Identification of a heat shock cognate protein 70 gene in Chinese soft-shell turtle (Pelodiscus sinensis) and its expression profiles under thermal stress

The heat shock cognate protein 70 (Hsc70) is a member of a 70-kDa heat shock protein (HSP70) family that functions as molecular chaperones.In this study,a novel Hsc70 gene from Chinese soft-shelled turtle (Pelodiscus sinensis) (tHsc70) was identified.The tHsc70 full-length complementary DNA (cDNA) is 2 272 bp long with a 1 941-bp open reading frame (ORF) encoding 646 amino acids.Three characteristic signature regions of the HSP70 family,two major domains of an adenosine triphosphate (ATP)/guanosine triphosphate (GTP) binding domain (ABD),and a substrate-binding domain (SBD) were present in the predicted tHsc70 amino acid sequence.The tHsc70 gene was expressed in Escherichia coli BL21 and the expression product reacted with the anti-Hsc70 mouse monoclonal antibody by Western blotting.Homology analysis revealed that tHsc70 shared identity from 53.9% to 87.7% at the nucleotide level,and 49.1% to 99.5% at the amino acid level with the known Hsc70s.Phylogenetic analysis showed that tHsc70 was clustered together with the Hsc70 gene of another reptile species (Alligator mississippiensis).The tHsc70 was expressed in the liver,lung,heart,and skeletal muscle.The expression patterns of tHsc70 messenger RNA (mRNA) differed among different tissues under different durations of heat stress at 40 °C.Adaptation at 25 °C for 1 h after heat stress was also different among tissues and length of heat stress.Irrespective of different profiles of expression under heat stress,tHsc70 may play roles in protecting turtles from thermal stress.

Stress protein expression in early phase spinal cord ischemia/reperfusion injury

Spinal cord ischemia/reperfusion injury is a stress injury to the spinal cord. Our previous studies using differential proteomics identified 21 differentially expressed proteins （n 〉 2） in rabbits with spinal cord ischemia/reperfusion injury. Of these proteins, stress-related proteins included protein disulfide isomerase A3, stress-induced-phosphoprotein 1 and heat shock cognate protein 70. In this study, we established New Zealand rabbit models of spinal cord ischemia/reperfusion injury by abdominal aorta occlusion. Results demonstrated that hind limb function initially improved after spinal cord ischemia/reperfusion injury, but then deteriorated. The pathological morphology of the spinal cord became aggravated, but lessened 24 hours after reperfusion. However, the numbers of motor neurons and interneurons in the spinal cord gradually decreased. The expression of protein disulfide isomerase A3, stress-induced-phosphoprotein 1 and heat shock cognate protein 70 was induced by ischemia/reperfusion injury. The expression of these proteins increased within 12 hours after reperfusion, and then decreased, reached a minimum at 24 hours, but subsequently increased again to similar levels seen at 6-12 hours, showing a characterization of induction-inhibition-induc- tion. These three proteins were expressed only in cytoplasm but not in the nuclei. Moreover, the expression was higher in interneurons than in motor neurons, and the survival rate of interneurons was greater than that of motor neurons. It is assumed that the expression of stress-related proteins exhibited a protective effect on neurons.