Cloning And Functional Analysis Of ACBP1 Gene From Puccinellia Tenuiflora

Acyl-CoA-binding proteins(ACBPs)are lipid carrier proteins involved in binding and transporting lipids.ACBPs play an important role in plant growth and development,as well as chilling,drought,oxidation and heavy metal stresses responses.However,the salt response mechanism is still unclear.Puccinellia tenuiflora,a monocotyledonous halophyte species belongs to Gramineae,which is an excellent model to study plant salt tolerance.In this study,PutACBPl was cloned,using molecular genetics technique in.tenuiflora.The open reading frame of PutACBPl is 981 bp,encoded 326 amino acids.PutACBPl belongs to ACBP Class II subfamily,including an acyl-CoA-binding ACB domain and an Ankyrin domain.RT-qPCR analysis showed that PutACBPl accumulated in all tissues.PutACBPl showed the highest expression levels in leaves,while the lowest levels in stems.The expression PutACBPl under treatments of stresses(200 mmol/L NaCl,100 mmol/L NaHCO3,75 mmol/L Na2CO3 stress,160 ?mol/L CdCl2,10 mmol/L H2O2 and,4?)were monitored using RT-qPCR analyses.The PutACBP1 was obviously induced in roots and leaves,which implied PutACBPl may play essential roles in plant response to saline-alkali,chilling,heavy metal and oxidative stresses.Arabidopsis seedlings and transient expression in tobacco leaves indicated that PutACBPl localized in cytoplasm.(His)6-PutACBPl recombinant proteins was expressed and purified in E.coli.Filter-binding assays indicated that(His)6-PutACBPl could bind phosphatidylcholine(PC)in vitro.In order to investigate the function of PutACBPl,we constructed PutACBP1-overexpression Arabidopsis under control of CaMV 35S promoter.Salt-tolerance analysis of PutACBP1 transgenic arabidopsis showed that the root length and fresh weight of overexpression PutACBP1 were better than wild type under the stress of 100 mmol/L and 125 mmol/L NaCl.Salt-tolerance analysis of Arabidopsis mutant At ACBP 2(Salk-023077)showed that root length and fresh weight of the mutant Arabidopsis were lower than wild type.Our results indicated that overexpression of PutACBP1 in wild-type Arabidopsis enhanced the root resistance to saline stresses.And PutACBPl could bind PC to regulate transportation from cytoplasm to membrane,and participate in plant growth and development.These results provide new evidences for understanding the molecular mechanisms of saline response in halophyte.