Ubiquitin-like (UBX)-domain-containing protein, UBXN2A, promotes cell death by interfering with the p53-Mortalin interactions in colon cancer cells

Sane, Sanam; Abdullah, Ammara; Boudreau, Donald A.; Autenried, Rebecca; Gupta, Brij K.; Wang, Xu-Chen; Wang, H.; Schlenker, Evelyn H.; Zhang, Dong; Telleria, Carlos; Huang, Lan; Chauhan, Subhash C.; Rezvani, Khosrow

doi:10.1038/cddis.2014.100

Cited by 45 publications

(72 citation statements)

References 59 publications

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“…Most recently, it was shown to activate telomerase and hnRNP-K proteins and contribute to malignant phenotype of cancer cells (23). In agreement with these reports, knockdown of mortalin in cancer cells was shown to activate p53 function and cause their growth arrest or apoptosis (17,18,(24)(25)(26)(27). Some studies have shown correlation of mortalin expression level with metastatic potential and tumor recurrence in case of hepatocellular carcinoma, suggesting clinical application of mortalin as a chemotherapeutic drug target (17,28,29).…”

Section: Introductionsupporting

confidence: 68%

Stress Chaperone Mortalin Contributes to Epithelial-to-Mesenchymal Transition and Cancer Metastasis

Kaul

Ryu

et al. 2016

Cancer Research

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Mortalin/mthsp70 (HSPA9) is a stress chaperone enriched in many cancers that has been implicated in carcinogenesis by promoting cell proliferation and survival. In this study, we examined the clinical relevance of mortalin upregulation in carcinogenesis. Consistent with high mortalin expression in various human tumors and cell lines, we found that mortalin overexpression increased the migration and invasiveness of breast cancer cells. Expression analyses revealed that proteins involved in focal adhesion, PI3K-Akt, and JAK-STAT signaling, all known to play key roles in cell migration and epithelial-tomesenchymal transition (EMT), were upregulated in mortalinexpressing cancer cells. We further determined that expression levels of the mesenchymal markers vimentin (VIM), fibronectin (FN1), b-catenin (CTNNB1), CK14 (KRT14), and hnRNP-K were also increased upon mortalin overexpression, whereas the epithelial markers E-cadherin (CDH1), CK8 (KRT8), and CK18 (KRT18) were downregulated. Furthermore, shRNAmediated and pharmacologic inhibition of mortalin suppressed the migration and invasive capacity of cancer cells and was associated with a diminished EMT gene signature. Taken together, these findings support a role for mortalin in the induction of EMT, prompting further investigation of its therapeutic value in metastatic disease models. Cancer Res; 76(9);

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Section: Introductionsupporting

confidence: 68%

Stress Chaperone Mortalin Contributes to Epithelial-to-Mesenchymal Transition and Cancer Metastasis

Kaul

Ryu

et al. 2016

Cancer Research

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“…Colony formation assay Transfected HCT-116 cells expressing (His) 6 -TYG-tagged UBXN2A or (His) 6 -TYG-tagged empty vector were selected with Zeocin (100 μg/ml) for 2 days as described previously (Sane et al 2014). Transfected cells were seeded (1000 cells per disc) in a 100-mm culture disc for 9 days for cells to form colonies.…”

Section: Molecular Modelingmentioning

confidence: 99%

“…3) using Lipofectamine 2000 (Life Technologies). In a set of experiments, (His) 6 -TYG-tagged UBXN2A or GFP-UBXN2A cloned in pAcGFP1-C1 (Clontech, Mountain View, CA) (Sane et al 2014) were cotransfected with (His) 6 -CHIP E3 ubiquitin ligase followed by WB analysis or immunocytochemical (Sane et al 2014) detection of p53.…”

Section: Molecular Modelingmentioning

confidence: 99%

“…Mortalin does not quite fit with other members of the HSP70 family due to its non-inducibility to heat shocks (Kaul et al 2007). Besides inactivation of p53 during the progression of cancer (Lu et al 2011;Sane et al 2014). mortalin enhances cancer proliferation and metastasis through other pathways as follows: (i) Mortalin negatively regulates the Raf/MEK/ERK pathway, which triggers innate tumorsuppressive mechanisms.…”

Section: Introductionmentioning

confidence: 99%

“…We discovered UBXN2A binds to and inactivates mortalin. By binding to mortalin, UBXN2A reactivates wild-type (WT)-p53 tumor suppressor protein (Sane et al 2014) and induces apoptosis at the cellular level and in tumor xenografts (Abdullah et al 2015b). In addition, we discovered that induction of UBXN2A by its enhancer (Abdullah et al 2015a).…”

Section: Introductionmentioning

confidence: 99%

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Structural studies of UBXN2A and mortalin interaction and the putative role of silenced UBXN2A in preventing response to chemotherapy

