Abstract
Salvador promotes both cell cycle exit and apoptosis through the modulation of both cyclin E and Drosophila inhibitor of apoptosis protein in Drosophila. However, the cellular function of human Salvador (hSav1) is rarely reported. To screen for novel binding proteins that interact with hSav1, the cDNA of hSav1 was cloned into a bait protein plasmid, and positive clones were screened from a human fetal liver cDNA library by the yeast two-hybrid system. hSav1 mRNA was expressed in yeast and there was no self-activation and toxicity in the yeast strain AH109. Twenty proteins were found to interact with hSav1, including HS1 (haematopoietic cell specific protein1)-associated protein X-1 (HAX-1); neural precursor cell expressed, developmentally down-regulated 9, pyruvate kinase, liver and RBC, cytochrome c oxidase subunit Vb, enoyl coenzyme A hydratase short chain 1, and NADH dehydrogenase (ubiquinone) 1 beta subcomplex, demonstrating that the yeast two-hybrid system is an efficient method for investigating protein interactions. Among the identified proteins, there were many mitochondrial proteins, indicating that hSav1 may play a role in mitochondrial function. We also confirmed the interaction of HAX-1 and hSav1 in mammalian cells. This investigation provides functional clues for further exploration of potential apoptosis-related proteins in disease biotherapy.
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Acknowledgments
This work was partly supported by National Natural Science Foundation of China (NSFC: No. 30872472, No. 30800569, No. 30973496), Natural Science Foundation of Hubei Province (2008CDB174 and 2009CDB239) and National Basic Research Program of China (973: No. 2009CB521802). We are grateful to prof. Ma ding’s laboratory for their technical assistance.
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11033_2012_1670_MOESM1_ESM.tif
Supplemental Fig. 1 The construction of bait plasmid pGBKT7-hSav1. lane 1: DNA Marker, lane 2: PCR product, lane 3: pGBKT7-hSav1 plasmid before digestion, lane 4: pGBKT7-hSav1 plasmid digested with restriction endonucleases (EcoRI and BamHI), lane 5: DNA marker) (TIFF 175 kb)
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Li, X., Luo, X., Li, Z. et al. Screening of binding proteins that interact with human Salvador 1 in a human fetal liver cDNA library by the yeast two-hybrid system. Mol Biol Rep 39, 8225–8230 (2012). https://doi.org/10.1007/s11033-012-1670-4
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DOI: https://doi.org/10.1007/s11033-012-1670-4