Table S1 lines. (DOCX) Table S3 variables. (DOCX) References The fluorescent protein tagged to the CDC25AQ110del dominated upon overlap. (TIF) CDC25A cDNA clones retrieved from NSCLC cell Table S2 CDC25AQ110del expression in NSCLC tumor tissue and overall survival. (DOCX) Tumor CDC25AQ110del expression and demographic 1. Kim AW, Boffa DJ, Wang Z, Detterbeck FC (2012) An analysis, systematic review, and meta-analysis of the perioperative mortality after neoadjuvant therapy and pneumonectomy for non-small cell lung cancer. J Thorac Cardiovasc Surg 143: 55–63. 2. Alvarez M, Roman E, Santos ES, Raez LE (2007) New targets for non-small-cell lung cancer therapy. Expert Rev Anticancer Ther 7: 1423–1437. 3. Mori S, Ito G, Usami N, Yoshioka H, Ueda Y, et al. (2004) p53 apoptotic pathway molecules are frequently and simultaneously altered in nonsmall cell lung carcinoma. Cancer 100: 1673–1682. 4. Caputi M, Russo G, Esposito V, Mancini A, Giordano A (2005) Role of cell- cycle regulators in lung cancer. J Cell Physiol 205: 319–327. 5. Baldi A, Esposito V, De Luca A, Fu Y, Meoli I, et al. (1997) Differential expression of Rb2/p130 and p107 in normal human tissues and in primary lung cancer. Clin Cancer Res 3: 1691–1697. 6. Shapiro GI, Edwards CD, Ewen ME, Rollins BJ (1998) p16INK4A participates in a G1 arrest checkpoint in response to DNA damage. Mol Cell Biol 18: 378– 387. 7. Wu W, Fan YH, Kemp BL, Walsh G, Mao L (1998) Overexpression of cdc25A and cdc25B is frequent in primary non-small cell lung cancer but is not associated with overexpression of c-myc. Cancer Res 58: 4082–4085. 8. Busino L, Chiesa M, Draetta GF, Donzelli M (2004) Cdc25A phosphatase: combinatorial phosphorylation, ubiquitylation and proteolysis. Oncogene 23: 2050–2056. 9. Galaktionov K, Lee AK, Eckstein J, Draetta G, Meckler J, et al. (1995) CDC25 phosphatases as potential human oncogenes. Science 269: 1575–1577. 10. Bartek J, Lukas C, Lukas J (2004) Checking on DNA damage in S phase. Nature Reviews Molecular Cell Biology 5: 792–804. 11. Mailand N, Podtelejnikov AV, Groth A, Mann M, Bartek J, et al. (2002) Regulation of G(2)/M events by Cdc25A through phosphorylation-dependent modulation of its stability. EMBO J 21: 5911–5920. 12. Kristjansdottir K, Rudolph J (2004) Cdc25 phosphatases and cancer. Chem Biol 11: 1043–1051. 13. Bhawal UK, Sugiyama M, Nomura Y, Sawajiri M, Tsukinoki K, et al. (2007) High-risk human papillomavirus type 16 E7 oncogene associates with Cdc25A over-expression in oral squamous cell carcinoma. Virchows Arch 450: 65–71. 14. Boutros R, Lobjois V, Ducommun B (2007) CDC25 phosphatases in cancer cells: key players? Good targets? Nature Reviews Cancer 7: 495–507. 15. Xu X, Yamamoto H, Sakon M, Yasui M, Ngan CY, et al. (2003) Overexpression of CDC25A phosphatase is associated with hypergrowth activity and poor prognosis of human hepatocellular carcinomas. Clin Cancer Res 9: 1764–1772. 16. Broggini M, Buraggi G, Brenna A, Riva L, Codegoni AM, et al. (2000) Cell cycle-related phosphatases CDC25A and B expression correlates with survival in ovarian cancer patients. Anticancer Res 20: 4835–4840. 17. Albert H, Santos S, Battaglia E, Brito M, Monteiro C, et al. (2011) Differential expression of CDC25 phosphatases splice variants in human breast cancer cells. Clin Chem Lab Med. 18. Hernandez S, Bessa X, Bea S, Hernandez L, Nadal A, et al. (2001) Differential expression of cdc25 cell-cycle-activating phosphatases in human colorectal carcinoma. Lab Invest 81: 465–473. 19. Hernandez S, Hernandez L, Bea S, Cazorla M, Fernandez PL, et al. (1998) cdc25 cell cycle-activating phosphatases and c-myc expression in human non- Hodgkin’s lymphomas. Cancer Res 58: 1762–1767. 20. Lo¨ffler H, Syljua˚sen RG, Bartkova J, Worm J, Lukas J, et al. (2003) Distinct modes of deregulation of the proto-oncogenic Cdc25A phosphatase in human breast cancer cell lines. Oncogene 22: 8063–8071. 21. Pereg Y, Liu BY, O’Rourke KM, Sagolla M, Dey A, et al. (2010) Ubiquitin hydrolase Dub3 promotes oncogenic transformation by stabilizing Cdc25A. Nat Cell Biol 12: 400–406. 