Abstract
Furin is a member of the proprotein convertase family, which is capable of cleaving the precursors of a wide variety of substrates including membrane-type 1 matrix metalloproteinase (MT1-MMP) proenzyme. c-Src is activated by growth factors, and has been linked with a poor prognosis in pancreatic cancer (PCa). Both c-Src and Furin play crucial roles in tumorigenesis, and the mechanism controlling their association is not understood. Modulation of the association between Furin and pro-MT1-MMP by c-Src inhibitor PP2 was evaluated by western blotting, assay of in vitro enzyme, co-immunoprecipitation (co-IP), and confocal immunofluorescence microscopy. Human platelet-derived growth factor BB (PDGF-BB) activated c-Src and induced c-Src-dependent association of Furin with pro-MT1-MMP in HPAC pancreatic cancer cells. Co-IP and confocal immunofluorescence assays revealed that c-Src interacts with Furin in vivo. The SH2 domain appeared to be important for c-Src interaction with Furin. In addition, we showed that Furin protein is tyrosine phosphorylated. Association between Furin and MT1-MMP is regulated by the tyrosine kinase c-Src.
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Acknowledgment
This work was supported by a grant (30873033) from the National Natural Science Foundation of PR China and grants from Major State Basic Research Development Program(2009CB521800; 2010CB529400).
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Chong Shi and Yongchao Ma have contributed equally to this work.
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11010_2011_1128_MOESM1_ESM.tif
Fig. S1. Furin inhibitor reduces pro-MT1-MMP cleavage. HPAC cells were cultured in RPMI 1640 containing 10% FBS at 37°C. Cells were treated with control or Furin inhibitor I (10 μM) for 2 h and Western blot was performed to detect MT1-MMP cleavage. The level of mature form of MT1-MMP decreased significantly in Furin inhibitor I-treated cells. (TIFF 157 kb)
11010_2011_1128_MOESM2_ESM.tif
Fig. S2. Furin inhibitor reduced cell surface expression of MT1-MMP. Control and Furin inhibitor I-treated HPAC cells were incubated with anti-MT1-MMP anti-goat antibody, and stained with FITC-labeled anti-goat second antibody (Green). DAPI was used for nuclear staining. The cells were observed with a confocal microscope (Leica Microsystems, LAS AF-TCS SP5). Scale bars, 25 μM. (TIFF 434 kb)
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Shi, C., Ma, Y., Liu, H. et al. The non-receptor tyrosine kinase c-Src mediates the PDGF-induced association between Furin and pro-MT1-MMP in HPAC pancreatic cells. Mol Cell Biochem 362, 65–70 (2012). https://doi.org/10.1007/s11010-011-1128-3
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DOI: https://doi.org/10.1007/s11010-011-1128-3