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BMC Cancer
Published in: BMC Cancer 1/2014

Open Access 01-12-2014 | Research article

Heat shock proteins HSP70 and MRJ cooperatively regulate cell adhesion and migration through urokinase receptor

Authors: Yuli Lin, Nana Peng, Hongqin Zhuang, Di Zhang, Yao Wang, Zi-Chun Hua

Published in: BMC Cancer | Issue 1/2014

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Abstract

Background

The urokinase-type plasminogen activator receptor (uPAR) is an important regulator of ECM proteolysis, cell-ECM interactions and cell signaling. uPAR and heat shock proteins HSP70 and MRJ (DNAJB6) have been implicated in tumor growth and metastasis. We have reported recently that MRJ (DNAJB6, a heat shock protein) can interact with uPAR and enhance cell adhesion. Here, we identified another heat shock protein HSP70 as a novel uPAR-interacting protein.

Methods

We performed co-immunoprecipitation in human embryonic kidney (HEK) 293 and colon cancer HCT116 cells as well as immunofluorence assays in HEK293 cells stably transfected with uPAR to investigate the association of suPAR with HSP70/MRJ. To understand the biological functions of the triple complex of suPAR/HSP70/MRJ, we determined whether HSP70 and/or MRJ regulated uPAR-mediated cell invasion, migration, adhesion to vitronectin and MAPK pathway in two pair of human tumor cells (uPAR negative HEK293 cells vs HEK293 cells stably transfected with uPAR and HCT116 cells stably transfected with antisense-uPAR vs HCT116 mock cells transfected with vector only) using transwell assay, wound healing assay, quantitative RT-PCR analyzing mmp2 and mmp9 transcription levels, cell adhesion assay and Western blotting assay.

Results

HSP70 and MRJ formed a triple complex with uPAR and over-expression of MRJ enhanced the interaction between HSP70 and uPAR, while knockdown of MRJ decreased soluble uPAR in HCT116 cells (P < 0.05) and reduced the formation of the triple complex, suggesting that MRJ may act as an uPAR-specific adaptor protein to link uPAR to HSP70. Further experiments showed that knockdown of HSP70 and/or MRJ by siRNA inhibited uPAR-mediated cell adhesion to vitronectin as well as suppressed cell invasion and migration. Knockdown of HSP70 and/or MRJ inhibited expression of invasion related genes mmp2 and mmp9. Finally, HSP70 and/or MRJ up-regulated phosphorylation levels of ERK1/2 and FAK suggesting MAPK pathway was involved. All the biological function experiments in cell level showed an additive effect when HSP70 and MRJ were regulated simultaneously indicating their collaborated regulation effects on uPAR.

