Mechanism Of Sucrose Non-fermentation Related Protein Kinase 1 Family(SnRK1s)Controlling Low Potassium Tolerance In Arabidopsis

Plant growth and development require adequate mineral nutrition,which needed for plant mainly come from the soil.Studies have shown that carbon uptake by plant photosynthesis determines the uptake of mineral nutrients by roots.When confronted with adverse conditions such as lack of light,drought,waterlogging,extreme temperatures,etc.,plants can maintain and restore the energy balance through a series of carbohydrate metabolism regulating mechanisms.However,the mechanism of changes in carbohydrate metabolism affecting the absorption of mineral nutrients remains to be further studied.In plant,SnRK1(sucrose non-fermenting-1-related protein kinases)is a central regulator of metabolic balance in plants,which plays a vital role in plant growth and response to different stresses.SnRK are widely present in plants and have three subfamilies:SnRK1,SnRK2 and SnRK3.SnRK1 have higher sequence similarity with yeast SNF1 and mammalian AMPK,which plays an important role in energy metabolism balance,while SnRK2 and SnRK3 are plant-specific protein kinases.SnRK2 and SnRK3 are mainly involved in ABA signaling pathways and mineral nutrient absorption pathways in plants.Therefore,in this paper,we want to explore the uptake of mineral nutrients by plants in the growth environment of barren nutrition through the study of SnRK1.This paper used model plant Arabidopsis as research materia1.We first identified two T-DNA mutants:snrk1.1 and snrk1.2 of the SnRK1 family of Arabidopsis thaliana.By comparing the phenotypic changes of the mutants in the absence of nitrogen,phosphorus and potassium-the three major nutrients necessary for plants-respectively,it was found that the mutants snrk1.1 and snrk1.2 were more difficult to adapt to the low potassium environment than the wild type.Next,the mechanism of low-potassium tolerance in Arabidopsis affected by the sucrose non-fermentation-associated protein kinase 1 family(SnRK1s)was analyzed using techniques such as plant molecular genetics,yeast two-hybrid,and bimolecular fluorescence complementarity(BIFC).The main results are as follows:1.SnRK1s affects the growth and development of Arabidopsis in low potassium environment.To explore the dependence of SnRK1 on K+,we designed a series of K+concentration gradient experiments.The snrk1.1 and snrk1.2 mutant exhibited a more sensitive phenotype than the wild-type at K+concentrations of 0.01 mM to 1 mM.In order to further confirm that the low potassium sensitive phenotype is caused by mutations of the SnRK1 gene,two SnRK1.1 overexpression lines were constructed.It was found that SnRK1.1 overexpressing plants had a low potassium tolerance phenotype under low potassium conditions.Through K+concentration gradient experiments,it was found that the phenotypic difference between SnRK1.1 overexpression and wild type gradually disappeared as the concentration of K+in the medium increased.2.The regulation of the low potassium tolerance of Arabidopsis by SnRK1s is affected by sucrose.First,we replaced 1%sucrose(w/v)with 1%glucose(w/v),1%fructose(w/v),and 1%mannitol(w/v)in the medium,and observed whether the snk1.1 and snrk1.2 mutant has a low potassium-sensitive phenotype.It was found that the low potassium phenotype of the snrk1.1 and snrk1.2 mutants had sucrose dependence.Next,by adding different concentrations of sucrose to the culture medium,we found that the low potassium-sensitive phenotypes of the snrk1,1 and snrk1,2 mutants were sucrose-concentration-dependent and SnRK1.1,SnRK1.2 mainly played a role at low sucrose concentrations.The tolerance to low potassium of SnRK1.1 overexpression line was inhibited by high sucrose.Therefore,it was confirmed that the regulation of SnRK1s on low potassium tolerance was affected by sucrose.3.SnRK1.1 affects Arabidopsis potassium uptake through potassium channel AKT1.In order to verify that the difference in low potassium adaptation is indeed caused by the changed potassium level in mutant plants,we use inductively coupled plasma mass spectrometry(ICP-MS)to determine the potassium content of wild type,snrk1.1 and SnRK1.1 overexpressed line in different potassium concentration.It was found that the potassium ion content of the snk1.1 was lower than that of the wild type under the condition of potassium deficiency,and the overexpression of SnRK1.1 helps plant to absorb more potassium in the low potassium environment and alleviate the symptoms of potassium deficiency.Therefore,SnRK1.1 does affect the potassium ion content in plants.Further effort was done to explore which pathway does SnRK1.1 regulate Arabidopsis low potassium tolerance.First,the physical interaction of SnRK1.1 and AKT1 was demonstrated by yeast two-hybrid experiments,and then the interaction between SnRK1.1 and AKT1 in vivo was verified by two-molecule fluorescence complementary(BIFC)experiments.And SnRK1.1 overexpression line based on the aktl mutant background was constructed.Through genetic experiments,it was proved that SnRK1.1 affects plants potassium uptake through potassium channel AKT1 at low potassium condition.In summary,in addition to plays an important role in energy metabolism balance,SnRK1 also in response to low potassium condition,and affects plant potassium absorption by interacting with the potassium channel AKT1.This finding establishes a link between carbohydrate metabolism and K+nutrient uptake through SnRK1,which is expected to provide theoretical guidance and experimental basis for molecular breeding of highly resistant crops.