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Colocalization and identification of interaction sites between IGFBP-3 and GalNAc-T14 |
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| Authors: | Wu Chen Ma Si-Si Ge Jian-Feng Wang Yuan-Yuan Tian Huan-Na Liu Xiao-Bo Zhang Bo Liu Fang-Ming Zhang Xiao-Kang Li Qin-Jian |
| Institution: | College of Life Sciences, Hebei University, Baoding, Hebei 071002, PR China. dawnwuchen@163.com |
| Abstract: | GalNAc-T14 was identified as a novel IGFBP-3 binding partner in previous studies. Here, we furtherly confirmed the interaction between them by confocal microscopy, and identified the binding domain and probable interaction sites of GalNAc-T14 with IGFBP-3. The result of subcellular localization indicated that GalNAc-T14 was distributed in the cytosol, whereas IGFBP-3 existed in the cytosol and nucleolus. Confocal analyses demonstrated that IGFBP-3 and GalNAc-T14 colocalized in the cytosol. The result from yeast two hybrid assay showed that the C terminus of GalNAc-T14 (408-552aa) was essential for the interaction between GalNAc-T14 and IGFBP-3, especially Tyr(408), Pro(409), and Glu(410) of GalNAc-T14 may play key roles in the interaction with IGFBP-3. In conclusion, these studies demonstrated that IGFBP-3 and GalNAc-T14 are colocalized in MCF-7 cells and confirmed the interaction between IGFBP-3 and GalNAc-T14. This interaction may play an important role in the functional regulation of IGFBP-3. |
| Keywords: | IGFBP-3 Insulin-like growth factor binding protein 3 IGFBP Insulin-like growth factor binding protein GalNAc-T14 N-acetylgalactosaminyltransferase 14 GalNAc-Tases N-acetylgalactosaminyltransferases IGF-independent Insulin-like growth factor-independent IGF-dependent Insulin-like growth factor-dependent MCF-7 human breast cancer cell line MCF-7 cells Rpb3 RNA polymerase II binding subunit 3 AMF/PGI autocrine motility factor/phosphoglucose isomerase LB Luria-Bertani AA Amino Acid E coli Escherichia coli YPD Yeast Extract Peptone Dextrose Medium DMEM Dulbecco modified Eagle medium FCS fetal calf serum 3-AT 3-amino-1 2 4-triazole SD/-Trp-Leu Complete Synthetic Defined Media without tryptophan and leucine SD/-Trp-Leu-His Complete Synthetic Defined Media without tryptophan leucine and histidine SD/-Trp Complete Synthetic Defined Media without Trp PBS Phosphate Buffered Saline GST Glutathione-S-transferase ORF open reading frame GT1 glucosyltransferase 1 |
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