Conclusions

These findings may offer a novel insight into the interactions between uPAR and HSP70/MRJ and their functions in cell adhesion and migration may provide more understanding of the roles in regulating cancer metastasis.
Appendix
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Literature
1.
Blasi F, Carmeliet P: uPAR: a versatile signalling orchestrator. Nat Rev Mol Cell Biol. 2002, 3 (12): 932-943. 10.1038/nrm977.CrossRefPubMed
2.
Cohen RL, Xi XP, Crowley CW, Lucas BK, Levinson AD, Shuman MA: Effects of urokinase receptor occupancy on plasmin generation and proteolysis of basement membrane by human tumor cells. Blood. 1991, 78 (2): 479-487.PubMed
3.
Zhou HM, Nichols A, Meda P, Vassalli JD: Urokinase-type plasminogen activator and its receptor synergize to promote pathogenic proteolysis. EMBO J. 2000, 19 (17): 4817-4826. 10.1093/emboj/19.17.4817.CrossRefPubMedPubMedCentral
4.
Wang YJDL, Johnson K, Selhamer JJ, Doe WF: Structure of the human urokinase receptor geneand its similarity to CD59 and the Ly-6 family. Eur J Biochem. 1994, 227 (94 1175/1): 6-
5.
Ploug M, Ellis V: Structure-function relationships in the receptor for urokinase-type plasminogen activator. Comparison to other members of the Ly-6 family and snake venom alpha-neurotoxins. FEBS Lett. 1994, 349 (2): 163-168. 10.1016/0014-5793(94)00674-1.CrossRefPubMed
6.
Smith HW, Marshall CJ: Regulation of cell signalling by uPAR. Nat Rev Mol Cell Biol. 2010, 11 (1): 23-36. 10.1038/nrm2821.CrossRefPubMed
7.
Ossowski L, Aguirre-Ghiso JA: Urokinase receptor and integrin partnership: coordination of signaling for cell adhesion, migration and growth. Curr Opin Cell Biol. 2000, 12 (5): 613-620. 10.1016/S0955-0674(00)00140-X.CrossRefPubMed
8.
Madsen CD, Sidenius N: The interaction between urokinase receptor and vitronectin in cell adhesion and signalling. Eur J Cell Biol. 2008, 87 (8–9): 617-629.CrossRefPubMed
9.
Thurison T, Lomholt AF, Rasch MG, Lund IK, Nielsen HJ, Christensen IJ, Hoyer-Hansen G: A new assay for measurement of the liberated domain I of the urokinase receptor in plasma improves the prediction of survival in colorectal cancer. Clin Chem. 2010, 56 (10): 1636-1640. 10.1373/clinchem.2010.144410.CrossRefPubMed
10.
Mustjoki S, Sidenius N, Sier CF, Blasi F, Elonen E, Alitalo R, Vaheri A: Soluble urokinase receptor levels correlate with number of circulating tumor cells in acute myeloid leukemia and decrease rapidly during chemotherapy. Cancer Res. 2000, 60 (24): 7126-7132.PubMed
11.
Chavakis T, Kanse SM, Yutzy B, Lijnen HR, Preissner KT: Vitronectin concentrates proteolytic activity on the cell surface and extracellular matrix by trapping soluble urokinase receptor-urokinase complexes. Blood. 1998, 91 (7): 2305-2312.PubMed
12.
Ploug M, Eriksen J, Plesner T, Hansen NE, Dano K: A soluble form of the glycolipid-anchored receptor for urokinase-type plasminogen activator is secreted from peripheral blood leukocytes from patients with paroxysmal nocturnal hemoglobinuria. Eur J Biochem. 1992, 208 (2): 397-404. 10.1111/j.1432-1033.1992.tb17200.x.CrossRefPubMed
13.
Connolly BM, Choi EY, Gardsvoll H, Bey AL, Currie BM, Chavakis T, Liu S, Molinolo A, Ploug M, Leppla SH, Buqqe TH: Selective abrogation of the uPA-uPAR interaction in vivo reveals a novel role in suppression of fibrin-associated inflammation. Blood. 2010, 116 (9): 1593-1603. 10.1182/blood-2010-03-276642.CrossRefPubMedPubMedCentral
