Nano‐LC based proteomic approach identifies that E6AP interacts with ENO1 and targets it for degradation in breast cancer cells

Mishra, Mukul; Sharma, Akshay; Thacker, Gatha; Trivedi, Arun Kumar

doi:10.1002/iub.2132

Cited by 6 publications

(3 citation statements)

References 27 publications

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“…Up-regulated ENO1can promote various types of human cancer, such as bladder cancer cells, oral squamous cell carcinoma, and breast cancer, by regulating β-catenin [12][13][14][15][16][17][18][19][30][31][32][33]. However, the mechanism of ENO1 regulating proliferation and cell cycle in the HCC remains unknown.Recent studies have shown that ENO1 interacts with a diverse range of proteins and plays a key role in regulating the occurrence and progression of various diseases [29,34,35]. However, there is no report on ENO1-interacting proteins regulating the proliferation and cell cycle of HCC.…”

Section: Discussionmentioning

confidence: 99%

Exploring the Role of ENO1 in Regulating Cell Proliferation and Cell Cycle in Hepatocellular Carcinoma: Insights from Proteomic Analysis

Ma,

Yang,

Luo

et al. 2024

Preprint

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Background and Aims: Hepatocellular carcinoma (HCC), a prevalent solid organ malignancy, is intricately linked to the glycolytic enzyme Alpha-enolase (ENO1). ENO1's overexpression is associated with malignant tumor progression and poor clinical outcomes, yet its role in HCC cell proliferation and differentiation warrants further investigation. This study focuses on identifying ENO1-interacting proteins in HCC, exploring their involvement in cell proliferation and the cell cycle. Materials and Methods: We employed RNA interference to downregulate ENO1 in HepG2 cells, evaluating effects on cell proliferation, cycle progression, and key signaling pathways using MTT assays, flow cytometry, and Western blotting. Identifying ENO1-interacting proteins involved combining RNAi and co-immunoprecipitation with isobaric tags (iTRAQ). We confirmed five proteins central to proliferation and cell cycle regulation through immunofluorescence microscopy and additional co-immunoprecipitation. Pathway analysis and follow-up experiments were conducted for further elucidation. Results: ENO1 silencing significantly inhibited HepG2 cell proliferation and extended G2/M and S phases. We identified 40 ENO1-interacting proteins with notable expression changes, including PCNA, HSP90, PCBP1, PSME1, and PEBP1. Significant downregulation was observed in PCNA, various cyclins, and phosphorylated ERK1/2, MAPK, and AKT, alongside an upregulation of P21. However, levels of CDK1, CDK2, ERK1/2, MAPK, and AKT remained unchanged. Conclusions: This study highlights the glycolytic pathway's crucial role in HCC development. The identified ENO1-interacting proteins like PCNA, HSP90, PCBP1, PSME1, and PEBP1 are key in tumor development, differentiation, invasion, and metastasis. The interaction between ENO1 and PCNA, influencing cell cycle regulators and phosphorylation processes, offers new insights into HCC molecular dynamics and potential therapeutic avenues.

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Section: Discussionmentioning

confidence: 99%

Exploring the Role of ENO1 in Regulating Cell Proliferation and Cell Cycle in Hepatocellular Carcinoma: Insights from Proteomic Analysis

Ma,

Yang,

Luo

et al. 2024

Preprint

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“…E6AP ubiquitinates and degrades AIB1 in breast cancer cell lines [ 62 ]. E6AP also ubiquitinates and degrades Enolase1 (ENO1), a glycolytic enzyme [ 63 ]. ENO1 expression is increased in breast cancer [ 64 ], which is likely to play a role in cancer-specific alterations in energy metabolism, i.e., Warburg effects, and also in metastasis via enzymatic degradation of extracellular matrix [ 65 , 66 ].…”

Section: E6ap In Cancermentioning

confidence: 99%

“…E6AP ubiquitinates and degrades AIB1 in breast cancer cell lines [62]. E6AP also ubiquitinates and degrades Enolase1 (ENO1), a glycolytic enzyme [63].…”

