Role Of Rice Lysophosphatidic Acid Acyltransferase 2 In Response To Hyperosmotic Stress

Lysophosphatidic acid acyltransferase 2(LPAT2)acylates lysophosphatidic acid(LPA)at the sn-2 position to produce phosphatidic acid(PA),which is an important lipid intermediate involved in glycerolipid biosynthesis and acts as a signaling molecule in various biological processes.The role of LPAT2 in lipid metabolism,signaling and postembryo growth remains unknown due to the lethality of female gametophyte in knockout mutant of Arabidopsis LPAT2.Our preliminary study showed that LPAT2 is involved in osmotic stress response.This study focused on the functional characterization of rice LPAT2 through multiple approaches such as genetic complementation,biochemical assay,physiological test,cellular observation,molecular interaction,and RNA-sequencing.The major results are as following:1.Rice LPAT2 mRNA was detectable in all tissues tested with relatively higher abundance in leaves and roots,and was induced by salt,drought and ABA treatment,suggesting its possible roles in growth and osmotic stress response.Moreover,LPAT2 is localized the endoplasmic reticulum(ER)membrane,implicating its involvement in lipid synthesis in the eukaryotic pathway.2.A LPAT2 knockdown mutant,namely lpat2 with a substantially reduced transcript level of LPAT2,was isolated from rice,which provides a possibility for functional characterization of LPAT2 in post-embryo study.Moreover,the genetic LPAT2 complementation rice plants were generated to confirm its function.3.The lpat2 mutant was more sensitive to osmotic stress caused by salt and drought stress,while genetic complementation restored the lpat2 mutant phenotype to wild type plants.The results suggest that LPAT2 plays a positive role in osmotic tolerance in rice.4.LPAT2 is capable of catalyzing the PA formation using acyl-Co A and lysophosphatidic acid(LPA)as substrates.Lipid analysis showed that suppressed LPAT2 led to a substantial reduction of PA in plants under salt and drought stress.The PA application restored the phenotypes of the lpat2 mutant to wild type plants when they were subjected to ABA and salt stress treatments.The results suggest that LPAT2 and its PA are important for plant tolerance to osmotic stress.Moreover,lipid-protein interaction showed that PA binds to salt overly sensitive 3(SOS3),suggesting that PA may be also involved in SOS pathway to regulate ion homeostasis under osmotic stress.5.Lipid profiling showed that suppressed LPAT2 also led to reduced phosphatidylcholine,(PC),phosphatidylethanolamine(PE)and lysophosphatidylcholine(lyso PC)in some extent under salt stress,suggesting that LPAT2 has contribution to phospholipid metabolism under osmotic stress.6.The transcriptome results revealed that suppressed LPAT2 led to substantial alteration of differentially expressed genes(DEGs)involved in diverse pathways,implicating its biological significance.In particular,DEGs were enriched the pathways involved in ion transport and hormone signaling in response to salt stress.The results suggest that LPAT2 and its PA are important for hormone response and ion homeostasis in response to salt stress.In summary,rice LPAT2 was localized to the ER to catalyze the PA synthesis.The suppressed LPAT2 resulted in substantial reduced PA in plants,which conferred plants less sensitive to ABA and more sensitive to osmotic stress.The transcriptome results revealed that suppressed LPAT2 led to a large of DEGs involved in diverse pathways,particularly,in hormone signaling and ion homeostasis in response to salt stress.Taken together,the LPAT2 and its PA play positive roles in osmotic stress tolerance via hormone response.