IP7-SPX Domain Interaction Controls Fungal Virulence by Stabilizing Phosphate Signaling Machinery

In the human-pathogenic fungus Cryptococcus neoformans, the inositol polyphosphate signaling pathway is critical for virulence. We recently demonstrated the key role of the inositol pyrophosphate IP7 (isomer 5-PP-IP5) in driving fungal virulence; however, the mechanism of action remains elusive. Using genetic and biochemical approaches, and mouse infection models, we show that IP7 synthesized by Kcs1 regulates fungal virulence by binding to a conserved lysine surface cluster in the SPX domain of Pho81. Pho81 is the cyclin-dependent kinase (CDK) inhibitor of the phosphate signaling (PHO) pathway. We also provide novel mechanistic insight into the role of IP7 in PHO pathway regulation by demonstrating that IP7 functions as an intermolecular "glue" to stabilize Pho81 association with Pho85/Pho80 and, hence, promote PHO pathway activation and phosphate acquisition. Blocking IP7-Pho81 interaction using site-directed mutagenesis led to a dramatic loss of fungal virulence in a mouse infection model, and the effect was similar to that observed following PHO81 gene deletion, highlighting the key importance of Pho81 in fungal virulence. Furthermore, our findings provide additional evidence of evolutionary divergence in PHO pathway regulation in fungi by demonstrating that IP7 isomers have evolved different roles in PHO pathway control in C. neoformans and nonpathogenic yeast.IMPORTANCE Invasive fungal diseases pose a serious threat to human health globally with >1.5 million deaths occurring annually, 180,000 of which are attributable to the AIDS-related pathogen, Cryptococcus neoformans Here, we demonstrate that interaction of the inositol pyrophosphate, IP7, with the CDK inhibitor protein, Pho81, is instrumental in promoting fungal virulence. IP7-Pho81 interaction stabilizes Pho81 association with other CDK complex components to promote PHO pathway activation and phosphate acquisition. Our data demonstrating that blocking IP7-Pho81 interaction or preventing Pho81 production leads to a dramatic loss in fungal virulence, coupled with Pho81 having no homologue in humans, highlights Pho81 function as a potential target for the development of urgently needed antifungal drugs

Endothelin potentiates TRPV1 via ETA receptor-mediated activation of protein kinase C

© 2007 Plant et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution Licens

Raw proteomic data for Triplexed affinity reagents to sample the mammalian inositol pyrophosphate interactome

HCT116 and HEK293T cell lysates were enriched over different inositol polyphosphate affinity reagents. Both data sets are generated with Perseus 1.6.7.

Cyclic enecarbamates as precursors of alpha,beta-unsaturated iminium ions: reactivity and synthesis of 6,6-spirocyclic ring systems

The scalable synthesis of cyclic enecarbamates and their use as convenient precursors of α,β-unsaturated N-acyl iminium ions is reported. The newly developed route overcomes synthetic and reactivity difficulties in previously reported methods, is readily scaled up, and proceeds through stable intermediates suitable for long-term storage if required. Preliminary investigations probing the reactivity of cyclic α,β-unsaturated N-acyl iminium ions as dienophiles in Diels-Alder reactions and electrophilic alkylating agents are described. In the presence of Lewis and Brønsted acids, iminium precursor 22a underwent efficient Diels-Alder cycloaddition with a range of simple and complex dienes, culminating in the synthesis of 6,6-spirocyclic ring systems possessing the same relative stereochemistry as the spirocyclic imine present in the marine natural product gymnodimine 1.Zhanwei Wang, Niels Krogsgaard-Larsen, Benjamin Daniels, Daniel. P. Furkert and Margaret A. Brimbl

Total Synthesis of the Macrocyclic <i>N</i>‑Methyl Enamides Palmyrolide A and 2<i>S</i>‑Sanctolide A

Full details of the total syntheses of the initially reported and revised structures of the neuroprotective agent palmyrolide A are reported. The key macrocyclization step was achieved using a sequential ring-closing metathesis/olefin isomerization reaction. Furthermore, the total synthesis of the related macrolide (2<i>S</i>)-sanctolide A is reported. The synthesis used key elements from the synthesis of palmyrolide A, including the RCM/olefin isomerization sequence. The synthetic work described herein serves to facilitate the assignment of stereochemistry of the natural product sanctolide A and demonstrates the utility of this approach for the synthesis of macrocyclic tertiary enamide natural products

Hair growth induction by substance P

In vitro, some neuropeptides, including the tachykinin, substance P (SP), act as growth factors. The cyclic growth of the richly innervated hair follicle offers a model for probing such functions in a complex, developmentally regulated tissue interaction system under physiologic conditions. Dissecting the role of neuropeptides in this system may also reveal as yet obscure neural mechanisms of hair growth control. The neuropeptide-releasing neurotoxin, capsaicin was injected intradermally, or SP slow-release formulations were implanted subcutaneously in the back skin of C57BL/6 mice with all follicles in the resting stage of the hair cycle (telogen) in order to see whether this induced hair growth (anagen). In addition, the endogenous SP skin concentration and the activity of the main SP-degrading enzyme, neutral endopeptidase, were determined during the induced murine hair cycle by high performance liquid chromatography-controlled radioimmuno-assay (SP) or by fluorometry (neutral endopeptidase). Both capsaicin and SP induced significant hair growth (anagen) in the back skin of telogen mice. This was associated with substantial mast cell degranulation. The endogenous SP skin concentration showed significant, hair cycle-dependent fluctuations during the induced murine hair cycle, which were largely independent of the activity of neutral endopeptidase. SP may play a role in the neural control of hair growth. Whereas this pilot study does not address the underlying mechanisms of action, it demonstrates that SP has potential as a hair growth-stimulatory agent in vivo, and serves as a basis for exploring the role of tachykinins in epithelial-mesenchymal-neuroectodermal interaction systems like the hair follicle

Total Synthesis of the Initially Reported and Revised Structures of the Neuroprotective Agent Palmyrolide A

The total syntheses of the initially reported and revised structures of the neuroprotective agent palmyrolide A are reported. The key macrocyclization step was achieved using a sequential ring closing metathesis/olefin isomerization reaction. The synthetic work described herein serves to confirm the recent structural revision of this unusual natural product