| Lipid metabolites are carriers of bioenergy storage such as triacylglycerol,which are the precursors of plant self-protective substance?wax?and signal molecules in stress reactions.They play a key role in growth,development and nutrient accumulation of plant.Therefore,it is crucial to dissect the regulation mechanisms of lipid synthesis,transformation and decomposition and to detect key lipid-regulated genes in abiotic stress for increasing corn yield,improving grain quality and enhancing resistance.Based our previous research,we have identified several candidate genes dynamically associated with expression pattern of 26oil-related genes by using the genome-wide dynamic liquid association?LA?model.Moreover,a key LA-scouting gene ZmNAC77 was screened that regulating multiple co-expression pattern gene pairs.We further performed candidate genes association analyses,expression analyses,transcriptional activation activity analyses,and transgenic validation of ZmNAC77,to preliminarily investigate the biological functions of this gene in lipid metabolism and abiotic stress.The main results were presented as follows:1.According to the previous study,the X=26 oil-related genes,Y=26 oil-related genes,and Z=whole genome genes were used in the genome-wide LA.A total of 58,318 pairs of significant genes dynamically associated with co-expression patterns with the genome-wide P<1.89×10-6.For the 190 important LA-scouting genes,we performed Gene Ontology?GO?enrichment analyses and discovered Z genes were mainly enriched in the metabolic process.Hence,Z gene with the largest absolute LA value that encodes the NAC77 protein was named ZmNAC77.This gene negatively regulated co-expression genes of LACS and DAGL(LA=-0.45,P=1.87×10-4).All SNPs in the region of 100 kb upstream and downstream of ZmNAC77 were extracted for linkage disequilibrium analyses,and 89 significant SNPs correlation with oil content were identified.It was preliminarily verified that this gene involved in the regulation of lipid metabolism pathway.2.The overexpression vector of ZmNAC77 gene was constructed and transferred into the Colombian Arabidopsis Col-0.The fatty acid contents of homozygous transgenic plants were determined by using GC-MS.The content of the fatty acid in transgenic positive lines with overexpressing ZmNAC77 was lower than that of the wild type,which proved that ZmNAC77is a negative regulator in the lipid metabolism pathway.The results of LA analysis by transgenic Arabidopsis presented that the gene had no significant correlation with the expression pattern of LACS and DAGL.The results showed that ZmNAC77 regulates the lipid metabolism pathway by negatively regulating the expression of oil-related genes.3.After detecting the gene structure of ZmNAC77,we found the open reading frame?ORF?of ZmNAC77 contained 1983 base pairs encoding 660 amino acids.Out of these amino acids,there were 23 ones can constitute a transmembrane domain.With the phylogenetic analysis,amino acid sequence analysis,and protein tertiary structure prediction analysis of ZmNAC77 gene,the results shown that ZmNAC77 has the structural features of the NAC transcription factor family which is most closely related to the ANAC072 in Arabidopsis.The expression pattern analysis showed that ZmNAC77 was highly expressed in the ear of maize on the flowering stage.Subcellular localization indicated that ZmNAC77 was localized in the cell membrane and nucleus without yeast autoactivation activity.Overexpression of ZmNAC77in Arabidopsis enhanced salt and drought tolerance compared with that of the wild type,which may suggest that ZmNAC77 play important roles in multiple stress resistance in plants. |
PDF Full Text Request