22. Kang T, Wei Y, Honaker Y, Yamaguchi H, Appella E, et al. (2008) GSK-3 beta targets Cdc25A for ubiquitin-mediated proteolysis, and GSK-3 beta inactivation correlates with Cdc25A overproduction in human cancers. Cancer Cell 13: 36– 47. CDC25A-Q110del Novel Isoform Role in Lung Cancer Table S4 CDC25Awt in NSCLC tumor versus normal tissue pair in correlation to overall patient survival. (DOCX) Table S5 CDC25Awt in NSCLC tumor versus normal tissue pair and demographic variables. (DOCX) Author Contributions Conceived and designed the experiments: RHY HR LM. Performed the experiments: RHY WC RL RX YM HR. Analyzed the data: RHY WC HR LM ZL. Contributed reagents/materials/analysis tools: RHY HR LM ZL. Wrote the paper: RHY HR LM MJE. 23. Li N, Zhong X, Lin X, Guo J, Zou L, et al. (2012) Lin-28 homologue A (LIN28A) promotes cell cycle progression via regulation of cyclin-dependent kinase 2 (CDK2), cyclin D1 (CCND1), and cell division cycle 25 homolog A (CDC25A) expression in cancer. J Biol Chem 287: 17386–17397. 24. Wang P, Zou F, Zhang X, Li H, Dulak A, et al. (2009) microRNA-21 negatively regulates Cdc25A and cell cycle progression in colon cancer cells. Cancer Res 69: 8157–8165. 25. Ramirez RD, Sheridan S, Girard L, Sato M, Kim Y, et al. (2004) Immortalization of human bronchial epithelial cells in the absence of viral oncoproteins. Cancer Res 64: 9027–9034. 26. Ray D, Kiyokawa H (2007) CDC25A levels determine the balance of proliferation and checkpoint response. Cell Cycle 6: 3039–3042. 27. Xiao Z, Chen Z, Gunasekera AH, Sowin TJ, Rosenberg SH, et al. (2003) Chk1 mediates S and G2 arrests through Cdc25A degradation in response to DNA- damaging agents. J Biol Chem 278: 21767–21773. 28. Fernandez-Vidal A, Mazars A, Manenti S (2008) CDC25A: a rebel within the CDC25 phosphatases family? Anticancer Agents Med Chem 8: 825–831. 29. Kasahara K, Goto H, Enomoto M, Tomono Y, Kiyono T, et al. (2010) 14-3- 3gamma mediates Cdc25A proteolysis to block premature mitotic entry after DNA damage. EMBO J 29: 2802–2812. 30. Badie C, Itzhaki JE, Sullivan MJ, Carpenter AJ, Porter AC (2000) Repression of CDK1 and other genes with CDE and CHR promoter elements during DNA damage-induced G(2)/M arrest in human cells. Mol Cell Biol 20: 2358–2366. 31. Rother K, Kirschner R, Sanger K, Bohlig L, Mossner J, et al. (2007) p53 downregulates expression of the G1/S cell cycle phosphatase Cdc25A. Oncogene 26: 1949–1953. 32. Demidova AR, Aau MY, Zhuang L, Yu Q (2009) Dual regulation of Cdc25A by Chk1 and p53-ATF3 in DNA replication checkpoint control. J Biol Chem 284: 4132–4139. 33. Fotedar R, Bendjennat M, Fotedar A (2004) Role of p21WAF1 in the cellular response to UV. Cell Cycle 3: 134–137. 34. Wegener S, Hampe W, Herrmann D, Schaller HC (2000) Alternative splicing in the regulatory region of the human phosphatases CDC25A and CDC25C. Eur J Cell Biol 79: 810–815. 35. Mailand N, Falck J, Lukas C, Syljuasen RG, Welcker M, et al. (2000) Rapid destruction of human Cdc25A in response to DNA damage. Science 288: 1425– 1429. 36. Khaled AR, Bulavin DV, Kittipatarin C, Li WQ, Alvarez M, et al. (2005) Cytokine-driven cell cycling is mediated through Cdc25A. J Cell Biol 169: 755– 763. 37. Falck J, Mailand N, Syljuasen RG, Bartek J, Lukas J (2001) The ATM-Chk2- Cdc25A checkpoint pathway guards against radioresistant DNA synthesis. Nature 410: 842–847. 38. Zhao H, Watkins JL, Piwnica-Worms H (2002) Disruption of the checkpoint kinase 1/cell division cycle 25A pathway abrogates ionizing radiation-induced S and G2 checkpoints. Proc Natl Acad Sci U S A 99: 14795–14800. 39. Shreeram S, Hee WK, Bulavin DV (2008) Cdc25A serine 123 phosphorylation couples centrosome duplication with DNA replication and regulates tumorigen- esis. Mol Cell Biol 28: 7442–7450. 40. Reinhardt HC, Aslanian AS, Lees JA, Yaffe MB (2007) p53-deficient cells rely on ATM- and ATR-mediated checkpoint signaling through the p38MAPK/ MK2 pathway for survival after DNA damage. Cancer Cell 11: 175–189. 41. Chen MS, Ryan CE, Piwnica-Worms H (2003) Chk1 kinase negatively regulates mitotic function of Cdc25A phosphatase through 14-3-3 binding. Mol Cell Biol 23: 7488–7497. 42. Kallstrom H, Lindqvist A, Pospisil V, Lundgren A, Karlssonrosenthal C (2005) Cdc25A localisation and shuttling: characterisation of sequences mediating nuclear export and import. Experimental Cell Research 303: 89–100. 43. Yaffe MB, Rittinger K, Volinia S, Caron PR, Aitken A, et al. (1997) The structural basis for 14-3-3:phosphopeptide binding specificity. Cell 91: 961–971. 44. Carbone DP (2009) Profiles in variation: lung carcinogenesis. Cancer Prev Res (Phila) 2: 695–697. PLOS ONE | www.plosone.org 10 October 2012 | Volume 7 | Issue 10 | e46464