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ASNA1, an ATPase targeting tail-anchored proteins, regulates melanoma cell growth and sensitivity to cisplatin and arsenite
Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Surgery.
Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Surgery.
Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Surgery.
Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Surgery.
2009 (English)In: Cancer Chemotherapy and Pharmacology, ISSN 0344-5704, E-ISSN 1432-0843, Vol. 63, no 3, 491-499 p.Article in journal (Refereed) Published
Abstract [en]

Purpose ASNA1 is homologous to E. coli ArsA, a well characterized ATPase involved in efflux of arsenite and antimonite. Cells resistant to arsenite and antimonite are cross-resistant to the chemotherapeutic drug cisplatin. ASNA1 is also an essential ATPase for the insertion of tail-anchored proteins into ER membranes and a novel regulator of insulin secretion. The aim of this study was to determine if altered ASNA1 levels influenced growth and sensitivity to arsenite and cisplatin in human melanoma cells.

Methods Cultured melanoma T289 cells were transfected with plasmids containing sense or antisense ASNA1. Cells were exposed to cisplatin, arsenite and zinc. Cell growth and chemosensitivity were evaluated by the MTT assay and apoptosis by a TUNEL assay.

Results ASNA1 expression was necessary for growth. T289 clones with decreased ASNA1 expression exhibited 51 ± 5% longer doubling times than wildtype T289 (P = 0.0091). After exposure to cisplatin, ASNA1 downregulated cells displayed a significant increase in apoptosis. The cisplatin IC50 in ASNA1 underexpressing cells was 41.7 ± 1.8% compared to wildtype (P = 0.00097) and the arsenite IC50 was 59.9 ± 3.2% of wildtype IC50 (P = 0.0067).

Conclusions Reduced ASNA1 expression is associated with significant inhibition of cell growth, increased apoptosis and increased sensitivity to cisplatin and arsenite.

Place, publisher, year, edition, pages
Springer , 2009. Vol. 63, no 3, 491-499 p.
Keyword [en]
ASNA1, Cisplatin, Arsenite, Melanoma, Drug resistance
National Category
Cancer and Oncology Surgery Pharmaceutical Sciences
Research subject
cellforskning; Pharmaceutical Biochemistry; Oncology
URN: urn:nbn:se:umu:diva-10210DOI: 10.1007/s00280-008-0762-2PubMedID: 18478230OAI: diva2:149881
Available from: 2008-11-18 Created: 2008-11-18 Last updated: 2013-11-29Bibliographically approved
In thesis
1. ASNA1 and cisplatin resistance: studies in C. elegans and in human tumor cells
Open this publication in new window or tab >>ASNA1 and cisplatin resistance: studies in C. elegans and in human tumor cells
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Platinum based chemotherapy is widely used to treat cancer. Cisplatin (diamminedichloroplatinum) combination treatments provide cure for metastatic testicular cancer and prolong survival for patients suffering from ovarian, head and neck, bladder and non small cell lung cancer. Tumors that initially respond to treatment may eventually acquire resistance, resulting in treatment failure. Cisplatin resistant cells are crossresistant to arsenite and antimonite and these metalloids are exported from bacteria by the ars-operon.

In this thesis, we describe the human ArsA homolog, ASNA1, as a protein involved in a novel resistance mechanism to cisplatin, arsenite and antimonite. ASNA1 was downregulated by antisense and siRNA techniques in human melanoma and ovarian carcinoma cell lines. These cells displayed increased sensitivity to arsenite and the platinum based drugs cisplatin, carboplatin and oxaliplatin. In both melanoma and ovarian carcinoma, cisplatin resistant cells overexpressed ASNA1.

Blockage of ASNA1 resulted in increased apoptosis and retarded growth, complicating further characterization of ASNA1 in human cell lines. ASNA1 also promotes insulin signaling and mediates membrane insertion of tail-anchored proteins. To explore different aspects of ASNA1 function with respect to cisplatin resistance, we used the model organism C. elegans.

In the nematode C. elegans, asna-1 (rnai) treated larvae were hypersensitive to cisplatin, arsenite and antimonite. Adult asna-1 mutant worms were cisplatin sensitive and this hypersensitivity was seen even when apoptosis was blocked. Expression of human ASNA1 rescued the cisplatin hypersensitivity in asna-1 mutants, showing conservation of function. Transgene expression of mutated forms of asna-1 separated the cisplatin hypersensitivity phenotype from the insulin signaling phenotype of asna-1 mutants. Three ASNA-1 residues, His164, Cys285 and Cys288 were identified as essential for ASNA-1 promoted cisplatin resistance but not for insulin signaling. Finally, studies of the C. elegans germline revealed increased numbers of apoptotic cells in asna-1 mutants.

In conclusion, C. elegans is a suitable model organism to identify and characterize cisplatin response mechanisms. A targeted therapy against ASNA1 could sensitize cisplatin resistant cells and improve outcome for cancer patients.

Place, publisher, year, edition, pages
Umeå: Umeå Universitet, 2010. 78 p.
Umeå University medical dissertations, ISSN 0346-6612 ; 1390
ASNA1, Cisplatin, C. elegans, Drug resistance, Apoptosis
National Category
Surgery Cancer and Oncology
Research subject
Surgery; Oncology; cellforskning
urn:nbn:se:umu:diva-37661 (URN)978-91-7459-115-6 (ISBN)
Public defence
2010-12-10, Sal B, 9tr, Tandläkarhögskolan, Norrlands Universitetssjukhus, Umeå, 11:17 (Swedish)
Swedish Research Council, K2008-68X-20803-01-3
Available from: 2010-11-19 Created: 2010-11-11 Last updated: 2010-11-19Bibliographically approved

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