Abstract
Quantification of secreted factors is most often measured with enzyme-linked immunosorbent assay (ELISA), Western Blot, or more recently with antibody arrays. However, some of these, like low-molecular-weight fibroblast growth factor-2 (LMW FGF-2; the 18 kDa form), exemplify a set of secreted but almost non-diffusible molecular actors. It has been proposed that phosphorylated FGF-2 is secreted via a non-vesicular mechanism and that heparan sulfate proteoglycans function as extracellular reservoir but also as actors for its secretion. Heparan sulfate is a linear sulfated polysaccharide present on proteoglycans found in the extracellular matrix or anchored in the plasma membrane (syndecan). Moreover the LMW FGF-2 secretion appears to be activated upon FGF-1 treatment. In order to estimate quantification of such factor export across the plasma membrane, technical approaches are presented (evaluation of LMW FGF-2: (1) secretion, (2) extracellular matrix reservoir, and (3) secretion modulation by surrounding factors) and the importance of such procedures in the comprehension of the biology of these growth factors is underlined.
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References
Touriol C, Bornes S, Bonnal S, Audigier S, Prats H, Prats AC, Vagner S (2003) Generation of protein isoform diversity by alternative initiation of translation at non-AUG codons. Biol Cell 95(3-4):169–178
Bikfalvi A, Klein S, Pintucci G, Rifkin DB (1997) Biological roles of fibroblast growth factor-2. Endocr Rev 18(1):26–45. doi:10.1210/edrv.18.1.0292
Schafer T, Zentgraf H, Zehe C, Brugger B, Bernhagen J, Nickel W (2004) Unconventional secretion of fibroblast growth factor 2 is mediated by direct translocation across the plasma membrane of mammalian cells. J Biol Chem 279(8):6244–6251. doi:10.1074/jbc.M310500200
Zehe C, Engling A, Wegehingel S, Schafer T, Nickel W (2006) Cell-surface heparan sulfate proteoglycans are essential components of the unconventional export machinery of FGF-2. Proc Natl Acad Sci U S A 103(42):15479–15484. doi:10.1073/pnas.0605997103
Ebert AD, Laussmann M, Wegehingel S, Kaderali L, Erfle H, Reichert J, Lechner J, Beer HD, Pepperkok R, Nickel W (2010) Tec-kinase-mediated phosphorylation of fibroblast growth factor 2 is essential for unconventional secretion. Traffic 11(6):813–826. doi:10.1111/j.1600-0854.2010.01059.x
Meunier S, Navarro MG, Bossard C, Laurell H, Touriol C, Lacazette E, Prats H (2009) Pivotal role of translokin/CEP57 in the unconventional secretion versus nuclear translocation of FGF2. Traffic 10(12):1765–1772. doi:10.1111/j.1600-0854.2009.00985.x
Szebenyi G, Fallon JF (1999) Fibroblast growth factors as multifunctional signaling factors. Int Rev Cytol 185:45–106
Trudel C, Faure-Desire V, Florkiewicz RZ, Baird A (2000) Translocation of FGF2 to the cell surface without release into conditioned media. J Cell Physiol 185(2):260–268. doi:10.1002/1097-652(200011)185:2<260::AID-JCP11>3.0.CO;2-X
Batchelor TT, Sorensen AG, di Tomaso E, Zhang WT, Duda DG, Cohen KS, Kozak KR, Cahill DP, Chen PJ, Zhu M, Ancukiewicz M, Mrugala MM, Plotkin S, Drappatz J, Louis DN, Ivy P, Scadden DT, Benner T, Loeffler JS, Wen PY, Jain RK (2007) AZD2171, a pan-VEGF receptor tyrosine kinase inhibitor, normalizes tumor vasculature and alleviates edema in glioblastoma patients. Cancer Cell 11(1):83–95. doi:10.1016/j.ccr.2006.11.021