14.
Wei C, Moller CC, Altintas MM, Li J, Schwarz K, Zacchigna S, Xie L, Henger A, Schmid H, Rastaldi MP, Cowan P, Kretzler M, Parrilla R, Bendayan M, Gupta V, Nikolic B, Kalluri R, Carmeliet P, Mundel P, Reiser J: Modification of kidney barrier function by the urokinase receptor. Nat Med. 2008, 14 (1): 55-63. 10.1038/nm1696.CrossRefPubMed
15.
Stephens RW, Nielsen HJ, Christensen IJ, Thorlacius-Ussing O, Sorensen S, Dano K, Brunner N: Plasma urokinase receptor levels in patients with colorectal cancer: relationship to prognosis. J Natl Cancer Inst. 1999, 91 (10): 869-874. 10.1093/jnci/91.10.869.CrossRefPubMed
16.
Konno H, Baba M, Shoji T, Ohta M, Suzuki S, Nakamura S: Cyclooxygenase-2 expression correlates with uPAR levels and is responsible for poor prognosis of colorectal cancer. Clin Exp Metastasis. 2002, 19 (6): 527-534. 10.1023/A:1020392309715.CrossRefPubMed
17.
Wei C, El Hindi S, Li J, Fornoni A, Goes N, Sageshima J, Maiguel D, Karumanchi SA, Yap HK, Saleem M, Zhang Q, Nikolic B, Chaudhuri A, Daftarian P, Salido E, Torres A, Salifu M, Sarwal MM, Schaefer F, Morath C, Schwenger V, Zeier M, Gupta V, Roth D, Rastaldi MP, Burke G, Ruiz P, Reiser J: Circulating urokinase receptor as a cause of focal segmental glomerulosclerosis. Nat Med. 2011, 17 (8): 952-960. 10.1038/nm.2411.CrossRefPubMedPubMedCentral
18.
Riisbro R, Christensen IJ, Piironen T, Greenall M, Larsen B, Stephens RW, Han C, Hoyer-Hansen G, Smith K, Brunner N, Harris AL: Prognostic significance of soluble urokinase plasminogen activator receptor in serum and cytosol of tumor tissue from patients with primary breast cancer. Clin Cancer Res. 2002, 8 (5): 1132-1141.PubMed
19.
Carmeliet P, Moons L, Lijnen R, Baes M, Lemaitre V, Tipping P, Drew A, Eeckhout Y, Shapiro S, Lupu F, Collen D: Urokinase-generated plasmin activates matrix metalloproteinases during aneurysm formation. Nat Genet. 1997, 17 (4): 439-444. 10.1038/ng1297-439.CrossRefPubMed
20.
Wei Y, Lukashev M, Simon DI, Bodary SC, Rosenberg S, Doyle MV, Chapman HA: Regulation of integrin function by the urokinase receptor. Science. 1996, 273 (5281): 1551-1555. 10.1126/science.273.5281.1551.CrossRefPubMed
21.
Madsen CD, Ferraris GM, Andolfo A, Cunningham O, Sidenius N: uPAR-induced cell adhesion and migration: vitronectin provides the key. J Cell Biol. 2007, 177 (5): 927-939. 10.1083/jcb.200612058.CrossRefPubMedPubMedCentral
22.
Wei Y, Waltz DA, Rao N, Drummond RJ, Rosenberg S, Chapman HA: Identification of the urokinase receptor as an adhesion receptor for vitronectin. J Biol Chem. 1994, 269 (51): 32380-32388.PubMed
23.
De Bock CE, Lin Z, Mekkawy AH, Byrne JA, Wang Y: Interaction between urokinase receptor and heat shock protein MRJ enhances cell adhesion. Int J Oncol. 2010, 36 (5): 1155-1163.PubMed
24.
Dai YS, Xu J, Molkentin JD: The DnaJ-related factor Mrj interacts with nuclear factor of activated T cells c3 and mediates transcriptional repression through class II histone deacetylase recruitment. Mol Cell Biol. 2005, 25 (22): 9936-9948. 10.1128/MCB.25.22.9936-9948.2005.CrossRefPubMedPubMedCentral
25.
Kelley WL: The J-domain family and the recruitment of chaperone power. Trends Biochem Sci. 1998, 23 (6): 222-227. 10.1016/S0968-0004(98)01215-8.CrossRefPubMed
