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Synthesis and characterization of non-hydrolysable diphosphoinositol polyphosphate second messengers.

ABSTRACT: The diphosphoinositol polyphosphates (PP-IPs) are a central group of eukaryotic second messengers. They regulate numerous processes, including cellular energy homeostasis and adaptation to environmental stresses. To date, most of the molecular details in PP-IP signalling have remained elusive, due to a lack of appropriate methods and reagents. Here we describe the expedient synthesis of methylene-bisphosphonate PP-IP analogues. Their characterization revealed that the analogues exhibit significant stability and mimic their natural counterparts very well. This was further confirmed in two independent biochemical assays, in which our analogues potently inhibited phosphorylation of the protein kinase Akt and hydrolytic activity of the Ddp1 phosphohydrolase. The non-hydrolysable PP-IPs thus emerge as important tools and hold great promise for a variety of applications.

SUBMITTER: Wu M

PROVIDER: S-EPMC3558982 | biostudies-literature | 2013 Jan

REPOSITORIES: biostudies-literature

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Synthesis and characterization of non-hydrolysable diphosphoinositol polyphosphate second messengers.

Wu Mingxuan M Dul Barbara E BE Trevisan Alexandra J AJ Fiedler Dorothea D

Chemical science 20130101 1

The diphosphoinositol polyphosphates (PP-IPs) are a central group of eukaryotic second messengers. They regulate numerous processes, including cellular energy homeostasis and adaptation to environmental stresses. To date, most of the molecular details in PP-IP signalling have remained elusive, due to a lack of appropriate methods and reagents. Here we describe the expedient synthesis of methylene-bisphosphonate PP-IP analogues. Their characterization revealed that the analogues exhibit significa ...[more]

PMID: 23378892

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Project description:UNLABELLED:Biofilms are surface-attached multicellular communities. Using single-cell tracking microscopy, we showed that a pilY1 mutant of Pseudomonas aeruginosa is defective in early biofilm formation. We leveraged the observation that PilY1 protein levels increase on a surface to perform a genetic screen to identify mutants altered in surface-grown expression of this protein. Based on our genetic studies, we found that soon after initiating surface growth, cyclic AMP (cAMP) levels increase, dependent on PilJ, a chemoreceptor-like protein of the Pil-Chp complex, and the type IV pilus (TFP). cAMP and its receptor protein Vfr, together with the FimS-AlgR two-component system (TCS), upregulate the expression of PilY1 upon surface growth. FimS and PilJ interact, suggesting a mechanism by which Pil-Chp can regulate FimS function. The subsequent secretion of PilY1 is dependent on the TFP assembly system; thus, PilY1 is not deployed until the pilus is assembled, allowing an ordered signaling cascade. Cell surface-associated PilY1 in turn signals through the TFP alignment complex PilMNOP and the diguanylate cyclase SadC to activate downstream cyclic di-GMP (c-di-GMP) production, thereby repressing swarming motility. Overall, our data support a model whereby P. aeruginosa senses the surface through the Pil-Chp chemotaxis-like complex, TFP, and PilY1 to regulate cAMP and c-di-GMP production, thereby employing a hierarchical regulatory cascade of second messengers to coordinate its program of surface behaviors. IMPORTANCE:Biofilms are surface-attached multicellular communities. Here, we show that a stepwise regulatory circuit, involving ordered signaling via two different second messengers, is required for Pseudomonas aeruginosa to control early events in cell-surface interactions. We propose that our studies have uncovered a multilayered "surface-sensing" system that allows P. aeruginosa to effectively coordinate its surface-associated behaviors. Understanding how cells transition into the biofilm state on a surface may provide new approaches to prevent formation of these communities.

Dataset Information

Synthesis and characterization of non-hydrolysable diphosphoinositol polyphosphate second messengers.

Publications

Synthesis and characterization of non-hydrolysable diphosphoinositol polyphosphate second messengers.

Similar Datasets

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