Molecular Characterization Of Phospholipase D Gene Family In Cotton And Functional Analysis Of GhPLD?1

Phospholipase D?PLD?is a major phospholipase family in plants that catalyzes the hydrolysis of phospholipids to produce phosphatidic acid and free polar head groups?e.g.,choline or ethanolamine?.Plant PLD not only regulates the metabolism of phospholipids in cell membranes,but also plays versatile roles in plant growth,development,and stress responses.However,until now,our knowledge concerning the PLD gene family members in cotton is limited,and their functions in cotton fiber development are also unrevealed.In this study,we performed the genome-wide analysis of PLD gene family in cotton.Also,the function of GhPLD?1 in secondary cell wall thickening was explored.Firstly,40,20 and 19 PLD genes were identified in the genomes of Gossypium hirsutum,Gossypium raimondii and Gossypium arboreum,respectively.All of these cotton PLDs were unevenly classified into six numbered subgroups:?,?/?,?,?,?and?.In the same subgroup,the PLDs shared the similar exon-intron structures,the same composition of functional domains and the subgroup-specific conserved amino acid residues.The expansion of cotton PLD gene families were mainly attributed to 16segmental duplication evnets and 3 tandem duplication events.Furthermore,purifying selection played a critical role in the evolution of duplicated PLD genes in cotton,increasing the functional redundancy of cotton PLD gene families.Then,the detailed expression patterns of GhPLD genes were revealed.These GhPLD genes were differently expressed in tissues.GhPLD?1A/D had the most abundant expression level in cotton fibers.The expression patterns of selected GhPLD genes in developing fibers suggested that GhPLD?1A/D might play an important role in the stage of secondary cell wall thickening.Finally,GhPLD?1 gene was successfully cloned in Gossypium hirsutum.GhPLD?1 was a typical plant PLD?gene.GhPLD?1 was distributed in both cell membranes and the cytoplasm.And it had the PIP₂-independent phospholipid hydrolysis activity.During cotton fiber development,the changing trend of PLD?activity in fibers was very similar to those of H₂O₂ level and cellulose content in fiber cells.The inhibition of PLD?activity obviously decreased the H₂O₂ and cellulose contents of in vitro-cultured cotton fibers.Furthermore,the overexpression of GhPLD?1 significantly increased the H₂O₂ content in Arabidopsis leaves under the induction of ABA.Based on these results,a possible model for the relationship of GhPLD?1 with secondary cell wall thickening in cotton fiber was proposed.At the onset of secondary cell wall thickening,GhPLD?1 was involved in regulating the continuous and high H₂O₂ accumulation.Then,the rosette stability of cellulose synthase complexes might be enhanced by the high H₂O₂ level,ultimately promoting cellulose accumulation.Our study made a contribution of revealing the function of PLD in cotton fiber development,and provided valuable information for understanding the secondary cell wall thickening.