Functional Analysis Of SsNRT1.7 And SsNRT2.5 In Suaeda Salsa During Nitrogen Starvation

Suaeda salsa L.is an euhalophyte with high salt tolerance which widely grows on coastal areas and inland saline soils of China,and it can be used for the remediation of saline soils.The content of NO₃^-in saline soils is low in field,while S.salsa grows well in high salinity and low nitrogen environments.Up to now,it is not clear how S.salsa adapts to high salinity and low nitrogen habitats.Therefore,in order to understand the adaptive mechanism of S.salsa to low nitrogen environment,the function of nitrate transporter(NRT)genes SsNRT1.7 and SsNRT2.5 in S.salsa and transgenic Arabidopsis plants during nitrogen starvation were analyzed.The results are as follows:1.Physiological and molecular mechanisms of nitrate nitrogen remobilization in S.salsaSeedlings of S.salsa were pre-cultured with 1 m M NO₃^--N nutrient solution combined with 200 m M NaCl for 30 days,then seedlings were watered with 1 m M NO₃^--N(N normal treatment)and N-free nutrient solution(N starvation treatment)for additional 10 days.The biomass of the shoots and roots,NO₃^-and chlorophyll content and net photosynthetic rate of leaves in different leaf positions(old,mature and young leaves),and the expression of SsNRT1.7,SsNRT2.5 in the leaves and SsNRT2.1 and SsNRT2.5 in the roots of S.salsa were determined.After N starvation for 10 days,compared with the control(N normal treatment),the shoot biomass of S.salsa significantly decreased,but the root biomass increased.N-starvation treatment may increase the nitrogen absorption by inducing the root growth of S.salsa,which may be an adaptive mechanism of S.salsa to low-nitrogen environment.The contents of NO₃^-and chlorophyll in the old,mature and young leaves of S.salsa significantly decreased after N starvation for 3 and 10 days,as well as the net photosynthetic rate after N starvation for 10 days.The expression of SsNRT1.7 and SsNRT2.5 in old leaves was significantly up-regulated after 3 and 10 days during N starvation,especially on the 10th day,suggesting that SsNRT1.7 and SsNRT2.5 may play an important role in the remobilization of NO₃^-in the old leaves of S.salsa.The expression of SsNRT2.1 in the root was up-regulated after N starvation for 3 days and down-regulated on the 10th day;the expression of SsNRT2.5 in the root was not significantly up-regulated after N starvation for 3 days,while down-regulated on the 10th day.It indicates that SsNRT2.1may play a role in absorbing NO₃^-by the roots in the initial stage of N starvation.However,due to the severe lack of nitrate nitrogen in the rhizosphere during long-term N starvation,both SsNRT2.1 and SsNRT2.5 in the roots were significantly down-regulated in S.salsa.2.Functional verification of SsNRT1.7 and SsNRT2.5 in ArabidopsisThe selected seeds of overexpression homozygous plants SsNRT1.7-OE12,OE14,SsNRT2.5-OE15,OE35 and wild-type Arabidopsis(WT)plants were sown on 1/2 MS medium for 3 weeks,then seedlings were transferred to a new 1/2 MS medium(N normal treatment)and N-free medium(N starvation treatment)for additional 3 days,and the physiological indexes were measured.Both under N normal and N starvation treatments,the biomass of SsNRT1.7-OE12,14 and SsNRT2.5-OE15,OE35 were higher than WT,indicating that SsNRT1.7 and SsNRT2.5 may promote the growth of Arabidopsis through promoting N absorption and/or remobilization in Arabidopsis.Under N normal treatment,the NO₃^-content in old and young leaves of SsNRT1.7-OE12,14 and SsNRT2.5-OE15,35were higher than that of WT.Under N starvation treatment,the NO₃^-content in old leaves of SsNRT1.7-OE12,14 and SsNRT2.5-OE15,35 was lower than WT,and the NO₃^-content in young leaves of SsNRT1.7-OE14 increased compared to WT,but the NO₃^-content in young leaves of other lines was not significantly different from WT.This indicates that SsNRT1.7 may be involved in transferring NO₃^-from old leaves to young leaves,thereby increasing the NO₃^-content in young leaves.The above results need to be further verified.