A. pompejana p53 homolog is based on the translation of EST N72937.
Fig. 6 Reshaping of the hydrophobic core of p53 family proteins during evolution. A sequence alignment of vertebrate and invertebrate p53 family DBDs shows a switch of the packing pattern of the DBD hydrophobic core via residues 255 and 270. The aromatic swap illustrated in Fig. 5A appears to have occurred at the transition from invertebrates to vertebrates. Also highlighted is an unsaturated hydrogen-bond pair in the hydrophobic core of vertebrate p53 DBDs (residues 236 and 253, human p53 numbering). UniProt accession numbers are given after the name of each species. The numbering of the A. pompejana p53 homolog is based on the translation of EST N72937.CC BY · PubMed Central
Fig. 7 Conservation of DNA-binding properties between Alvinella pompejana and human p53 family DBDs, measured by electrophoretic mobility shift assay. p53 family DBDs were incubated with Con1 DNA for 1 h at 4 °C (A) and 20 °C (B) at a nominal protein concentration of 200, 800, and 2400 nM, and then run on a 6% polyacrylamide gel (37.5:1 acrylamide: bisacrylamide ratio) at the same temperature as the incubation. The DNA-binding deficient p53 cancer mutant R273H was used as a negative control.
Fig. 7 Conservation of DNA-binding properties between Alvinella pompejana and human p53 family DBDs, measured by electrophoretic mobility shift assay. p53 family DBDs were incubated with Con1 DNA for 1 h at 4 °C (A) and 20 °C (B) at a nominal protein concentration of 200, 800, and 2400 nM, and then run on a 6% polyacrylamide gel (37.5:1 acrylamide: bisacrylamide ratio) at the same temperature as the incubation. The DNA-binding deficient p53 cancer mutant R273H was used as a negative control.CC BY · PubMed Central

Verification

Claims the paper makes

0 Valid · 0 Not valid · 34 Not checked · 57 Own results and interpretations

50 more claims

Create a free account to see all 91 claims and to verify the 54 references still open.

See all claims — free account
  • It may state two separate claims as one. 32
  • Probably not a substantial claim. 8

Create a free account to see these claims.

Create a free account to see these assumptions.

Citations

Cites

Show all 89 references

Cited by

See all 14

Related

See all

Metadata

Authors
Zhang, Q., Balourdas, D.-I., Baron, B., Senitzki, A., Haran, T. E., Wiman, K. G., Soussi, T., & Joerger, A. C.
Affiliations
  • Karolinska Institutet (SE)
  • Goethe University Frankfurt (DE)
  • Institut Pasteur (FR)
  • Technion – Israel Institute of Technology (IL)
  • Uppsala University (SE)
Corresponding author
  • Haran, Tali E.
  • Wiman, Klas G.
  • Soussi, Thierry
  • Joerger, Andreas C.
ORCID
Venue
Cell Death & Disease
Publisher
Springer Science and Business Media LLC
Published
2022-03-07
Volume
13
Issue
3
Pages
214-214
ISSN
2041-4889 (ISSN-L)
DOI
10.1038/s41419-022-04653-8
PMC
PMC8901663
OpenAlex
W4220757956
Semantic Scholar
bf5f595770d49249744fbf1e35d0155149c37e25
Cited by
14
References
89
Citations by year
2025: 3 · 2024: 4 · 2023: 6 · 2022: 1
FWCI
1.55
Type
journal article
Language
en
Open access
Licence
Locations
  • Cell Death and Disease (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • PubMed Central (publishedVersion, cc-by)
  • HAL (submittedVersion, cc-by)
  • DORA PSI (Paul Scherrer Institute) (submittedVersion, other-oa)
  • KTH Publication Database DiVA (KTH Royal Institute of Technology) (submittedVersion, other-oa)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • Publication Server of Goethe University Frankfurt am Main (Goethe University Frankfurt) (submittedVersion, cc-by)
Funders
  • Israel Science Foundation (1517/14)
  • Cancerfonden
  • Deutsche Forschungsgemeinschaft (JO 1473/1-3)
Topics
  • Health Sciences › Medicine › Oncology › Cancer-related Molecular Pathways
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Heat shock proteins research
Keywords
p53 family, DNA-binding domain, Alvinella pompejana, extremophiles, hydrothermal vents, protein thermal stability, p63, protein stability, genome stability, tumor suppressor, expressed sequence tags, phylogenetic analysis, crystal structure
MeSH terms
Protein Domains, Animals, DNA, Phylogeny, Polychaeta, Tumor Suppressor Protein p53

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

A language model drew the claims from the paper on 2026-11-15, under version 3 of the claim definition (model deepseek-v4.1-flash, prompt extract-v3).

A second model labelled each claim on 2026-11-15 (model jev-1.13-20260917, definition revision gold-guideline-v0.2).

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review

How to cite

Cite CoChat for the checks on this page, and the paper for its own findings.

This page
CoChat. (2026). Citation check: Evolutionary history of the p53 family DNA-binding domain: insights from an Alvinella pompejana homolog. https://app.cochat.ai/papers/10.1038/s41419-022-04653-8
The paper
Zhang, Q., Balourdas, D.-I., Baron, B., Senitzki, A., Haran, T. E., Wiman, K. G., Soussi, T., & Joerger, A. C. (2022). Evolutionary history of the p53 family DNA-binding domain: insights from an Alvinella pompejana homolog. Cell Death & Disease. https://doi.org/10.1038/s41419-022-04653-8

Data sources

Bibliographic data: Semantic Scholar, OpenAlex, Crossref.

Contains information from the Semantic Scholar Academic Graph Datasets, made available under the ODC Attribution License (ODC-BY 1.0).

Semantic Scholar’s data is described in: Kinney, R., et al. (2023). The Semantic Scholar Open Data Platform. arXiv:2301.10140.