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Midkine binds to 37-kDa laminin binding protein precursor, leading to nuclear transport of the complex
R H Salama1, H Muramatsu, K Zou
1Department of Biochemistry, Nagoya University School of Medicine, 65 Tsurumai-cho, Nagoya, Showa-ku, 466-8550, Japan.
Abstract:
Midkine (MK) is a heparin binding multifunctional protein that promotes cell survival and cell migration. MK was found to bind to 37-kDa laminin binding protein precursor (LBP), a precursor of 67-kDa laminin receptor, with K(d) of 1.1 nM between MK and LBP-glutathione-S-transferase fusion protein. The binding was inhibited by laminin, anti-LBP, amyloid beta-peptide, and heparin; the latter two are known to bind to MK. In CMT-93 mouse rectal carcinoma cells, LBP was mostly located in the cytoplasm as revealed by immunostaining with anti-LBP antibody. That a portion of LBP or 67-kDa laminin receptor was located at the surface of these cells was verified by inhibition of cell attachment to laminin-coated dishes by anti-LBP antibody. When MK was added to culture medium of these cells, a part of LBP migrated to the nucleus. The movement occurred concomitantly with nuclear transport of biotin-labeled MK. These findings suggested that the binding of MK to LBP caused nuclear translocation of the molecular complex.
Insights
Midkine (MK) protein binds to laminin binding protein precursor (LBP). This interaction causes LBP to move into the nucleus within cancer cells, suggesting a novel molecular mechanism.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Midkine (MK) is a multifunctional protein involved in cell survival and migration.
- Laminin binding protein precursor (LBP) is a precursor to the 67-kDa laminin receptor.
- LBP is typically found in the cytoplasm, with some presence at the cell surface.
Purpose of the Study:
- To investigate the interaction between Midkine (MK) and laminin binding protein precursor (LBP).
- To determine the effect of MK binding on LBP localization within CMT-93 mouse rectal carcinoma cells.
Main Methods:
- Binding assays using MK and LBP-glutathione-S-transferase fusion protein.
- Immunostaining with anti-LBP antibody to determine LBP localization.
- Cell attachment assays to verify cell surface LBP.
- Observation of LBP and MK localization after MK addition to cell cultures.
Main Results:
- MK binds to LBP with high affinity (K(d) = 1.1 nM).
- Binding is inhibited by laminin, anti-LBP, amyloid beta-peptide, and heparin.
- In CMT-93 cells, MK addition induced nuclear translocation of LBP, coinciding with MK's nuclear transport.
- A portion of LBP was confirmed to be at the cell surface, mediating cell attachment to laminin.
Conclusions:
- Midkine directly interacts with laminin binding protein precursor.
- The MK-LBP interaction facilitates the translocation of LBP into the nucleus.
- This molecular complex movement suggests a novel signaling pathway involving MK and LBP.