ADAM12-L is a direct target of the miR-29 and miR-200 families in breast cancer

BMC Cancer

Sara Duhachek-Muggy¹  Anna Zolkiewska¹ 

[1] Department of Biochemistry and Molecular Biophysics, Kansas State University, 141 Chalmers Hall, Manhattan 66506, KS, USA

关键词: Epithelial-to-mesenchymal transition;    Claudin-low tumors;    Breast cancer;    Post-transcriptional gene regulation;    microRNA;    Alternative splicing;    Disintegrin;    Metalloproteinase;   

Others  :  1136250
DOI  :  10.1186/s12885-015-1108-1

 received in 2014-09-02, accepted in 2015-02-20,  发布年份 2015

【 摘 要 】

Background

ADAM12-L and ADAM12-S represent two major splice variants of human metalloproteinase-disintegrin 12 mRNA, which differ in their 3′-untranslated regions (3′UTRs). ADAM12-L, but not ADAM12-S, has prognostic and chemopredictive values in breast cancer. Expression levels of the two ADAM12 splice variants in clinical samples are highly discordant, suggesting post-transcriptional regulation of the ADAM12 gene. The miR-29, miR-30, and miR-200 families have potential target sites in the ADAM12-L 3′UTR and they may negatively regulate ADAM12-L expression.

Methods

miR-29b/c, miR-30b/d, miR-200b/c, or control miRNA mimics were transfected into SUM159PT, BT549, SUM1315MO2, or Hs578T breast cancer cells. ADAM12-L and ADAM12-S mRNA levels were measured by qRT-PCR, and ADAM12-L protein was detected by Western blotting. Direct targeting of the ADAM12-L 3′UTR by miRNAs was tested using an ADAM12-L 3′UTR luciferase reporter. The rate of ADAM12-L translation was evaluated by metabolic labeling of cells with ³⁵S cysteine/methionine. The roles of endogenous miR-29b and miR-200c were tested by transfecting cells with miRNA hairpin inhibitors.

Results

Transfection of miR-29b/c mimics strongly decreased ADAM12-L mRNA levels in SUM159PT and BT549 cells, whereas ADAM12-S levels were not changed. ADAM12-L, but not ADAM12-S, levels were also significantly diminished by miR-200b/c in SUM1315MO2 cells. In Hs578T cells, miR-200b/c mimics impeded translation of ADAM12-L mRNA. Importantly, both miR-29b/c and miR-200b/c strongly decreased steady state levels of ADAM12-L protein in all breast cancer cell lines tested. miR-29b/c and miR-200b/c also significantly decreased the activity of an ADAM12-L 3′UTR reporter, and this effect was abolished when miR-29b/c and miR-200b/c target sequences were mutated. In contrast, miR-30b/d did not elicit consistent and significant effects on ADAM12-L expression. Analysis of a publicly available gene expression dataset for 100 breast tumors revealed a statistically significant negative correlation between ADAM12-L and both miR-29b and miR-200c. Inhibition of endogenous miR-29b and miR-200c in SUM149PT and SUM102PT cells led to increased ADAM12-L expression.

Conclusions

The ADAM12-L 3′UTR is a direct target of miR-29 and miR-200 family members. Since the miR-29 and miR-200 families play important roles in breast cancer progression, these results may help explain the different prognostic and chemopredictive values of ADAM12-L and ADAM12-S in breast cancer.

