The subcellular distribution of eukaryotic translation initiation factor, eIF-5A, in cultured cells.
To gain insight into the role of the eukaryotic translation initiation factor, eIF-5A, we investigated the subcellular distribution of this protein in several cultured cell types and at different stages of the cell cycle using a highly potent monospecific polyclonal antibody to eIF-5A. Studies using indirect immunofluorescence and confocal microscopy in conjunction with subcellular fractionation demonstrate that eIF-5A is primarily localized in the cytoplasm of cells. This cytoplasmic location of eIF-5A is not significantly altered in different stages of the cell cycle and the subcellular distribution pattern of eIF-5A is not changed by viral oncogene transformation. Cell fractionation experiments identified two populations of eIF-5A in the cytoplasm, a soluble fraction and a fraction bound to internal membranes. By double immunofluorescence staining with an antibody against calnexin, a resident protein of the endoplasmic reticulum (ER), we demonstrate that the membrane-bound fraction of eIF-5A colocalizes with the ER and not with the cytoskeleton. Expression of Rev, a regulatory protein of human immunodeficiency virus type 1 (HIV-1), does not alter the subcellular distribution of endogenous eIF-5A in these cells. eIF-5A is detected in all tissues and cells examined including extracts prepared from Xenopus oocytes. Our results indicate that eIF-5A is a ubiquitous cytoplasmic protein and suggest that a site of eIF-5A function is likely to be in association with the ER.
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Department of Biological Chemistry, Merck Research Laboratories, West Point, Pennsylvania 19486, USA.
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D016475:3T3 Cells / Q000737:chemistry / Q000648:ultrastructure*; D000163:Acquired Immunodeficiency Syndrome / Q000821:virology*; D000818:Animals; D000918:Antibody Specificity; D015153:Blotting, Western; D002453:Cell Cycle / Q000502:physiology; D003599:Cytoskeleton / Q000737:chemistry; D004721:Endoplasmic Reticulum / Q000737:chemistry; D015870:Gene Expression / Q000502:physiology; D015694:Gene Products, rev / Q000235:genetics; D015497:HIV-1 / Q000235:genetics*; D051379:Mice; D018613:Microscopy, Confocal; D010448:Peptide Initiation Factors / Q000032:analysis* / Q000276:immunology; D014176:Protein Biosynthesis / Q000502:physiology; D016601:RNA-Binding Proteins; D013347:Subcellular Fractions / Q000737:chemistry*; D014760:Viral Fusion Proteins / Q000235:genetics; D054321:rev Gene Products, Human Immunodeficiency Virus; D000097574:Eukaryotic Translation Initiation Factor 5A
D015694:Gene Products, rev; D010448:Peptide Initiation Factors; D016601:RNA-Binding Proteins; D014760:Viral Fusion Proteins; D054321:rev Gene Products, Human Immunodeficiency Virus; D000097574:Eukaryotic Translation Initiation Factor 5A