Study On The Synergistic Effect Of GOGAT And GS During Ammonia Assimilation Of Sugar Beet(beta Vulgaris L.)

The diploid beet KWS0143 was selected as the test materials, to study the GS and GOGAT enzyme activity, gene expression and protein content in beet leaves and roots treated with different NO3- / NH4 + ratios of 4: 0,2: 1,1: 2,3: 1,1: 3,1: 1,0: 4 for 24 h. Screening the best proportion of nitrogen, at this scale, sugar beet seedlings were treated with Azaserine(GOGAT inhibitors) and PPT( GS inhibitor) at different concentrations and times. Respectively, the leaves and roots samples were taken after processing for 6h, 9h, 12 h and 24 h. Then by q RT-PCR method to detect the mRNA expression of GS1, GS2 and GOGAT of the seedlings. Simultaneously detecting the enzyme activity and protein content of GS and GOGAT. Further to elucidate the synergy mechanism of GS and GOGAT from the perspective of the enzyme activity, gene expression and protein levels. The results are as follows:(1)The GS activity in beet leaves effected by NO3- / NH4 + proportion is 2: 1> 1: 1> 3: 1> 4: 0> 1: 2: 1: 3, and effected by beet roots is 2: 1> 3: 1> 100: 0> 1: 1> 1: 2> 1: 3> 0: 4. The GOGAT activity in beet leaves effected by NO3- / NH4 + proportion is 2: 1> 4: 0> 1: 1> 3: 1> 1: 2> 1: 3> 0: 4. The GS activity in beet roots effected by NO3- / NH4 + + proportion is 2: 1> 4: 0> 1: 1>: 3: 1> 1: 2> 1: 3> 0: 4. The optimum nitrogen ratio of NO3-: NH4 + is 2: 1.(2)When azaserine concentration of 0.5μmol · L-1 was applied, GOGAT activity in beet leaves treated for 6 hours was inhibited, GS activity was inhibited when treated with azaserine for 9 hours. In Beetroot, after treatment with azaserine for 2 hours, the GOGAT activity was inhibited. Also, the decreased GS activity was seen at 9h after treatment. The inhibition of GOGAT activity occurred before GS activity, indicating that the activity of GS and GOGAT showed synergistic trends.(3)Under the treatment of azaserine in beet leaves, GOGAT gene expressions was up-regulated at 2-6 h, and the expression from 6h to 24 h showed a descending trend overall. The GS2 gene expression showed the low- high- low trend from the time of 2h to 24 h. Whereas the GS1 gene expression began to decrease at 2h, and reached the minimum at 24 h. The enzymes synergy in Beet leaves occurs in GS2 and GOGAT. In beetroot, the gene expression of GS1 and GOGAT gene showed the same trend, indicating that the synergy in beetroot GS and GOGAT occurs between GS1 and GOGAT. Besides, the symergy was seen in protein content when treating beet leaves and roots with azaserine.(4)Treating beet leaves with 0.5μmol · L-1 PPT, the GS and GOGAT activity were both inhibited at 9h. In the beet root, the GS activity was inhibited at 6h under the same PPT concentration. For GOGAT activity, the inhibited time was 6 hours. The inhibition of GS activity occurred before GOGAT, indicating that GOGAT and GS showed synergistic trends in activity.(5)From the view point of gene expression, when processing beet with PPT at different times, GOGAT and GS2 gene expression at 2-6 h were on the rise, and that of 6-24 h showed a downward trend.The GOGAT and GS2 gene expression experienced synergistic changes during the processing time. Those results indicated that the GS and GOGAT synergies in leaf beet under PPT treatment was dorminated by mainly GS2 / GOGAT. The assimilation of NH4 + in leaves was conducted through GS2 and GOGAT synergy.(6)In beetroot treated with PPT, the gene expression of GOGAT and GS1 both showed a trend of high to low, indicating the synergies was mainly GS1 / GOGAT based, the assimilation of NH4 +was conducted by GOGAT and GS1. Besides, the symergy was seen in protein content when treating beet leaves and roots with azaserine.(7)We can conclude that the inhibition of azaserine and PPT on GOGAT and GS in beet leaves and roots occurred at the enzyme activity, protein content and transcriptional levels.