26.
Calderwood SK, Khaleque MA, Sawyer DB, Ciocca DR: Heat shock proteins in cancer: chaperones of tumorigenesis. Trends Biochem Sci. 2006, 31 (3): 164-172. 10.1016/j.tibs.2006.01.006.CrossRefPubMed
27.
Wang Y, Liang X, Wu S, Murrell GA, Doe WF: Inhibition of colon cancer metastasis by a 3’- end antisense urokinase receptor mRNA in a nude mouse model. Int J Cancer. 2001, 92 (2): 257-262. 10.1002/1097-0215(200102)9999:9999<::AID-IJC1178>3.0.CO;2-6.CrossRefPubMed
28.
Zhuang H, Jiang W, Zhang X, Qiu F, Gan Z, Cheng W, Zhang J, Guan S, Tang B, Huang Q, Wu X, Huang X, Jiang W, Hu Q, Lu M, Hua ZC: Suppression of HSP70 expression sensitizes NSCLC cell lines to TRAIL-induced apoptosis by upregulating DR4 and DR5 and downregulating c-FLIP-L expressions. J Mol Med (Berl). 2013, 91 (2): 219-235. 10.1007/s00109-012-0947-3.CrossRef
29.
Liang X, Yang X, Tang Y, Zhou H, Liu X, Xiao L, Gao J, Mao Z: RNAi-mediated downregulation of urokinase plasminogen activator receptor inhibits proliferation, adhesion, migration and invasion in oral cancer cells. Oral Oncol. 2008, 44 (12): 1172-1180. 10.1016/j.oraloncology.2008.03.004.CrossRefPubMed
30.
Gondi CS, Lakka SS, Dinh DH, Olivero WC, Gujrati M, Rao JS: RNAi-mediated inhibition of cathepsin B and uPAR leads to decreased cell invasion, angiogenesis and tumor growth in gliomas. Oncogene. 2004, 23 (52): 8486-8496. 10.1038/sj.onc.1207879.CrossRefPubMed
31.
Zhou H, Tang Y, Liang X, Yang X, Yang J, Zhu G, Zheng M, Zhang C: RNAi targeting urokinase-type plasminogen activator receptor inhibits metastasis and progression of oral squamous cell carcinoma in vivo. Int J Cancer. 2009, 125 (2): 453-462. 10.1002/ijc.24360.CrossRefPubMed
32.
Gondi CS, Kandhukuri N, Dinh DH, Gujrati M, Rao JS: Down-regulation of uPAR and uPA activates caspase-mediated apoptosis and inhibits the PI3K/AKT pathway. Int J Oncol. 2007, 31 (1): 19-27.PubMedPubMedCentral
33.
Vial E, Sahai E, Marshall CJ: ERK-MAPK signaling coordinately regulates activity of Rac1 and RhoA for tumor cell motility. Cancer Cell. 2003, 4 (1): 67-79. 10.1016/S1535-6108(03)00162-4.CrossRefPubMed
34.
Nguyen DH, Catling AD, Webb DJ, Sankovic M, Walker LA, Somlyo AV, Weber MJ, Gonias SL: Myosin light chain kinase functions downstream of Ras/ERK to promote migration of urokinase-type plasminogen activator-stimulated cells in an integrin-selective manner. J Cell Biol. 1999, 146 (1): 149-164. 10.1083/jcb.146.1.149.CrossRefPubMedPubMedCentral
35.
Jo M, Thomas KS, O'Donnell DM, Gonias SL: Epidermal growth factor receptor-dependent and -independent cell-signaling pathways originating from the urokinase receptor. J Biol Chem. 2003, 278 (3): 1642-1646. 10.1074/jbc.M210877200.CrossRefPubMed
36.
Nguyen DH, Webb DJ, Catling AD, Song Q, Dhakephalkar A, Weber MJ, Ravichandran KS, Gonias SL: Urokinase-type plasminogen activator stimulates the Ras/Extracellular signal-regulated kinase (ERK) signaling pathway and MCF-7 cell migration by a mechanism that requires focal adhesion kinase, Src, and Shc. Rapid dissociation of GRB2/Sps-Shc complex is associated with the transient phosphorylation of ERK in urokinase-treated cells. J Biol Chem. 2000, 275 (25): 19382-19388. 10.1074/jbc.M909575199.CrossRefPubMed
37.
