IGFBP-5 Promotes Fibrosis Independently of Its Translocation to the Nucleus and Its Interaction with Nucleolin and IGF

Overview

Journal PLoS One

Specialties General Medicine
Science

Date 2015 Jun 24

PMID 26103640

Citations 12

Authors

Yunyun Su

Tetsuya Nishimoto

Carol Feghali-Bostwick

Affiliations

Soon will be listed here.

Abstract

Background: Insulin-like growth factor binding protein (IGFBP)-5 levels are increased in systemic sclerosis (SSc) skin and lung. We previously reported that IGFBP-5 is a pro-fibrotic factor that induces extracellular matrix (ECM) production and deposition. Since IGFBP-5 contains a nuclear localization signal (NLS) that facilitates its nuclear translocation, we sought to examine the role of nuclear translocation on the fibrotic activity of IGFBP-5 and identify IGFBP-5 binding partners relevant for its nuclear compartmentalization.

Methods: We generated functional wild type IGFBP-5 and IGFBP-5 with a mutated NLS or a mutated IGF binding site. Abrogation of nuclear translocation in the NLS mutant was confirmed using immunofluorescence and immunoblotting of nuclear and cytoplasmic cellular extracts. Abrogation of IGF binding was confirmed using western ligand blot. The fibrotic activity of wild type and mutant IGFBP-5 was examined in vitro in primary human fibroblasts and ex vivo in human skin. We identified IGFBP-5 binding partners using immunoprecipitation and mass spectrometry. We examined the effect of nucleolin on IGFBP-5 localization and function via sequence-specific silencing in primary human fibroblasts.

Results: Our results show that IGFBP-5-induced ECM production in vitro in primary human fibroblasts is independent of its nuclear translocation. The NLS-mutant also induced fibrosis ex vivo in human skin, thus confirming and extending the in vitro findings. Similar findings were obtained with the IGF-binding mutant. Nucleolin, a nucleolar protein that can serve as a nuclear receptor, was identified as an IGFBP-5 binding partner. Silencing nucleolin reduced IGFBP-5 translocation to the nucleus but did not block the ability of IGFBP-5 to induce ECM production and a fibrotic phenotype.

Conclusions: IGFBP-5 transport to the nucleus requires an intact NLS and nucleolin. However, nuclear translocation is not necessary for IGFBP-5 fibrotic activity; neither is IGF binding. Our data provide further insights into the role of cellular compartmentalization in IGFBP-5-induced fibrosis.

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References

Hermani A, Shukla A, Medunjanin S, Werner H, Mayer D . Insulin-like growth factor binding protein-4 and -5 modulate ligand-dependent estrogen receptor-α activation in breast cancer cells in an IGF-independent manner. Cell Signal. 2013; 25(6):1395-402. DOI: 10.1016/j.cellsig.2013.02.018. View

Parker A, Rees C, Clarke J, Busby Jr W, Clemmons D . Binding of insulin-like growth factor (IGF)-binding protein-5 to smooth-muscle cell extracellular matrix is a major determinant of the cellular response to IGF-I. Mol Biol Cell. 1998; 9(9):2383-92. PMC: 25505. DOI: 10.1091/mbc.9.9.2383. View

Perks C, McCaig C, Holly J . Differential insulin-like growth factor (IGF)-independent interactions of IGF binding protein-3 and IGF binding protein-5 on apoptosis in human breast cancer cells. Involvement of the mitochondria. J Cell Biochem. 2000; 80(2):248-58. DOI: 10.1002/1097-4644(20010201)80:2<248::aid-jcb140>3.0.co;2-4. View

Seinsoth S, Uhlmann-Schiffler H, Stahl H . Bidirectional DNA unwinding by a ternary complex of T antigen, nucleolin and topoisomerase I. EMBO Rep. 2003; 4(3):263-8. PMC: 1315898. DOI: 10.1038/sj.embor.embor770. View

Sureshbabu A, Okajima H, Yamanaka D, Tonner E, Shastri S, Maycock J . IGFBP5 induces cell adhesion, increases cell survival and inhibits cell migration in MCF-7 human breast cancer cells. J Cell Sci. 2012; 125(Pt 7):1693-705. DOI: 10.1242/jcs.092882. View

Schedlich L, Le Page S, Firth S, BRIGGS L, Jans D, BAXTER R . Nuclear import of insulin-like growth factor-binding protein-3 and -5 is mediated by the importin beta subunit. J Biol Chem. 2000; 275(31):23462-70. DOI: 10.1074/jbc.M002208200. View