Sane

Abdullah

Nelson

et al. 2016

Cell Stress and Chaperones

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Overexpression of the oncoprotein mortalin in cancer cells and its protein partners enables mortalin to promote multiple oncogenic signaling pathways and effectively antagonize chemotherapy-induced cell death. A UBX-domaincontaining protein, UBXN2A, acts as a potential mortalin inhibitor. This current study determines whether UBXN2A effectively binds to and occupies mortalin's binding pocket, resulting in a direct improvement in the tumor's sensitivity to chemotherapy. Molecular modeling of human mortalin's binding pocket and its binding to the SEP domain of UBXN2A followed by yeast two-hybrid and His-tag pulldown assays revealed that three amino acids (PRO442, ILE558, and LYS555) within the substrate-binding domain of mortalin are crucial for UBXN2A binding to mortalin. As revealed by chase experiments in the presence of cycloheximide, overexpression of UBXN2A seems to interfere with the mortalin-CHIP E3 ubiquitin ligase and consequently suppresses the C-terminus of the HSC70-interacting protein (CHIP)-mediated destabilization of p53, resulting in its stabilization in the cytoplasm and upregulation in the nucleus. Overexpression of UBXN2A causes a significant inhibition of cell proliferation and the migration of colon cancer cells. We silenced UBXN2A in the human osteosarcoma U2OS cell line, an enriched mortalin cancer cell, followed by a clinical dosage of the chemotherapeutic agent 5-fluorouracil (5-FU). The UBXN2A knockout U2OS cells revealed that UBXNA is essential for the cytotoxic effect achieved by 5-FU. UBXN2A overexpression markedly increased the apoptotic response of U2OS cells to the 5-FU. In addition, silencing of UBXN2A protein suppresses apoptosis enhanced by UBXN2A overexpression in U2OS. The knowledge gained from this study provides insights into the mechanistic role of UBXN2A as a potent mortalin inhibitor and as a potential chemotherapy sensitizer for clinical application.

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UBXN2A enhances CHIP‐mediated proteasomal degradation of oncoprotein mortalin‐2 in cancer cells

et al. 2018

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Overexpression of oncoproteins is a major cause of treatment failure using current chemotherapeutic drugs. Drug‐induced degradation of oncoproteins is feasible and can improve clinical outcomes in diverse types of cancers. Mortalin‐2 (mot‐2) is a dominant oncoprotein in several tumors, including colorectal cancer (CRC). In addition to inactivating the p53 tumor suppressor protein, mot‐2 enhances tumor cell invasion and migration. Thus, mot‐2 is considered a potential therapeutic target in several cancer types. The current study investigated the biological role of a ubiquitin‐like protein called UBXN2A in the regulation of mot‐2 turnover. An orthogonal ubiquitin transfer technology followed by immunoprecipitation, in vitro ubiquitination, and Magnetic Beads TUBE2 pull‐down experiments revealed that UBXN2A promotes carboxyl terminus of the HSP70‐interacting protein (CHIP)‐dependent ubiquitination of mot‐2. We subsequently showed that UBXN2A increases proteasomal degradation of mot‐2. A subcellular compartmentalization experiment revealed that induced UBXN2A decreases the level of mot‐2 and its chaperone partner, HSP60. Pharmacological upregulation of UBXN2A using a small molecule, veratridine (VTD), decreases the level of mot‐2 in cancer cells. Consistent with the in vitro results, UBXN2A+/− mice exhibited selective elevation of mot‐2 in colon tissues. An in vitro Anti‐K48 TUBE isolation approach showed that recombinant UBXN2A enhances proteasomal degradation of mot‐2 in mouse colon tissues. Finally, we observed enhanced association of CHIP with the UBXN2A‐mot‐2 complex in tumors in an azoxymethane/dextran sulfate sodium‐induced mouse CRC model. The existence of a multiprotein complex containing UBXN2A, CHIP, and mot‐2 suggests a synergistic tumor suppressor activity of UBXN2A and CHIP in mot‐2‐enriched tumors. This finding validates the UBXN2A‐CHIP axis as a novel and potential therapeutic target in CRC.

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Ubiquitin-like (UBX)-domain-containing protein, UBXN2A, promotes cell death by interfering with the p53-Mortalin interactions in colon cancer cells

Cited by 45 publications

References 59 publications

Stress Chaperone Mortalin Contributes to Epithelial-to-Mesenchymal Transition and Cancer Metastasis

Stress Chaperone Mortalin Contributes to Epithelial-to-Mesenchymal Transition and Cancer Metastasis

Structural studies of UBXN2A and mortalin interaction and the putative role of silenced UBXN2A in preventing response to chemotherapy

UBXN2A enhances CHIP‐mediated proteasomal degradation of oncoprotein mortalin‐2 in cancer cells

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