Caifang Chen1, Yaoyao Shen1, Weiliang Shen2, Jing He1, Zhihua Lin3, Yinghui Dong4. 1. Zhejiang Key Laboratory of Aquatic Germplasm Resources, Zhejiang Wanli University, Ningbo, Zhejiang, 315100, People's Republic of China. 2. Ningbo Academy of Oceanology and Fishery, Ningbo, 315012, People's Republic of China. 3. Zhejiang Key Laboratory of Aquatic Germplasm Resources, Zhejiang Wanli University, Ningbo, Zhejiang, 315100, People's Republic of China. zhihua9988@126.com. 4. Zhejiang Key Laboratory of Aquatic Germplasm Resources, Zhejiang Wanli University, Ningbo, Zhejiang, 315100, People's Republic of China. dongyinghui118@126.com.
Abstract
BACKGROUND: Sulfide is a well-known toxicant widely distributed in the culture environment. As a representative burrowing benthic bivalve, the razor clam Sinonovacula constricta is highly sulfide tolerant. Mitochondrial sulfide oxidation is an important way for sulfide detoxification, where sulfur dioxygenase (SDO) is the second key enzyme. OBJECTIVE: To investigate the mechanism of sulfide tolerance in S. constricta, the molecular characterization of its SDO (designated as ScSDO) was studied. METHODS: The cDNA sequence of ScSDO was cloned by RACE technique. The response of ScSDO in gills and livers of S. constricta was investigated during sulfide exposure (50, 150, and 300 μM sulfide) for 0, 3, 6, 12, 24, 48, 72, and 96 h by qRT-PCR. Moreover, the temporal expression of ScSDO protein in S. constricta gills after exposure to 150 μM sulfide was detected by Western blot. The subcellular location of ScSDO was identified by TargetP 1.1 prediction and Western Blot analysis. RESULTS: The full-length cDNA of ScSDO was 2914 bp, encoding a protein of 304 amino acids. The deduced ScSDO protein was highly conserved, containing the signature HXHXDH motif of the metallo-β-lactamase superfamily and two metal-binding sites, of which metal-binding site I is known to be the catalytically active center. Subcellular localization confirmed that ScSDO was located only in the mitochondria. Responding to the sulfide exposure, distinct time-dependent increases in ScSDO expression were detected at both mRNA and protein levels. Moreover, the gills exhibited a higher ScSDO expression level than the livers. CONCLUSIONS: All of our results suggest that ScSDO plays an important role in mitochondrial sulfide oxidation during sulfide stress, making S. constricta highly sulfide tolerant. In addition, as a respiratory tissue, the gills play a more critical role in sulfide detoxification.
BACKGROUND: Sulfide is a well-known toxicant widely distributed in the culture environment. As a representative burrowing benthic bivalve, the razor clam Sinonovacula constricta is highly sulfide tolerant. Mitochondrial sulfide oxidation is an important way for sulfide detoxification, where sulfur dioxygenase (SDO) is the second key enzyme. OBJECTIVE: To investigate the mechanism of sulfide tolerance in S. constricta, the molecular characterization of its SDO (designated as ScSDO) was studied. METHODS: The cDNA sequence of ScSDO was cloned by RACE technique. The response of ScSDO in gills and livers of S. constricta was investigated during sulfide exposure (50, 150, and 300 μM sulfide) for 0, 3, 6, 12, 24, 48, 72, and 96 h by qRT-PCR. Moreover, the temporal expression of ScSDO protein in S. constricta gills after exposure to 150 μM sulfide was detected by Western blot. The subcellular location of ScSDO was identified by TargetP 1.1 prediction and Western Blot analysis. RESULTS: The full-length cDNA of ScSDO was 2914 bp, encoding a protein of 304 amino acids. The deduced ScSDO protein was highly conserved, containing the signature HXHXDH motif of the metallo-β-lactamase superfamily and two metal-binding sites, of which metal-binding site I is known to be the catalytically active center. Subcellular localization confirmed that ScSDO was located only in the mitochondria. Responding to the sulfide exposure, distinct time-dependent increases in ScSDO expression were detected at both mRNA and protein levels. Moreover, the gills exhibited a higher ScSDO expression level than the livers. CONCLUSIONS: All of our results suggest that ScSDO plays an important role in mitochondrial sulfide oxidation during sulfide stress, making S. constricta highly sulfide tolerant. In addition, as a respiratory tissue, the gills play a more critical role in sulfide detoxification.
Authors: Meghan M Holdorf; Heather A Owen; Sarah Rhee Lieber; Li Yuan; Nicole Adams; Carole Dabney-Smith; Christopher A Makaroff Journal: Plant Physiol Date: 2012-07-10 Impact factor: 8.340