Physiological Roles Of Phloridzin In Apple (Malus × Domestica)

In apple(Malus × domestica),phloridzin(phloretin 2’-O-glucoside)is the predominant dihydrochalcone(DHC)and comprises up to 90% of soluble phenolic compounds and 18% of the total dry weight in leaves.This makes apple unique in the plant kingdom because phloridzin does not accumulate in such huge amounts outside Malus.However,the physiological roles of phloridzin in planta remains largely elusive.Herein,we identified a key P2’GT(UDP-glucose: phloretin 2’-O-glucosyltransferase),MdUGT88F1,converting phloretin into phloridzin in apple based on an investigation of DHCs profiles in Malus germplasm.Subsequently,we analyzed the physiological roles of phloridzin in apple development and stress responses using MdUGT88F1 transgenic apple lines(including overexpressing and silencing lines)and Malus germplasm.The followings are the main results.1.A large-scale investigation of DHCs profiles in leaves from 411 Malus accessions was conducted.In Malus,phloridzin was the predominant DHC,and its accumulation followed a Gaussian distribution.Profile of DHC-glucosides(i.e.phloridzin,trilobatin and sieboldin)would be jointly determined by the activity of both P2’GT and P4’GT.Moreover,a key P2’GT gene,UGT88F1,was identified by performing genome-wide association analysis of the expression patterns of UGTs(UDP-dependent glycosyltransferases)and phloridzin accumulations.Also,characterizations of UGT88 F subfamily found that MdUGT88F4,a paralog of MdUGT88F1,was involved in the conversion of phloretin into phloridzin in apple.2.MdUGT88F1 and MdUGT88F4 were localized in both nucleus and cytoplasm via analysis of subcellular location.In the development of apple leaves,MdUGT88F1 and MdUGT88F4 could be transcriptionally regulated to maintain phloridzin biosynthesis via a feedback loop.Decreasing phloridzin biosynthesis in apple lines by RNA silencing of MdUGT88F1(and MdUGT88F4)led to trilobatin accumulation and a series of severe phenotypic changes(i.e.,severe stunting,reduced internode length,spindly leaf shape,more stems,and weak adventitious roots).Moreover,it was revealed that decreased phloridzin biosynthesis indirectly caused lignin reduction via modulating phenylpropanoid pathway flux,and gave rise to disorders of cell wall polysaccharides via indirectly changing myo-inositol metabolism.Ultimately,in MdUGT88F1-RNAi lines,the interfered cell wall deposition was characterized by a severe dwarf.3.MdUGT88F1-mediated phloridzin biosynthesis was negatively involved in Valsa canker resistance in apple.After infection with Valsa mali,a higher level of phloridzin compromised the Valsa canker resistance slightly by directly promoting growth and toxins production of the pathogen in MdUGT88F1-OE(over-expressing)apple lines.In contrast,in MdUGT88F1-RNAi lines,decreased phloridzin biosynthesis not only limited growth and infection of V.mali but also stimulated accumulation of SA and ROS via interference of cell wall deposition.Although modified cell wall deposition resulted in growth reduction,reinforces in tissue(e.g.,compact epidermis and thickened bark region)inhibited infection by V.mali along with increases in SA and ROS.4.MdUGT88F1-mediated phloridzin biosynthesis was closely associated with high light tolerance in apple.In MdUGT88F1-OE apple lines,increased phloridzin biosynthesis disturbed nitrogen metabolism and resulted in chlorophyll reductions,thereby compromising tolerance of high light(or UV)under natural conditions.In contrast,in the MdUGT88F1-RNAi lines,decreased phloridzin biosynthesis improved nitrogen assimilation and recycling.It was suggested that decreased phloridzin biosynthesis would improve nitrogen metabolism and enhance high-light tolerance in apple under high-light stress.