Functional Analysis Of Voltage-dependent Anion Channels From Vitis Vinifera Cv. ‘Thompson Seedless’

As the worldwide important economic fruit tree,grapes are widely used in table grape,juice and wine industries.Vitis vinifera cv.‘Thompson Seedless’ is the most dominant grape species used for raisin and table grapes,with a significant role in the industry.Voltage-dependent anion channels are the most abundant proteins in the mitochondrial outer membrane,playing an important role in regulating metabolite exchange across mitochondrial outer membrane,mitochondrial energy metabolism,mitochondria-mediated apoptosis,growth and reproductive development.However,the study about function of grapevine VDAC is rare.To further clarify the function of VvVDAC proteins in V.vinifera cv.‘Thompson Seedless’,the transcription level,three-dimension structures and subcellular localization properties of the VvVDAC gene family and the biological function of VvVDAC transient over-expression with salycilic acid(SA)treatment were studied.The main results are described as followings:1.The 3D structures of VvVDAC proteins were predicted that most of the family members consist of β-barrel and α-helix N-terminus except for VvVDAC6.1 and VvVDAC6.2 lacking the α-helix N-terminus.The amounts of β-sheets among VvVDAC proteins were between 9-19.VvVDAC3 and VvVDAC5 were predicted to contain the MPS motif.However,the subcellular localization of VvVDAC proteins after transient transformation in protoplasts showed that the VvVDAC3 and VvVDAC4 localized in mitochondria,whereas the VvVDAC1.1 and VvVDAC5 localized in mitochondria and other region in the cytoplasm,indicating that the presence of the MPS motif cannot fully decide the specific mitochondrial localization,which might be affected by other motifs.2.The plant binary-vector carrying VvVDAC were transiently transformed into Nicotiana benthamiana leaves followed by SA treatment.The results indicated that the VvVDAC1.1 was the major mitochondrial porin corresponding to H2O2 accumulation induced by SA treatment.However,the accumulation of H2O2 could not induce VDAC-mediated apoptosis.3.The total RNA from different ‘Thompson Seedless’ grape tissues,including roots,stems,leaves,tendrils and inflorescences were extracted to test the transcription level of 6 genes from VvVDAC gene family using RT-PCR.The results indicated that the VvVDAC1,which alternatively transcribed VvVDAC1.1 and VvVDAC1.2,was the most frequently transcribed gene.In the stem tissue,only the VvVDAC6.2 was transcribed,indicating that the transcription of VvVDAC6.2 was tissue-specific.