【 授权许可】

   
2015 Duhachek-Muggy and Zolkiewska; licensee BioMed Central.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

【 参考文献 】

• [1]Kveiborg M, Albrechtsen R, Couchman JR, Wewer UM: Cellular roles of ADAM12 in health and disease. Int J Biochem Cell Biol 2008, 40:1685-702.
• [2]Nyren-Erickson EK, Jones JM, Srivastava DK, Mallik S: A disintegrin and metalloproteinase-12 (ADAM12): function, roles in disease progression, and clinical implications. Biochim Biophys Acta 1830, 2013:4445-55.
• [3]Kveiborg M, Frohlich C, Albrechtsen R, Tischler V, Dietrich N, Holck P, et al.: A role for ADAM12 in breast tumor progression and stromal cell apoptosis. Cancer Res 2005, 65:4754-61.
• [4]Frohlich C, Nehammer C, Albrechtsen R, Kronqvist P, Kveiborg M, Sehara-Fujisawa A, et al.: ADAM12 produced by tumor cells rather than stromal cells accelerates breast tumor progression. Mol Cancer Res 2011, 9:1449-61.
• [5]Dyczynska E, Syta E, Sun D, Zolkiewska A: Breast cancer-associated mutations in metalloprotease disintegrin ADAM12 interfere with the intracellular trafficking and processing of the protein. Int J Cancer 2008, 122:2634-40.
• [6]Stautz D, Wewer UM, Kveiborg M: Functional analysis of a breast cancer-associated mutation in the intracellular domain of the metalloprotease ADAM12. PLoS One 2012, 7:e37628.
• [7]Qi Y, Duhachek-Muggy S, Li H, Zolkiewska A: Phenotypic diversity of breast cancer-related mutations in metalloproteinase-disintegrin ADAM12. PLoS One 2014, 9:e92536.
• [8]Jacobsen J, Wewer UM: Targeting ADAM12 in human disease: head, body or tail? Curr Pharm Des. 2009, 15:2300-10.
• [9]Duhachek-Muggy S, Li H, Qi Y, Zolkiewska A: Alternative mRNA splicing generates two distinct ADAM12 prodomain variants. PLoS One 2013, 8:e75730.
• [10]Prat A, Parker JS, Karginova O, Fan C, Livasy C, Herschkowitz JI, et al.: Phenotypic and molecular characterization of the claudin-low intrinsic subtype of breast cancer. Breast Cancer Res 2010, 12:R68. BioMed Central Full Text
• [11]Harrell JC, Pfefferle AD, Zalles N, Prat A, Fan C, Khramtsov A, et al.: Endothelial-like properties of claudin-low breast cancer cells promote tumor vascular permeability and metastasis. Clin Exp Metastasis 2014, 31:33-45.
• [12]Li H, Duhachek-Muggy S, Dubnicka S, Zolkiewska A: Metalloproteinase-disintegrin ADAM12 is associated with a breast tumor-initiating cell phenotype. Breast Cancer Res Treat 2013, 139:691-703.
• [13]Sabatier R, Finetti P, Guille A, Adelaide J, Chaffanet M, Viens P, et al.: Claudin-low breast cancers: clinical, pathological, molecular and prognostic characterization. Mol Cancer 2014, 13:228. BioMed Central Full Text
• [14]Taube JH, Herschkowitz JI, Komurov K, Zhou AY, Gupta S, Yang J, et al.: Core epithelial-to-mesenchymal transition interactome gene-expression signature is associated with claudin-low and metaplastic breast cancer subtypes. Proc Natl Acad Sci U S A 2010, 107:15449-54.
• [15]Gupta PB, Onder TT, Jiang G, Tao K, Kuperwasser C, Weinberg RA, et al.: Identification of selective inhibitors of cancer stem cells by high-throughput screening. Cell 2009, 138:645-59.
• [16]Onder TT, Gupta PB, Mani SA, Yang J, Lander ES, Weinberg RA: Loss of E-cadherin promotes metastasis via multiple downstream transcriptional pathways. Cancer Res 2008, 68:3645-54.
• [17]Scheel C, Eaton EN, Li SH, Chaffer CL, Reinhardt F, Kah KJ, et al.: Paracrine and autocrine signals induce and maintain mesenchymal and stem cell states in the breast. Cell 2011, 145:926-40.
• [18]Dontu G, Abdallah WM, Foley JM, Jackson KW, Clarke MF, Kawamura MJ, et al.: In vitro propagation and transcriptional profiling of human mammary stem/progenitor cells. Genes Dev 2003, 17:1253-70.
• [19]Creighton CJ, Li X, Landis M, Dixon JM, Neumeister VM, Sjolund A, et al.: Residual breast cancers after conventional therapy display mesenchymal as well as tumor-initiating features. Proc Natl Acad Sci U S A 2009, 106:13820-5.
• [20]Miller WR, Larionov AA, Renshaw L, Anderson TJ, White S, Murray J, et al.: Changes in breast cancer transcriptional profiles after treatment with the aromatase inhibitor, letrozole. Pharmacogenet Genomics 2007, 17:813-26.
• [21]Farmer P, Bonnefoi H, Anderle P, Cameron D, Wirapati P, Becette V, et al.: A stroma-related gene signature predicts resistance to neoadjuvant chemotherapy in breast cancer. Nat Med 2009, 15:68-74.
• [22]Ayers M, Symmans WF, Stec J, Damokosh AI, Clark E, Hess K, et al.: Gene expression profiles predict complete pathologic response to neoadjuvant paclitaxel and fluorouracil, doxorubicin, and cyclophosphamide chemotherapy in breast cancer. J Clin Oncol 2004, 22:2284-93.
• [23]Popovici V, Chen W, Gallas BG, Hatzis C, Shi W, Samuelson FW, et al.: Effect of training-sample size and classification difficulty on the accuracy of genomic predictors. Breast Cancer Res 2010, 12:R5. BioMed Central Full Text
