Regulation of nitrogen starvation responses by the alarmone (p)ppGpp in rice

Nitrogen is an essential macronutrient for all living organisms and is critical for crop productivity and quality.In higher plants, inorganic nitrogen is absorbed through roots and then assimilated into amino acids by the highly conserved glutamine synthetase/glutamine:2-oxoglutarate aminotransferase(GS/GOGAT) cycle.How nitrogen metabolism and nitrogen starvation responses of plants are regulated remains largely unknown. Previous studies revealed that mutations in the rice ABNORMAL CYTOKININ RES...     

A multi-repeat adhesin of the phytopathogen, Pectobacterium atrosepticum, is secreted by a Type I pathway and is subject to complex regulation involving a non-canonical diguanylate cyclase

Cyclic diguanylate (c-di-GMP) is a second messenger controlling many important bacterial processes. The phytopathogen Pectobacterium atrosepticum SCRI1043 (Pba1043) possesses a Type I secretion system (T1SS) essential for the secretion of a proteinaceous multi-repeat adhesin (MRP) required for binding to the host plant. The genes encoding the MRP and the T1SS are tightly linked to genes encoding several putative c-di-GMP regulatory components. We show that c-di-GMP regulates secreted MRP levels in Pba1043 through the action of two genes encoding predicted diguanylate cyclase (DGC) and phosphodiesterase proteins (ECA3270 and ECA3271). Phenotypic analyses and quantification of c-di-GMP levels demonstrated that ECA3270 and ECA3271 regulate secreted MRP levels by increasing and decreasing, respectively, the intracellular levels of c-di-GMP. Moreover, ECA3270 represents the first active DGC reported to have an alternative active-site motif from the 'canonical' GG[D/E]EF. ECA3270 has an A-site motif of SGDEF and analysis of single amino acid replacements demonstrated that the first position of this motif can tolerate functional substitution. Serine in position one of the A-site is also observed in many other DGCs. Finally, another T1SS-linked regulator (ECA3265) also plays an important role in regulating secreted MRP, with an altered localization of MRP observed in an ECA3265 mutant background. Mutants defective in these three T1SS-linked regulators exhibit a reduction in root binding and virulence, confirming that this complex, finely tuned regulation system is crucial in the interaction with host plants.     

The Cek1 MAPK is a short-lived protein regulated by quorum sensing in the fungal pathogen Candida albicans

Mitogen activated protein kinase (MAPK) cascades are signal transduction mechanisms present in eukaryotic cells that allow adaptation to environmental changes. MAPK activity is mainly regulated by dual phosphorylation in a TXY motif present in the kinase subdomain VIII as well as dephosphorylation by specific phosphatases. The Cek1 MAPK is involved in filamentous growth in Candida albicans and is an important determinant of virulence in this microorganism; its activation is controlled by the Sho1 adaptor protein. Here we show that Cek1 phosphorylation is regulated by quorum sensing (QS). Cek1 phosphorylation is prevented by farnesol, a compound that also regulates the dimorphic transition in this fungus. Farnesol also induced the activation of Mkc1, the MAPK of the cell integrity pathway. The role of farnesol in Cek1 phosphorylation is independent of the Chk1 histidine kinase, a putative QS sensor, as revealed by genetic analysis. In addition, Cek1, not Hog1, is degraded by proteasome, as revealed by the use of a conditional lethal protein degradation mutant. Our data therefore describe two different mechanisms (QS and protein degradation) that control a MAPK pathway that regulates virulence in a fungal pathogen.     

Significance of accumulation of the alarmone (p)ppGpp in chloroplasts for controlling photosynthesis and metabolite balance during nitrogen starvation in Arabidopsis

Photosynthesis Research - The regulatory nucleotides, guanosine 5′-triphosphate 3′-diphosphate (pppGpp) and guanosine 5′-diphosphate 3′-diphosphate (ppGpp), were originally...     

Translational accuracy enhanced in vitro by (p)ppGpp

Summary The effect of (p)ppGpp on the accuracy of translation in vitro was studied with a system that has a missense error frequency similar to that of living bacteria. When poly (U)¹ is translated, limiation of the system in Phe increases the Leu missense error frequency. The introduction of (p)ppGpp to the Phelimited mixtures reduces significantly the missense errors as well as reduces the rate of translation. The introduction of (p)ppGpp to a full system has no effect on the accuracy of translation but does reduce its rate. The effects of (p)ppGpp on rate and accuracy of translation can be simulated in part by other inhibitors of translation such as GDPCP, fusidic acid and tetracycline. Furthermore, the presence of ppGpp or GDPCP in a Phe-limited system leads to an accumulation of Phe-tRNA, while a Phe-limited system that contains only GTP has negligibly small concentrations of Phe-tRNA. We conclude that one way in which (p)ppGpp improves the accuracy of translation is by permitting the system to maintain a favorable Phe-tRNA/Leu-tRNA ratio.     

Non-CDK-bound p27 (p27) is a marker for cell stress and is regulated through the Akt/PKB and AMPK-kinase pathways

p27Kip1 (p27) tumour suppressor protein is regulated by multiple mechanisms including its turnover, localization and complex formation with its key targets, cyclin-dependent kinases (CDK) and cyclins. We have earlier shown that p27 exists in cells in a form that lacks cyclin/CDK interactions (hence non-CDK, p27^NCDK) but the nature of p27^NCDK has remained unresolved. Here we demonstrate that the epitope recognized by the p27^NCDK-specific antibody resides in the p27 CDK-interaction domain and that p27^NCDK is regulated by the balance of CDK inhibitors and cyclin-CDK complexes. We find that signalling by cellular growth promoting pathways, like phosphoinositol 3-kinase (PI3K) and specifically Akt/PKB kinase, inversely correlates with p27^NCDK levels whereas total p27 levels are unaffected. p27^NCDK, but not total p27, is increased by cellular perturbations such as hyperosmotic and metabolic stress and activation of AMP-activated protein kinase (AMPK). By using AMPK catalytic subunit proficient and deficient cells we further demonstrate that the AMPK pathway governs p27^NCDK responses to metabolic stress and PI3K inhibition. These results indicate that p27^NCDK is a sensitive marker for both cell stress and proliferation over and above p27 and is regulated by Akt/PKB and AMPK pathways.