Identification and molecular analysis of a ferritin subunit from red drum (Sciaenops ocellatus).
Ferritin is a conserved iron binding protein existing ubiquitously in prokaryotes and eukaryotes. In this study, the gene encoding a ferritin M subunit homologue (SoFer1) was cloned from red drum (Sciaenops ocellatus) and analyzed at expression and functional levels. The open reading frame of SoFer1 is 531 bp and preceded by a 5'-untranslated region that contains a putative Iron Regulatory Element (IRE) preserved in many ferritins. The deduced amino acid sequence of SoFer1 possesses both the ferroxidase center of mammalian H ferritin and the iron nucleation site of mammalian L ferritin. Expression of SoFer1 was tissue specific and responded positively to experimental challenges with Gram-positive and Gram-negative fish pathogens. Treatment of red drum liver cells with iron, copper, and oxidant significantly upregulated the expression of SoFer1 in time-dependent manners. To further examine the potential role of SoFer1 in antioxidation, red drum liver cells transfected transiently with SoFer1 were prepared. Compared to control cells, SoFer1 transfectants exhibited reduced production of reactive oxygen species following H(2)O(2) challenge. Finally, to examine the iron binding potential of SoFer1, SoFer1 was expressed in and purified from Escherichia coli as a recombinant protein. Iron-chelating analysis showed that purified recombinant SoFer1 was capable of iron binding. Taken together, these results suggest that SoFer1 is likely to be a functional ferritin involved in iron sequestration, host immune defence against bacterial infection, and antioxidation.
Hu
Yong-hua
YH
Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, PR China.
Zheng
Wen-jiang
WJ
Sun
Li
L
D000595:Amino Acid Sequence; D000818:Animals; D001419:Bacteria; D001424:Bacterial Infections / Q000276:immunology / Q000662:veterinary; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005293:Ferritins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D005393:Fish Diseases / Q000276:immunology / Q000382:microbiology; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D007501:Iron / Q000378:metabolism; D008099:Liver / Q000276:immunology; D008969:Molecular Sequence Data; D018384:Oxidative Stress / Q000276:immunology; D010473:Perciformes / Q000145:classification / Q000235:genetics* / Q000276:immunology*; D010802:Phylogeny; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism; D016415:Sequence Alignment; D015854:Up-Regulation
D011994:Recombinant Proteins; D005293:Ferritins; D007501:Iron