Response Of Banana Seedling Roots To Potassium Deficiency Stress And Its Molecular Mechanism

Banana is a typical "potassium-loving" crop,which needs a lot of potash fertilizer during its growth period.The content of available potassium is low in some banana plantation soils in southern China,putting the banana plant under potassium deficiency stress,which affected banana production.Meanwhile,the management status of unreasonable topdressing potassium fertilizer aggravated this situation.In order to elucidate the physiological and molecular mechanism of banana in response to potassium-deficiency stress,in this study,pot experiments were conducted with banana seedlings(Musa acuminate L.AAA ground,cv.Brazilian)as experimental materials,which were treated with three concentrations of potassium,which are K0:0.00 mmol/L(low postassium),K1:0.03 mmol/L(low potassium),K2: 3.00 mmol/L(appropriate potassium)respectively.During the growth of the banana seedlings from 0 day to 35 days,the responses of banana seedling morphological,physiological and nutrient changes to potassium deficiency stress were investigated.Based on this,the combination of transcriptomics,proteomics and phosphorylated proteomics was employed to detect the key genes responding to the potassium-deficiency stress in banana seedling roots,and the function of the identified key genes were verified by K+ deficient yeast complementary experiments.The main results are as follows:1.With the growth of banana seedlings,the K0,K1 treatment of potassium deficiency inhibited the growth,development and activity of banana seedling roots.When the seedlings are 35 days old,their above ground biomass decreased 2.9%-3.2%,compared to the 30-day old banana seedlings,their root activity under K0,K1 treatment decreased 51.6%,33.9% respectively,compared to the K2 treatment.Meanwhile,K0 treatment obviously caused yellowing and withering of leaves.It was further observed from root tissue structure that the root apical cell membrane outline under K0 treatment was not clear,and there were gaps between cell walls,forming large intercellular spaces,which was not good for the transfer of the nutrient ion.Thus,it can be seen that the treatment of potassium deficiency affected the morphology of banana seedlings and the growth of banana seedling roots.The lower of K level,the more obvious symptom caused by the potassium deficiency.2.The conductivity of leaves and roots and MDA content in these two organs increased under potassium-deficiency treatments of K0 and K1.When the banana seedlings are 35 days old,the leaf and root conductivity under K0 treatment is 0.7%,3.5% respectively higher than that of K2 treatment,MDA content in leaf and root under K0 treatment is 6.5%,18.0% higher than that of K2 treatment,showing the potassium-deficiency stress led to cell membrane lipid peroxidation and resulted in cell membrane damage to a certain extent,and the damage to roots was more rapid and serious than to leaves.The activity of antioxidant enzymes(SOD,POD and CAT)in leaves and roots exhibited the same changing trend after the potassium-deficiency treatment.The antioxidant enzyme activity(SOD,POD and CAT)in banana seedlings changed little in the early stage,but the activity of POD,CAT significantly decreased in the later stage,indicating that the adaptive mechanism of banana seedlings to stress was initiated to eliminate ROS when they suffered from potassium-deficiency stress at the beginning.While with the exacerbating of potassium deficiency and the weakening of plant stress resistance,in the later stage,the physiological function in banana seedlings tended to be disordered.Besides,potassium-deficiency stress promoted the accumulation of NK,but inhibited the accumulation of P.The ratio of N:P:K in roots,shoots and the whole plants under K2 treatment tended to be reasonable,but potassium-deficiency stress(K0 and K1 treatment)caused the unbalance of N,P and K in banana seedlings.3.Transcriptome analysis showed there were 3350 DEGs(Differentially expressed genes)in banana seedling roots after 30 days of treatment,which were mainly enriched in 14 pathways involved in linolenic acid metabolism,unsaturated fatty acid biosynthesis,galactose metabolism,glycerol ester metabolism,and sphingolipid metabolism,etc.Genes such as NRT1.1,HKT2,IAA16,A-2b,78A4,pectinesterase2,playing key roles in responding to potassium deficiency stress were identified.Among them,the expression of NRT1.1(NO3-transporter gene)was significantly down-regulated.It can be inferred that the uptake and transport of NO3-could regulate the NPK balance in the cells of banana seedlings roots under potassium-deficiency stress.As the DEGs related to ion transports,transcription factors and cell wall showed the differential expression of genes responded to K0 and K1 treatments was not significant,only K0 treatment was selected as potassium-deficiency treatment in the following experiments.4.The proteomics approach identified 457 differentially expressed proteins in banana seedlings roots under K0 treatment,including 240 up-regulated and 217 down-regulated.Among them,the proteins related to glutathione metabolism were significantly down-regulated,which suggested that potassium-deficiency stimulated banana seedlings roots to produce ROS signals and activated the antioxidant defense systems to clear the excessive ROS,which temporarily mitigated the damage to banana seedling roots caused by potassium-deficiency stress,also inhibited the expression of the cell growth-related proteins.5.By using the phosphorylated proteomics,263 differential phosphorylation sites on the proteins in banana seedling roots under K0 treatment were revealed,which were located on the 215 stress resistant-proteins including PP2 C,glutathione transferase,pyruvate dehydrogenase,pyruvate kinase and acetyl coenzyme A synthase,etc,which were differentially phosphorylated proteins.Four proteins containing glutathione transferase conservative domain were phosphorylated on serine,and the phosphorylation levels were significantly up-regulated,demonstrating the phosphorylation modification of the glutathione transferase played an important part in the defense of banana seedling roots against potassium-deficiency stress.6.By utilizing the multi-omics analysis,two key genes(Ma QORH,Maznt A)in response to potassium-deficiency stress were obtained and their full-length c DNAs were cloned.Sequencing analysis showed the size of the Ma QORH,Maznt A were 1002 bp,2295 bp respectively,encoding333 and 764 amino acid respectively.The K+ deficient yeast complementary experiment was employed to verify the function of potassium-transporting of Ma QORH and Maznt A.Results showed that both Ma QORH and Maznt A recovered the growth of K+ deficient yeast on low potassium culture medium.In summary,potassium-deficiency stress not only affected the morphology of the above ground of banana seedlings and the growth of their roots,but also caused the unbalance of N,P,K.Simultaneously,multi-omics analysis found that the differential expression of numerous genes and proteins as well as proteins phosphorylation were involved in response of banana seedling roots to potassium-deficiency stress.The function of potassium-transporting of two identified key genes responding to potassium-deficiency stress was confirmed,which provided a source of genes and scientific evidences for cultivation of new low potassium tolerant banana variety.