Identification of 12-lipoxygenase interaction with cellular proteins by yeast two-hybrid screening.
The platelet isoform of 12-lipoxygenase (12-LOX) is expressed in a variety of human tumors. 12-LOX metabolizes arachidonic acid to 12(S)-hydroxyeicosateraenoic acid (12(S)-HETE), which induces a number of cellular responses associated with tumor progression and metastasis. Little is known about 12-LOX regulation and no direct regulators of 12-LOX activity have been identified. To identify potential regulators of 12-LOX, we isolated cDNAs encoding 12-LOX interacting proteins using the yeast two-hybrid system. We screened a yeast two-hybrid interaction library from human epidermoid carcinoma A431 cells and identified four cellular proteins that interact specifically with 12-LOX. We identified type II keratin 5, lamin A, the cytoplasmic domain of integrin beta4 subunit and a phosphoprotein C8FW as 12-LOX interacting proteins. Here, we demonstrated that keratin 5, a 58 kD protein required for formation of 8 nm intermediate filaments, binds to 12-LOX in human tumor cells and may contribute to the regulated trafficking of 12-LOX. We also showed that lamin A binds 12-LOX in human tumor cells. These proteins provide the first candidate regulators of 12-LOX.
Tang
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Department of Radiation Oncology and Pathology, Center for Molecular Medicine and Genetics, Wayne State University School of Medicine, and Karmanos Cancer Institute, Detroit, Michigan 48202, USA.
Finley
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D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism; D001092:Arachidonate 12-Lipoxygenase / Q000097:blood / Q000235:genetics* / Q000378:metabolism*; D001792:Blood Platelets / Q000201:enzymology; D003001:Cloning, Molecular; D015723:Gene Library; D006801:Humans; D039663:Integrin beta4; D016023:Integrins / Q000235:genetics / Q000378:metabolism; D007527:Isoenzymes / Q000097:blood / Q000235:genetics / Q000378:metabolism; D007633:Keratins / Q000235:genetics / Q000378:metabolism; D034904:Lamin Type A; D034882:Lamins; D009363:Neoplasm Proteins / Q000235:genetics* / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014407:Tumor Cells, Cultured; D020798:Two-Hybrid System Techniques
D015703:Antigens, CD; D039663:Integrin beta4; D016023:Integrins; D007527:Isoenzymes; D034904:Lamin Type A; D034882:Lamins; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D007633:Keratins; D001092:Arachidonate 12-Lipoxygenase