Project description:Hypoxia exerts major effects on gene expression, which is reconfigured to meet the needs of the stressed plant. In the recent years the mechanisms of oxygen sensing and transcriptional regulation of a cluster of anaerobic genes was described. We utilized a hypomorphic, post-transcriptional gene silencing defective ARGONAUTE1 (AGO1) mutant, ago1-27, to evaluate the contribution of AGO1 to plant tolerance to submergence and its effects on gene expression.
Project description:Microarray analysis of wild type plants and plants with reduced (ago1-27 and se-1) or increased miR156 levels (se-1 p35S:MIR156). Shoot apices were dissected from 20-day-old, short-day grown plants.
Project description:SNF1 RELATED PROTEIN KINASE 1 (SnRK1) is proposed as a central integrator of regulatory pathways in plant stress and energy starvation signaling. We observed in this study that the Arabidopsis SnRK1.1 dominant negative mutant (SnRK1.1K48M) had lower tolerance to submergence than the wild-type, suggesting that SnRK1.1-dependent phosphorylation of target proteins is important in energy starvation signaling triggered by submergence. To gain further insight into submergence signaling mechanisms, we determined the temporal response to energy starvation through AMP/ATP quantification and used quantitative phosphoproteomics to compare the global changes in phosphopeptides in Col-0 and SnRK1.1K48M. We found that the phosphorylation levels of 59 peptides increased and the levels of 96 peptides decreased in Col-0 within 0.5–3 h of submergence. Among the 59 peptides with increased phosphorylation in Col-0, 49 did not show increased phosphorylation levels in SnRK1.1K48M under submergence. These proteins are involved in sugar synthesis, glycolysis, osmotic regulation, ABA signaling, protein synthesis and ROS signaling. In particular, the phosphorylation of MAPK6, which is involved in regulating ROS responses under different abiotic stresses, was disrupted in the SnRK1.1K48M mutant. In addition, PTP1, a negative regulator of MAPK6 activity that directly dephosphorylates MAPK6, was also regulated by SnRK1.1. These results reveal insights into the function of SnRK1 and the downstream signaling factors of submergence.
Project description:SNF1 RELATED PROTEIN KINASE 1 (SnRK1) is proposed as a central integrator of regulatory pathways in plant stress and energy starvation signaling. We observed in this study that the Arabidopsis SnRK1.1 dominant negative mutant (SnRK1.1K48M) had lower tolerance to submergence than the wild-type, suggesting that SnRK1.1-dependent phosphorylation of target proteins is important in energy starvation signaling triggered by submergence. To gain further insight into submergence signaling mechanisms, we determined the temporal response to energy starvation through AMP/ATP quantification and used quantitative phosphoproteomics to compare the global changes in phosphopeptides in Col-0 and SnRK1.1K48M. We found that the phosphorylation levels of 59 peptides increased and the levels of 96 peptides decreased in Col-0 within 0.5�� h of submergence. Among the 59 peptides with increased phosphorylation in Col-0, 49 did not show increased phosphorylation levels in SnRK1.1K48M under submergence. These proteins are involved in sugar synthesis, glycolysis, osmotic regulation, ABA signaling, protein synthesis and ROS signaling. In particular, the phosphorylation of MAPK6, which is involved in regulating ROS responses under different abiotic stresses, was disrupted in the SnRK1.1K48M mutant. In addition, PTP1, a negative regulator of MAPK6 activity that directly dephosphorylates MAPK6, was also regulated by SnRK1.1. These results reveal insights into the function of SnRK1 and the downstream signaling factors of submergence.
Project description:Sequencing of small RNA libraries prepared from total RNA or RNA extracted from immuno affinity purified AGO1 from Col-0, rns2-2 and rns2-3 flowers. The aim of this experiment was to compare profiles of small RNA expression between Col-0 (wild type) and mutants of the ribonuclease T2 rns2-2 rns2-3 genotypes and characterize AGO1-bound small RNAs in the same genotypes. Two biological replicates of each genotype were prepared and analyzed.
Project description:Col-0 floral stem was grafted on the msh1 mutant (Col-0/msh1); on the dcl2,3,4,msh1 quadruple mutant (Col-0/dcl2,3,4,msh1); on Col-0 (Col-0/Col-0). Seeds were collected from the grafted Col-0 scion after grafts were established. Seed coming from the graft then were grown on the peat mix, leaf tissue was collected at the bolting and used for the total RNA sequencing.