• [24]Wang Y, Klijn JG, Zhang Y, Sieuwerts AM, Look MP, Yang F, et al.: Gene-expression profiles to predict distant metastasis of lymph-node-negative primary breast cancer. Lancet 2005, 365:671-9.
• [25]Minn AJ, Gupta GP, Padua D, Bos P, Nguyen DX, Nuyten D, et al.: Lung metastasis genes couple breast tumor size and metastatic spread. Proc Natl Acad Sci U S A 2007, 104:6740-5.
• [26]Desmedt C, Piette F, Loi S, Wang Y, Lallemand F, Haibe-Kains B, et al.: Strong time dependence of the 76-gene prognostic signature for node-negative breast cancer patients in the TRANSBIG multicenter independent validation series. Clin Cancer Res 2007, 13:3207-14.
• [27]Li H, Duhachek-Muggy S, Qi Y, Hong Y, Behbod F, Zolkiewska A: An essential role of metalloprotease-disintegrin ADAM12 in triple-negative breast cancer. Breast Cancer Res Treat 2012, 135:759-69.
• [28]Schwarzenbacher D, Balic M, Pichler M: The role of microRNAs in breast cancer stem cells. Int J Mol Sci 2013, 14:14712-23.
• [29]D’Amato NC, Howe EN, Richer JK: MicroRNA regulation of epithelial plasticity in cancer. Cancer Lett 2013, 341:46-55.
• [30]Shimono Y, Zabala M, Cho RW, Lobo N, Dalerba P, Qian D, et al.: Downregulation of miRNA-200c links breast cancer stem cells with normal stem cells. Cell 2009, 138:592-603.
• [31]Gregory PA, Bert AG, Paterson EL, Barry SC, Tsykin A, Farshid G, et al.: The miR-200 family and miR-205 regulate epithelial to mesenchymal transition by targeting ZEB1 and SIP1. Nat Cell Biol 2008, 10:593-601.
• [32]Gregory PA, Bracken CP, Smith E, Bert AG, Wright JA, Roslan S, et al.: An autocrine TGF-beta/ZEB/miR-200 signaling network regulates establishment and maintenance of epithelial-mesenchymal transition. Mol Biol Cell 2011, 22:1686-98.
• [33]Wright JA, Richer JK, Goodall GJ: microRNAs and EMT in mammary cells and breast cancer. J Mammary Gland Biol Neoplasia 2010, 15:213-23.
• [34]Chou J, Lin JH, Brenot A, Kim JW, Provot S, Werb Z: GATA3 suppresses metastasis and modulates the tumour microenvironment by regulating microRNA-29b expression. Nat Cell Biol 2013, 15:201-13.
• [35]Cittelly DM, Finlay-Schultz J, Howe EN, Spoelstra NS, Axlund SD, Hendricks P, et al.: Progestin suppression of miR-29 potentiates dedifferentiation of breast cancer cells via KLF4. Oncogene 2013, 32:2555-64.
• [36]Yu F, Deng H, Yao H, Liu Q, Su F, Song E: Mir-30 reduction maintains self-renewal and inhibits apoptosis in breast tumor-initiating cells. Oncogene 2010, 29:4194-204.
• [37]Ouzounova M, Vuong T, Ancey PB, Ferrand M, Durand G, Le-Calvez Kelm F, et al.: MicroRNA miR-30 family regulates non-attachment growth of breast cancer cells. BMC Genomics 2013, 14:139. BioMed Central Full Text
• [38]Riaz M, van Jaarsveld MT, Hollestelle A, der Smissen WJ P-v, Heine AA, Boersma AW, et al.: miRNA expression profiling of 51 human breast cancer cell lines reveals subtype and driver mutation-specific miRNAs. Breast Cancer Res 2013, 15:R33. BioMed Central Full Text
• [39]Prat A, Karginova O, Parker JS, Fan C, He X, Bixby L, et al.: Characterization of cell lines derived from breast cancers and normal mammary tissues for the study of the intrinsic molecular subtypes. Breast Cancer Res Treat 2013, 142:237-55.
• [40]Neve RM, Chin K, Fridlyand J, Yeh J, Baehner FL, Fevr T, et al.: A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. Cancer Cell 2006, 10:515-27.
• [41]Nygren MK, Tekle C, Ingebrigtsen VA, Makela R, Krohn M, Aure MR, et al.: Identifying microRNAs regulating B7-H3 in breast cancer: the clinical impact of microRNA-29c. Br J Cancer 2014, 110:2072-80.
• [42]Iorio MV, Ferracin M, Liu CG, Veronese A, Spizzo R, Sabbioni S, et al.: MicroRNA gene expression deregulation in human breast cancer. Cancer Res 2005, 65:7065-70.
• [43]Fabian MR, Sonenberg N, Filipowicz W: Regulation of mRNA translation and stability by microRNAs. Annu Rev Biochem 2010, 79:351-79.
• [44]Enerly E, Steinfeld I, Kleivi K, Leivonen SK, Aure MR, Russnes HG, et al.: miRNA-mRNA integrated analysis reveals roles for miRNAs in primary breast tumors. PLoS One 2011, 6:e16915.
• [45]Li H, Solomon E, Duhachek Muggy S, Sun D, Zolkiewska A: Metalloprotease-disintegrin ADAM12 expression is regulated by Notch signaling via microRNA-29. J Biol Chem 2011, 286:21500-10.
• [46]Ramdas V, McBride M, Denby L, Baker AH: Canonical transforming growth factor-beta signaling regulates disintegrin metalloprotease expression in experimental renal fibrosis via miR-29. Am J Pathol 2013, 183:1885-96.
• [47]Luna C, Li G, Qiu J, Epstein DL, Gonzalez P: Role of miR-29b on the regulation of the extracellular matrix in human trabecular meshwork cells under chronic oxidative stress. Mol Vis 2009, 15:2488-97.