Schedlich L, Young T, Firth S, BAXTER R . Insulin-like growth factor-binding protein (IGFBP)-3 and IGFBP-5 share a common nuclear transport pathway in T47D human breast carcinoma cells. J Biol Chem. 1998; 273(29):18347-52. DOI: 10.1074/jbc.273.29.18347. View

Nam T, Busby Jr W, Rees C, Clemmons D . Thrombospondin and osteopontin bind to insulin-like growth factor (IGF)-binding protein-5 leading to an alteration in IGF-I-stimulated cell growth. Endocrinology. 2000; 141(3):1100-6. DOI: 10.1210/endo.141.3.7386. View

Amaar Y, Thompson G, Linkhart T, Chen S, Baylink D, Mohan S . Insulin-like growth factor-binding protein 5 (IGFBP-5) interacts with a four and a half LIM protein 2 (FHL2). J Biol Chem. 2002; 277(14):12053-60. DOI: 10.1074/jbc.M110872200. View

10.

Bourbon H, Bugler B, Caizergues-Ferrer M, Amalric F . Role of phosphorylation on the maturation pathways of a 100 kDa nucleolar protein. FEBS Lett. 1983; 155(2):218-22. DOI: 10.1016/0014-5793(82)80606-6. View

11.

Santos A, Jung J, Aziz N, Kissil J, Pure E . Targeting fibroblast activation protein inhibits tumor stromagenesis and growth in mice. J Clin Invest. 2009; 119(12):3613-25. PMC: 2786791. DOI: 10.1172/JCI38988. View

12.

Barel M, Meibom K, Charbit A . Nucleolin, a shuttle protein promoting infection of human monocytes by Francisella tularensis. PLoS One. 2010; 5(12):e14193. PMC: 2995743. DOI: 10.1371/journal.pone.0014193. View

13.

Seurin D, Lombet A, Babajko S, Godeau F, Ricort J . Insulin-like growth factor binding proteins increase intracellular calcium levels in two different cell lines. PLoS One. 2013; 8(3):e59323. PMC: 3602172. DOI: 10.1371/journal.pone.0059323. View

14.

Lapeyre B, Bourbon H, Amalric F . Nucleolin, the major nucleolar protein of growing eukaryotic cells: an unusual protein structure revealed by the nucleotide sequence. Proc Natl Acad Sci U S A. 1987; 84(6):1472-6. PMC: 304456. DOI: 10.1073/pnas.84.6.1472. View

15.

Shi H, Huang Y, Zhou H, Song X, Yuan S, Fu Y . Nucleolin is a receptor that mediates antiangiogenic and antitumor activity of endostatin. Blood. 2007; 110(8):2899-906. DOI: 10.1182/blood-2007-01-064428. View

16.

Amaar Y, Baylink D, Mohan S . Ras-association domain family 1 protein, RASSF1C, is an IGFBP-5 binding partner and a potential regulator of osteoblast cell proliferation. J Bone Miner Res. 2005; 20(8):1430-9. PMC: 2897826. DOI: 10.1359/JBMR.050311. View

17.

Pilewski J, Liu L, Henry A, Knauer A, Feghali-Bostwick C . Insulin-like growth factor binding proteins 3 and 5 are overexpressed in idiopathic pulmonary fibrosis and contribute to extracellular matrix deposition. Am J Pathol. 2005; 166(2):399-407. PMC: 1602317. DOI: 10.1016/S0002-9440(10)62263-8. View

18.

Ma N, Matsunaga S, Takata H, Ono-Maniwa R, Uchiyama S, Fukui K . Nucleolin functions in nucleolus formation and chromosome congression. J Cell Sci. 2007; 120(Pt 12):2091-105. DOI: 10.1242/jcs.008771. View

19.

Overgaard M, Boldt H, Laursen L, Sottrup-Jensen L, Conover C, Oxvig C . Pregnancy-associated plasma protein-A2 (PAPP-A2), a novel insulin-like growth factor-binding protein-5 proteinase. J Biol Chem. 2001; 276(24):21849-53. DOI: 10.1074/jbc.M102191200. View

20.

Yasuoka H, Jukic D, Zhou Z, Choi A, Feghali-Bostwick C . Insulin-like growth factor binding protein 5 induces skin fibrosis: A novel murine model for dermal fibrosis. Arthritis Rheum. 2006; 54(9):3001-10. DOI: 10.1002/art.22084. View