Cenni E, Perut F, Granchi D, Avnet S, Amato I, Brandi ML, Giunti A, Baldini N (2007) Inhibition of angiogenesis via FGF-2 blockage in primitive and bone metastatic renal cell carcinoma. Anticancer Res 27(1A):315–319
Kopetz S, Hoff PM, Morris JS, Wolff RA, Eng C, Glover KY, Adinin R, Overman MJ, Valero V, Wen S, Lieu C, Yan S, Tran HT, Ellis LM, Abbruzzese JL, Heymach JV (2010) Phase II trial of infusional fluorouracil, irinotecan, and bevacizumab for metastatic colorectal cancer: efficacy and circulating angiogenic biomarkers associated with therapeutic resistance. J Clin Oncol 28(3):453–459. doi:10.1200/JCO.2009.24.8252
Porta C, Paglino C, Imarisio I, Ganini C, Sacchi L, Quaglini S, Giunta V, De Amici M (2013) Changes in circulating pro-angiogenic cytokines, other than VEGF, before progression to sunitinib therapy in advanced renal cell carcinoma patients. Oncology 84(2):115–122. doi:10.1159/000342099
Sharpe K, Stewart GD, Mackay A, Van Neste C, Rofe C, Berney D, Kayani I, Bex A, Wan E, O'Mahony FC, O'Donnell M, Chowdhury S, Doshi R, Ho-Yen C, Gerlinger M, Baker D, Smith N, Davies B, Sahdev A, Boleti E, De Meyer T, Van Criekinge W, Beltran L, Lu YJ, Harrison DJ, Reynolds AR, Powles T (2013) The effect of VEGF-targeted therapy on biomarker expression in sequential tissue from patients with metastatic clear cell renal cancer. Clin Cancer Res 19(24):6924–6934. doi:10.1158/1078-0432.CCR-13-1631
Arcondeguy T, Lacazette E, Millevoi S, Prats H, Touriol C (2013) VEGF-A mRNA processing, stability and translation: a paradigm for intricate regulation of gene expression at the post-transcriptional level. Nucleic Acids Res 41(17):7997–8010. doi:10.1093/nar/gkt539
Rapraeger AC, Krufka A, Olwin BB (1991) Requirement of heparan sulfate for bFGF-mediated fibroblast growth and myoblast differentiation. Science 252(5013):1705–1708
Rifkin DB, Quarto N, Mignatti P, Bizik J, Moscatelli D (1991) New observations on the intracellular localization and release of bFGF. Ann N Y Acad Sci 638:204–206
Haugsten EM, Sorensen V, Brech A, Olsnes S, Wesche J (2005) Different intracellular trafficking of FGF1 endocytosed by the four homologous FGF receptors. J Cell Sci 118(Pt 17):3869–3881. doi:10.1242/jcs.02509
Farley JR, Nakayama G, Cryns D, Segel IH (1978) Adenosine triphosphate sulfurylase from Penicillium chrysogenum equilibrium binding, substrate hydrolysis, and isotope exchange studies. Arch Biochem Biophys 185(2):376–390
Guimond S, Maccarana M, Olwin BB, Lindahl U, Rapraeger AC (1993) Activating and inhibitory heparin sequences for FGF-2 (basic FGF). Distinct requirements for FGF-1, FGF-2, and FGF-4. J Biol Chem 268(32):23906–23914
Vlodavsky I, Miao HQ, Medalion B, Danagher P, Ron D (1996) Involvement of heparan sulfate and related molecules in sequestration and growth promoting activity of fibroblast growth factor. Cancer Metastasis Rev 15(2):177–186
Acknowledgements
This work was supported by “La ligue contre le cancer” (2FI10869NEBD).
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Arcondéguy, T., Touriol, C., Lacazette, E. (2016). Quantification of a Non-conventional Protein Secretion: The Low-Molecular-Weight FGF-2 Example. In: Pompa, A., De Marchis, F. (eds) Unconventional Protein Secretion. Methods in Molecular Biology, vol 1459. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3804-9_8
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DOI: https://doi.org/10.1007/978-1-4939-3804-9_8
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