Doshi BM, Hightower LE, Lee J: The role of Hsp27 and actin in the regulation of movement in human cancer cells responding to heat shock. Cell Stress Chaperones. 2009, 14 (5): 445-457. 10.1007/s12192-008-0098-1.CrossRefPubMedPubMedCentral
38.
Olson D, Pollanen J, Hoyer-Hansen G, Ronne E, Sakaguchi K, Wun TC, Appella E, Dano K, Blasi F: Internalization of the urokinase-plasminogen activator inhibitor type-1 complex is mediated by the urokinase receptor. J Biol Chem. 1992, 267 (13): 9129-9133.PubMed
39.
Cubellis MV, Wun TC, Blasi F: Receptor-mediated internalization and degradation of urokinase is caused by its specific inhibitor PAI-1. Embo J. 1990, 9 (4): 1079-1085.PubMedPubMedCentral
40.
Fazioli F, Blasi F: Urokinase-type plasminogen activator and its receptor: new targets for anti-metastatic therapy?. Trends Pharmacol Sci. 1994, 15 (1): 25-29. 10.1016/0165-6147(94)90130-9.CrossRefPubMed
41.
Arispe N, Doh M, De Maio A: Lipid interaction differentiates the constitutive and stress-induced heat shock proteins Hsc70 and Hsp70. Cell Stress Chaperones. 2002, 7 (4): 330-338. 10.1379/1466-1268(2002)007<0330:LIDTCA>2.0.CO;2.CrossRefPubMedPubMedCentral
42.
Kanse SM, Kost C, Wilhelm OG, Andreasen PA, Preissner KT: The urokinase receptor is a major vitronectin-binding protein on endothelial cells. Exp Cell Res. 1996, 224 (2): 344-353. 10.1006/excr.1996.0144.CrossRefPubMed
43.
Waltz DA, Natkin LR, Fujita RM, Wei Y, Chapman HA: Plasmin and plasminogen activator inhibitor type 1 promote cellular motility by regulating the interaction between the urokinase receptor and vitronectin. J Clin Invest. 1997, 100 (1): 58-67. 10.1172/JCI119521.CrossRefPubMedPubMedCentral
44.
Stahl A, Mueller BM: Melanoma cell migration on vitronectin: regulation by components of the plasminogen activation system. Int J Cancer. 1997, 71 (1): 116-122. 10.1002/(SICI)1097-0215(19970328)71:1<116::AID-IJC19>3.0.CO;2-G.CrossRefPubMed
45.
Sitrin RG, Todd RF, Albrecht E, Gyetko MR: The urokinase receptor (CD87) facilitates CD11b/CD18-mediated adhesion of human monocytes. J Clin Invest. 1996, 97 (8): 1942-1951. 10.1172/JCI118626.CrossRefPubMedPubMedCentral
46.
Boroughs LK, Antonyak MA, Johnson JL, Cerione RA: A unique role for heat shock protein 70 and its binding partner tissue transglutaminase in cancer cell migration. J Biol Chem. 2011, 286 (43): 37094-37107. 10.1074/jbc.M111.242438.CrossRefPubMedPubMedCentral
47.
Sims JD, McCready J, Jay DG: Extracellular heat shock protein (Hsp)70 and Hsp90alpha assist in matrix metalloproteinase-2 activation and breast cancer cell migration and invasion. PLoS One. 2011, 6 (4): e18848-10.1371/journal.pone.0018848.CrossRefPubMedPubMedCentral
48.
Sidenius N, Blasi F: The urokinase plasminogen activator system in cancer: recent advances and implication for prognosis and therapy. Cancer Metastasis Rev. 2003, 22 (2–3): 205-222.CrossRefPubMed
Metadata
Title
Heat shock proteins HSP70 and MRJ cooperatively regulate cell adhesion and migration through urokinase receptor
Authors
Yuli Lin
Nana Peng
Hongqin Zhuang
Di Zhang
Yao Wang
Zi-Chun Hua
Publication date
01-12-2014
Publisher
BioMed Central
Published in
BMC Cancer / Issue 1/2014
Electronic ISSN: 1471-2407
DOI
https://doi.org/10.1186/1471-2407-14-639

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