Subcellular localization of 12(S)-hydroxy-5,8,10,14-eicosatetraenoic acid binding sites in Lewis lung carcinoma cells.
12(S)-Hydroxy-5,8,10,14-eicosatetraenoic acid (12(S)-HETE) stimulates both gene expression and cell surface expression of the heterodimeric integrin alpha IIb beta 3 in Lewis lung carcinoma cells. These cells contain high affinity binding sites which are specific for 12(S)-HETE. Analyses of the subcellular distribution and molecular size of these sites showed that cytosol was the fraction exhibiting the largest specific binding. On gel permeation chromatography the cytosolic 12(S)-HETE-binding component appeared to be slightly smaller than thyroglobulin (M(r) 669,000). The sedimentation coefficient (20.5 S, determined by sucrose density gradient centrifugation), on the other hand was 1 S unit higher than that of thyroglobulin. The radioactive material bound to the macromolecule was found to be unaltered 12(S)-HETE. Proteinase treatment disrupted the ligand/macromolecule complex, suggesting that a polypeptide component is essential. In addition to cytosol, mitochondria and nuclei also contained significant but lower amounts of specifically bound 12(S)-HETE. The biological significance of this is not clear, but the results are in agreement with observations that 12(S)-HETE exerts effects at several subcellular sites. Our results, to our knowledge for the first time, demonstrate a predominantly cytosolic localization of a recognition molecule for 12(S)-HETE. This localization is different from that of other eicosanoid receptors which are G-protein coupled plasma membrane proteins.
Herbertsson
H
H
Department of Cell Biology, Linköping University, Sweden.
Hammarström
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S
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D000818:Animals; D001665:Binding Sites; D018827:Carcinoma, Lewis Lung / Q000378:metabolism* / Q000648:ultrastructure; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D006893:Hydroxyeicosatetraenoic Acids / Q000378:metabolism*; D066298:In Vitro Techniques; D051379:Mice; D008928:Mitochondria / Q000378:metabolism; D014407:Tumor Cells, Cultured
D006893:Hydroxyeicosatetraenoic Acids; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid