Trophoblast Cell Surface Antigen 2 (Trop-2) Phosphorylation by Protein Kinase C α/δ (PKCα/δ) Enhances Cell Motility

Overview

Journal J Biol Chem

Specialty Biochemistry

Date 2019 Jun 10

PMID 31177095

Citations 15

Authors

Yugo Mori

Kaoru Akita

Kazuki Ojima

Shungo Iwamoto

Tomoko Yamashita

Eiichi Morii

Hiroshi Nakada

Affiliations

Soon will be listed here.

Abstract

Dysfunction of tight junctions is a critical step during the initial stage of tumor progression. Trophoblast cell surface antigen 2 (Trop-2) belongs to the family of tumor-associated calcium signal transducer () and is required for the stability of claudin-7 and claudin-1, which are often dysregulated or lost in carcinogenesis. Here, we investigated the effects of Trop-2 phosphorylation on cell motility. Analyses using HCT116 cells expressing WT Trop-2 (HCT116/WT) or Trop-2 alanine-substituted at Ser-303 (HCT116/S303A) or Ser-322 (HCT116/S322A) revealed that Trop-2 is phosphorylated at Ser-322. Furthermore, coimmunoprecipitation and Transwell assays indicated that Trop-2 S322A interacted with claudin-7 the strongest, and a phosphomimetic variant, Trop-2 S322E, the weakest and that HCT116/S322E cells have the highest motility and HCT116/S322A cells the lowest. All cell lines had similar levels of mRNA, but levels of claudin-7 protein were markedly decreased in the HCT116/S322E cells, suggesting posttranscriptional control of claudin-7. Moreover, claudin-7 was clearly localized to cell-cell borders in HCT116/S322A cells but was diffusely distributed on the membrane and partially localized in the cytoplasm of HCT116/S322E and HCT116/WT cells. These observations suggested that Trop-2 phosphorylation plays a role in the decrease or mislocalization of claudin-7. Using protein kinase C (PKC) inhibitors and PKC-specific siRNAs, we found that PKCα and PKCδ are responsible for Trop-2 phosphorylation. Of note, chemical PKC inhibition and PKCα- and PKCδ-specific siRNAs reduced motility. In summary, our findings provide evidence that Trop-2 is phosphorylated at Ser-322 by PKCα/δ and that this phosphorylation enhances cell motility and decreases claudin-7 localization to cellular borders.

Citing Articles

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Trop2-targeted therapy in breast cancer.

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The Relationship between Trop-2, Chemotherapeutic Drugs, and Chemoresistance.

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References

Tsujikawa M, Kurahashi H, Tanaka T, Nishida K, Shimomura Y, Tano Y . Identification of the gene responsible for gelatinous drop-like corneal dystrophy. Nat Genet. 1999; 21(4):420-3. DOI: 10.1038/7759. View

Gastl G, Spizzo G, Obrist P, Dunser M, Mikuz G . Ep-CAM overexpression in breast cancer as a predictor of survival. Lancet. 2000; 356(9246):1981-2. DOI: 10.1016/S0140-6736(00)03312-2. View

Moustafa A, Alaoui-Jamali M, Batist G, Hernandez-Perez M, Serruya C, Alpert L . Identification of genes associated with head and neck carcinogenesis by cDNA microarray comparison between matched primary normal epithelial and squamous carcinoma cells. Oncogene. 2002; 21(17):2634-40. DOI: 10.1038/sj.onc.1205351. View

Kleiger G, Grothe R, Mallick P, Eisenberg D . GXXXG and AXXXA: common alpha-helical interaction motifs in proteins, particularly in extremophiles. Biochemistry. 2002; 41(19):5990-7. DOI: 10.1021/bi0200763. View

Kominsky S, Argani P, Korz D, Evron E, Raman V, Garrett E . Loss of the tight junction protein claudin-7 correlates with histological grade in both ductal carcinoma in situ and invasive ductal carcinoma of the breast. Oncogene. 2003; 22(13):2021-33. DOI: 10.1038/sj.onc.1206199. View

Rangel L, Agarwal R, DSouza T, Pizer E, Alo P, Lancaster W . Tight junction proteins claudin-3 and claudin-4 are frequently overexpressed in ovarian cancer but not in ovarian cystadenomas. Clin Cancer Res. 2003; 9(7):2567-75. View