Section: Breast Cancermentioning

confidence: 99%

The HECT E3 Ligase E6AP/UBE3A as a Therapeutic Target in Cancer and Neurological Disorders

Owais

Mishra

Kiyokawa

2020

Cancers

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The HECT (Homologous to the E6-AP Carboxyl Terminus)-family protein E6AP (E6-associated protein), encoded by the UBE3A gene, is a multifaceted ubiquitin ligase that controls diverse signaling pathways involved in cancer and neurological disorders. The oncogenic role of E6AP in papillomavirus-induced cancers is well known, with its action to trigger p53 degradation in complex with the E6 viral oncoprotein. However, the roles of E6AP in non-viral cancers remain poorly defined. It is well established that loss-of-function alterations of the UBE3A gene cause Angelman syndrome, a severe neurodevelopmental disorder with autosomal dominant inheritance modified by genomic imprinting on chromosome 15q. Moreover, excess dosage of the UBE3A gene markedly increases the penetrance of autism spectrum disorders, suggesting that the expression level of UBE3A must be regulated tightly within a physiologically tolerated range during brain development. In this review, current the knowledge about the substrates of E6AP-mediated ubiquitination and their functions in cancer and neurological disorders is discussed, alongside with the ongoing efforts to pharmacologically modulate this ubiquitin ligase as a promising therapeutic target.

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Hakai, a novel Runx2 interacting protein, augments osteoblast differentiation by rescuing Runx2 from Smurf2‐mediated proteasome degradation

Upadhyay,

Sharma,

Sethi

et al. 2024

Journal Cellular Physiology

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Runt‐related transcription factor 2 (Runx2) is a key regulator of osteoblast differentiation and bone formation. In Runx2‐deficient embryos, skeletal development ceases at the cartilage anlage stage. These embryos die of respiratory failure upon birth and display a complete absence of bone and cartilage mineralization. Here, we identified Hakai, a type of E3 ubiquitin ligase as a potential Runx2 interacting partner through affinity pulldown‐based proteomic approach. Subsequently, we observed that similar to Runx2, Hakai was downregulated in osteopenic ovariectomized rats, suggesting its involvement in bone formation. Consistent with this observation, Hakai overexpression significantly enhanced osteoblast differentiation in mesenchyme‐like C3H10T1/2 as well as primary rat calvaria osteoblast (RCO) cells in vitro. Conversely, overexpression of a catalytically inactive Hakai mutant (C109A) exhibited minimal to no effect, whereas Hakai depletion markedly reduced endogenous Runx2 levels and impaired osteogenic differentiation in both C3H10T1/2 and RCOs. Mechanistically, Hakai physically interacts with Runx2 and enhances its protein turnover by rescuing it from Smad ubiquitination regulatory factor 2 (Smurf2)‐mediated proteasome degradation. Wild‐type Hakai but not Hakai‐C109A inhibited Smurf2 protein levels through proteasome‐mediated degradation. These findings underscore Hakai's functional role in bone formation, primarily through its positive modulation of Runx2 protein turnover by protecting it from Smurf2‐mediated ubiquitin‐proteasomal degradation. Collectively, our results demonstrate Hakai as a promising novel therapeutic target for osteoporosis.

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Nano‐LC based proteomic approach identifies that E6AP interacts with ENO1 and targets it for degradation in breast cancer cells

Cited by 6 publications

References 27 publications

Exploring the Role of ENO1 in Regulating Cell Proliferation and Cell Cycle in Hepatocellular Carcinoma: Insights from Proteomic Analysis

Exploring the Role of ENO1 in Regulating Cell Proliferation and Cell Cycle in Hepatocellular Carcinoma: Insights from Proteomic Analysis

The HECT E3 Ligase E6AP/UBE3A as a Therapeutic Target in Cancer and Neurological Disorders

Hakai, a novel Runx2 interacting protein, augments osteoblast differentiation by rescuing Runx2 from Smurf2‐mediated proteasome degradation

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