In Silico Analysis Of GDSL Genes In Arabidopsis And Brassica And The Investigation On Their Function In Seeds

Oilseed rape(Brassica napus L.) is the most important plant edible oil in China. For decades, rapeseed breeders have made great efforts to elevate the seed oil content (SOC) of rapeseed. To date, most of the efforts focused on the elevation of lipid biosynthesis efficiency, but little attention has been paid on the reducers of seed fatty acids (FAs). GDSL genes are an important family of hydrolyser, diminishing seed oil accumulation. However, so far there is a paucity of knowledge regarding to the number, distribution, expression pattern and function, in particular, the function in reducing seed FA of the GDSL genes in rapeseed. This study aims to know the feature of nucleotide sequence of the GDSL genes in the A genome of rapeseed as well as the genome of Arabidopsis, and the evolutionary relationship between the GDSL genes in rapeseed and Arabidopsis; to understand the expressional pattern and the effects of GDSL loss-of-function on SOC, and reveal the different expression profiles of GDSL genes between two Recombinant Inbred Lines (RILs), and two Near Isogenetic Lines (NILs), and the effect of temperature on GDSL expression and the SOC. The major results are as the following.(1) Using a variety of bioinformatic programs, we found141GDSL genes in the A genome of rapeseed and108GDSL genes in the genome of Arabidopsis, and analyzed the structures of the genes and constructed an evolutionary tree. Based on the analysis, we grouped the rapeseed GDSL genes into three clusters. The alignment of amino acid sequences between Arabidopsis and the rapeseed GDSL genes revealed a total of82orthologous pairs. We mapped all GDSL genes onto various chromosomes and linked the orthologs between rapeseed and Arabidopsis. We discover that GDSL genes distribute unevenly on different chromosomes and most of GDSL genes (116) can be identified with a role on a secretory pathway.(2) We selected a total of16GDSL gene from the five different chromosomes of Arabidopsis to investigate their expressional pattern and the effects of gene loss-of-function. Of these,9genes, namely AT1G23500, AT1G53900, AT1G54000, AT1G54790、AT1G75890, AT1G75900, AT2G03980, AT2G23540, AT3G0993, express in two or more tissues.3of them, namely AT2G19010, AT2G36325and AT5G42170, specifically express in only one kind of tissues. The expression of the rest4genes, namely AT1G54010, AT1G58020, AT2G19050and AT2G19060could not be detected in any of the tissues such as roots, leaves, buds, young and old siliques. The genes such as AT1G54000express highly in roots. The genes such as AT1G75000, AT2G03980, AT2G19010, AT2G36325and AT5G42170have more expression in buds. AT1G75900and AT2G23540have high expression in young siliques, whereas AT2G23540has high expression in old siliques.(3) Morphological observation did not show any obvious difference between the16gdsl mutants and wild-type (WT). However, there are significant differences in one-thousand-seed weight (TSW). Except for at2g19060, all other gdsl mutants had higher TSW than the WT (Col-0). The atlg53990even exceeded WT in TSW more than20%. Except for atlg53990and at5g42170, all the other14gdsl mutants had higher SOC than WT. The at2g19060exceeded WT in SOC more than30%. TEM observation reveals that higher SOC gdsl seeds features with a larger number of oil bodies and less proteins. In WT seed, the sum of linoleic and linolenic acids accounts for more than50%of total FAs, and dominates the FAs, whereas, the sum of linoleic and linolenic acids only accounts for less than50%of total seed FAs in most of the gdsl mutants.(4) There were significant differences for SOC between two recombinant inbred lines (namely, RIL-hoc or RIL-loc) at40or50days after pollination (DAP). At40DAP, a larger amount of proteins and less number of oil bodies in RIL-loc seeds than in RIL-hoc seeds were observed. RIL-hoc seeds had higher linoleic and linolenic acid content in RIL-loc. Most of the selected GDSL genes had a higher expressional level in RIL-hoc than in RIL-loc at30DAP, had a similar expressional level between RIL-hoc and RIL-loc at40DAP, but a lower expressional level in RIL-hoc than in RIL-loc at50DAP. On the other hand, most of GDSL genes (46out54) had a higher transcriptional level in low SOC near-isogenetic line (NIL-1) than in NIL-9. Only8GDSL genes had a higher transcriptional level in NIL-9than in NIL-1, suggesting the role of GDSL genes in decomposing seed FAs.(5) High temperature (T) induced the expression of numerous GDSL genes. More than80%of GDSL genes had higher expression in high T condition than in low T condition, indicating the role of temperature in inducing the expression of GDSL genes, and reducing seed FA accumulation. Despite of the temperature effect, we also noticed the significance of temperature and genotype interaction effect. Regardless of high, low or normal temperature conditions, there were always certain groups GDSL genes that expressed higher in NIL-1than in NIL-9.The above findings enrich our knowledge about the mechanism of seed FA accumulation in oil seed rape.