Identification And Functional Analysis Of PutACBP Gene Family In Puccinellia Tenuiflora

Acyl-CoA binding proteins,containing acyl-CoA binding domain,are involved in lipid synthesis and decomposition,acyl-CoA homeostasis and other physiological processes.ACBPs play an important role in plant growth and development as well as biotic and abiotic stress response.Puccinellia tenuiflora is a monocotyledonous plant with strong salt and alkali resistance.It is an ideal material for investigating the mechanism of plant salt and alkali response.In this study,bioinformatics and molecular genetics techniques were used to clone five PutACBP genes from P.tenuiflora.PutACBP1-PutACBP5 encode 89-655 amino acids,belong to Class ? subfamily(PutACBP1,PutACBP2),Class ? subfamily(PutACBP3),Class ? subfamily(PutACBP4)and Class ? subfamily(PutACBP5)respectively.According to the analysis of conserved motifs,it was found that PutACBPs motifs contain about 6-41 amino acids,each protein contains 3-8 motifs,and all PutACBPs contain motif 1 and motif 2.There are cis-acting elements in the promoter region of PutACBPs in response to hormone and abiotic stress,indicating that PutACBPs may be involved in the regulation of hormone signaling pathway and abiotic stress response.PutACBPs were located on 3 chromosomes of P.tenuiflora(Chr01?Chr03?Chr04).RT-qPCR analysis showed that PutACBPs were expressed in roots,stems,leaves,sheaths,flowers,spikes and seeds of P.tenuiflora.The expression level of PutACBP1 was the highest in spikes,and the expression level was the lowest in leaves.The expression level of PutACBP2 and PutACBP3 were the highest in leaves,and the expression level were the lowest in stems.The expression level of PutACBP4 was the highest in flowers,and the expression level was the lowest in seeds.The expression level of PutACBP5 was the highest in stems,and the expression level was the lowest in leaves.Under abiotic stresses(200 mmol/LNaCl,75 mmol/L Na2CO3,100 mmol/L NaHCO3,4?,10 mmol/L H2O2,160 ?mol/L CdCl2),PutACBPs in roots and leaves showed up-regulated expression pattern.It is suggested that PutACBPs may be involved in the response of salt,alkali,low temperature,oxidation and heavy metal stress.The subcellular localization analysis showed that PutACBP1 and PutACBP2 are located in the plasma membrane,cytoplasm and nucleus,PutACBP3,PutACBP4,PutACBP5 are located in the plasma membrane and cytoplasm.The result of NBT staining showed that,compared with WT,the superoxide anion free radicals(O2·-)in Arabidopsis overexpressing PutACBP3 was less under salt stress,indicated Arabidopsis overexpressing PutACBP3 maintained the ROS homeostasis to cope with salt stress.Under 100 mmol/L NaCl stress,PLD? in Arabidopsis overexpressing PutACBP3 showed a higher expression level than WT,suggesting that salt stress tolerance of PutACBP3 may be related to the enhancement of PLD?expression.In summary,PutACBP gene family may be involved in regulating the response to stresses of P.tenuifora.These results provide a new evidence for the in-depth study of the molecular regulation mechanism of the stress response in P.tenuiflora.