Tinsley J, Teasdale N, Yuan S . Involvement of PKCdelta and PKD in pulmonary microvascular endothelial cell hyperpermeability. Am J Physiol Cell Physiol. 2003; 286(1):C105-11. DOI: 10.1152/ajpcell.00340.2003. View

Michl P, Barth C, Buchholz M, Lerch M, Rolke M, Holzmann K . Claudin-4 expression decreases invasiveness and metastatic potential of pancreatic cancer. Cancer Res. 2003; 63(19):6265-71. View

Went P, Lugli A, Meier S, Bundi M, Mirlacher M, Sauter G . Frequent EpCam protein expression in human carcinomas. Hum Pathol. 2004; 35(1):122-8. DOI: 10.1016/j.humpath.2003.08.026. View

10.

Seligson D, Pantuck A, Liu X, Huang Y, Horvath S, Bui M . Epithelial cell adhesion molecule (KSA) expression: pathobiology and its role as an independent predictor of survival in renal cell carcinoma. Clin Cancer Res. 2004; 10(8):2659-69. DOI: 10.1158/1078-0432.ccr-1132-03. View

11.

Koivunen J, Aaltonen V, Koskela S, Lehenkari P, Laato M, Peltonen J . Protein kinase C alpha/beta inhibitor Go6976 promotes formation of cell junctions and inhibits invasion of urinary bladder carcinoma cells. Cancer Res. 2004; 64(16):5693-701. DOI: 10.1158/0008-5472.CAN-03-3511. View

12.

Schneider D, Engelman D . Motifs of two small residues can assist but are not sufficient to mediate transmembrane helix interactions. J Mol Biol. 2004; 343(4):799-804. DOI: 10.1016/j.jmb.2004.08.083. View

13.

Usami Y, Chiba H, Nakayama F, Ueda J, Matsuda Y, Sawada N . Reduced expression of claudin-7 correlates with invasion and metastasis in squamous cell carcinoma of the esophagus. Hum Pathol. 2006; 37(5):569-77. DOI: 10.1016/j.humpath.2005.12.018. View

14.

Olsen J, Blagoev B, Gnad F, Macek B, Kumar C, Mortensen P . Global, in vivo, and site-specific phosphorylation dynamics in signaling networks. Cell. 2006; 127(3):635-48. DOI: 10.1016/j.cell.2006.09.026. View

15.

Takaoka M, Nakamura T, Ban Y, Kinoshita S . Phenotypic investigation of cell junction-related proteins in gelatinous drop-like corneal dystrophy. Invest Ophthalmol Vis Sci. 2007; 48(3):1095-101. DOI: 10.1167/iovs.06-0740. View

16.

Wang J, Day R, Dong Y, Weintraub S, Michel L . Identification of Trop-2 as an oncogene and an attractive therapeutic target in colon cancers. Mol Cancer Ther. 2008; 7(2):280-5. DOI: 10.1158/1535-7163.MCT-07-2003. View

17.

Cubas R, Li M, Chen C, Yao Q . Trop2: a possible therapeutic target for late stage epithelial carcinomas. Biochim Biophys Acta. 2009; 1796(2):309-14. DOI: 10.1016/j.bbcan.2009.08.001. View

18.

Bignotti E, Todeschini P, Calza S, Falchetti M, Ravanini M, Tassi R . Trop-2 overexpression as an independent marker for poor overall survival in ovarian carcinoma patients. Eur J Cancer. 2010; 46(5):944-53. DOI: 10.1016/j.ejca.2009.12.019. View

19.

Nakatsukasa M, Kawasaki S, Yamasaki K, Fukuoka H, Matsuda A, Tsujikawa M . Tumor-associated calcium signal transducer 2 is required for the proper subcellular localization of claudin 1 and 7: implications in the pathogenesis of gelatinous drop-like corneal dystrophy. Am J Pathol. 2010; 177(3):1344-55. PMC: 2928967. DOI: 10.2353/ajpath.2010.100149. View

20.

Bornholdt J, Friis S, Godiksen S, Poulsen S, Santoni-Rugiu E, Bisgaard H . The level of claudin-7 is reduced as an early event in colorectal carcinogenesis. BMC Cancer. 2011; 11:65. PMC: 3045986. DOI: 10.1186/1471-2407-11-65. View