Novel protein contact points among <scp>TP53</scp> and minichromosome maintenance complex proteins 2, 3, and 5

Schaefer-Ramadan, Stephanie; Aleksic, Jovana; Al-Thani, Nayra M; Malek, Joel A.

doi:10.1002/cam4.4805

Cited by 6 publications

(4 citation statements)

References 69 publications

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“…In 8 independent experiments, we identified 33 proteins that interacted with G245D mutp53 in UM-SCC-1 cells (Supplementary Data 1 ). Many of them, such as the p63 40 , FAM83A 41 , FUBP1 42 , TGM2 43 , SNRPN 44 , MCM5 45 , and MCM7 46 , were previously shown to interact with wtp53 and/or mutp53s, which validated the reliability of our approach. Metascape enrichment analysis 47 of purified G245D mutp53-interacting proteins showed that the gene ontology (GO) term “regulation of DNA replication initiation” involving MCM5, MCM7, and WRNIP1 was significantly enriched (Fig.…”

Section: Resultssupporting

confidence: 66%

Mutant p53 gains oncogenic functions through a chromosomal instability-induced cytosolic DNA response

Zhao,

Wang,

Gleber-Netto

et al. 2024

Nat Commun

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Inactivating TP53 mutations leads to a loss of function of p53, but can also often result in oncogenic gain-of-function (GOF) of mutant p53 (mutp53) proteins which promotes tumor development and progression. The GOF activities of TP53 mutations are well documented, but the mechanisms involved remain poorly understood. Here, we study the mutp53 interactome and find that by targeting minichromosome maintenance complex components (MCMs), GOF mutp53 predisposes cells to replication stress and chromosomal instability (CIN), leading to a tumor cell-autonomous and cyclic GMP–AMP synthase (cGAS)-stimulator of interferon genes (STING)-dependent cytosolic DNA response that activates downstream non-canonical nuclear factor kappa light chain enhancer of activated B cell (NC-NF-κB) signaling. Consequently, GOF mutp53-MCMs-CIN-cytosolic DNA-cGAS-STING-NC-NF-κB signaling promotes tumor cell metastasis and an immunosuppressive tumor microenvironment through antagonizing interferon signaling and regulating genes associated with pro-tumorigenic inflammation. Our findings have important implications for understanding not only the GOF activities of TP53 mutations but also the genome-guardian role of p53 and its inactivation during tumor development and progression.

show abstract

Section: Resultssupporting

confidence: 66%

Mutant p53 gains oncogenic functions through a chromosomal instability-induced cytosolic DNA response

Zhao,

Wang,

Gleber-Netto

et al. 2024

Nat Commun

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“…The AVA-Seq method is a powerful tool to detect protein fragment-fragment interactions (8, 25, 29) and full-length protein-protein interactions. Here, we showed a novel application combining the AVA-Seq system and Oxford Nanopore Technologies MinION platform to detect full-length protein interactomes.…”

Section: Discussionmentioning

confidence: 99%

Long-read sequencing to detect full-length protein-protein interactions

Schaefer-Ramadan,

Guan,

Ahmed

et al. 2024

Preprint

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Given the increased predictions on interactome size and demand for protein function information, methods for detecting protein-protein interactions remain a significant development area. The all-vs.-all sequencing (AVA-Seq) method utilizes a convergent fusion plasmid design to make two-hybrid technology amenable to next-generation sequencing. Here, we further innovate to take advantage of synthetic DNA technologies and Oxford Nanopore Technologies long-read sequencing improvements to allow us to determine full-length protein-protein interactions. Here, using this approach we recovered 159 protein-protein interactions from a set of 57 human proteins using multiple forms of validation. Further, when referencing a human gold standard set of interactions, eight full-length protein-protein interactions were recovered from an expected 28 interaction pairs (28.6%), a typical recovery rate for two-hybrid technologies. The AVA-Seq, in combination with the ease of synthetic DNA production and the MinION platform, offers a low-cost, high-throughput alternative for determining protein-protein interactions, which can be utilized in research labs at all stages.3Key PointsFirst application of long-read sequencing for full-length protein-protein interaction studies.The recovery rate of the AVA-Seq method using full-length proteins is on par with other leading methods.Advances in synthetic biology and sequencing technologies make full-length protein interactomes affordable and accessible.GRAPHICAL ABSTRACT

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“…It is well known that various protein-protein interaction methods do not recover all interactions. So even well-studied pathways such as the WNT pathway would benefit from analysis with new methods [19, 20]. The all-vs-all sequencing (AVA-Seq) method is a novel approach for detecting PPIs.…”

Section: Introductionmentioning

confidence: 99%

Identifying novel interactions of the colon-cancer related APC protein with Wnt-pathway nuclear transcription factors

Al-Thani

Schaefer-Ramadan

Aleksic

et al. 2022

Preprint

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Background: Colon cancer is often driven by mutations of the adenomatous polyposis coli (APC) gene, an essential tumor suppressor gene of the Wnt β-catenin signaling pathway. APC and its interactions in the cytoplasm have been well studied, however various groups have also observed its presence in the nucleus. Identifying novel interactions of APC in the Wnt pathway will provide an opportunity to better understand the nuclear role of APC and ultimately identify potential cancer treatment targets. Methods: We used the all-vs-all sequencing (AVA-Seq) method to interrogate the interactome of protein fragments spanning most of the 60 Wnt β-catenin pathway proteins. Using protein fragments identified the interacting regions between the proteins with more resolution than a full-length protein approach. Pull- down assays were used to validate a subset of these interactions. Results: 74 known and 703 novel Wnt β-catenin pathway protein-protein interactions were recovered in this study. There were 8 known and 31 novel APC protein-protein interactions. Novel interactions of APC and nuclear transcription factors TCF7, JUN, FOSL1, and SOX17 were particularly interesting and confirmed in validation assays. Conclusion: Based on our findings of novel interactions between APC and transcription factors and previous evidence of APC localizing to the nucleus, we suggest APC may compete and repress CTNNB1. This would occur through the binding of the transcription factors (JUN, FOSL1, TCF7) to regulate the Wnt signaling pathway including through enhanced marking of CTNNB1 for degradation in the nucleus by APC binding with SOX17. Additional novel Wnt β-catenin pathway protein-protein interactions from this study could lead researchers to novel drug designs for cancer.

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Novel protein contact points among TP53 and minichromosome maintenance complex proteins 2, 3, and 5

Cited by 6 publications

References 69 publications

Mutant p53 gains oncogenic functions through a chromosomal instability-induced cytosolic DNA response

Mutant p53 gains oncogenic functions through a chromosomal instability-induced cytosolic DNA response

Long-read sequencing to detect full-length protein-protein interactions

Identifying novel interactions of the colon-cancer related APC protein with Wnt-pathway nuclear transcription factors

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