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Jani, N.

Publications and source records attributed to Jani, N..

2 recordsLinked to original sources
bioRxiv2018

Rapid Paediatric Sequencing (RaPS): Comprehensive real-life workflow for rapid diagnosis of critically ill children

BackgroundRare genetic conditions are frequent risk factors for, or direct causes of, organ failure requiring paediatric intensive care unit (PICU) support. Such conditions are frequently suspected but unidentified at PICU admission. Compassionate and effective care is greatly assisted by definitive diagnostic information. There is therefore a need to provide a rapid genetic diagnosis to inform clinical management.\n\nTo date, Whole Genome Sequencing (WGS) approaches have proved successful in diagnosing a proportion of children with rare diseases, but results may take months to report or require the use of equipment and practices not compatible with a clinical diagnostic setting. We describe an end-to-end workflow for the use of rapid WGS for diagnosis in critically ill children in a UK National Health Service (NHS) diagnostic setting.\n\nMethodsWe sought to establish a multidisciplinary Rapid Paediatric Sequencing (RaPS) team for case selection, trio WGS, a rapid bioinformatics pipeline for sequence analysis and a phased analysis and reporting system to prioritise genes with a high likelihood of being causal. Our workflow was iteratively developed prospectively during the analysis of the first 10 children and applied to the following 14 to assess its utility.\n\nFindingsTrio WGS in 24 critically ill children led to a molecular diagnosis in ten (42%) through the identification of causative genetic variants. In three of these ten individuals (30%) the diagnostic result had an immediate impact on the individuals clinical management. For the last 14 trios, the shortest time taken to reach a provisional diagnosis was four days (median 7 days).\n\nInterpretationRapid WGS can be used to diagnose and inform management of critically ill children using widely available off the shelf products within the constraints of an NHS clinical diagnostic setting. We provide a robust workflow that will inform and facilitate the rollout of rapid genome sequencing in the NHS and other healthcare systems globally.\n\nFundingThe study was funded by NIHR GOSH/UCL BRC: ormbrc-2012-1

genomics↗
bioRxiv2017

Phosphate Is The Third Nutrient Monitored By TOR In Candida albicans And Provides A Target For Fungal-Specific Indirect TOR Inhibition

The TOR pathway regulates morphogenesis and responses to host cells in the fungal pathogen Candida albicans. Eukaryotic TOR complex 1 (TORC1) induces growth and proliferation in response to nitrogen and carbon source availability. Our unbiased genetic approach seeking new components of TORC1 signaling in C. albicans revealed that the phosphate transporter Pho84 is required for normal TORC1 activity. We found that mutants in PHO84 are hypersensitive to rapamycin and, in response to phosphate feeding, generate less phosphorylated ribosomal protein S6 (P-S6) than wild type. The small GTPase Gtr1, a component of the TORC1-activating EGO complex, links Pho84 to TORC1. Mutants in Gtr1, but not in another TORC1-activating GTPase, Rhb1, are defective in the P-S6 response to phosphate. Overexpression of Gtr1 and of a constitutively active Gtr1Q67L mutant suppress TORC1-related defects. In S. cerevisiae pho84 mutants, constitutively active Gtr1 suppresses a TORC1 signaling defect but does not rescue rapamycin hypersensitivity. Hence connections from phosphate homeostasis to TORC1 may differ between C. albicans and S. cerevisiae. The converse direction of signaling, from TORC1 to the phosphate homeostasis (PHO) regulon, previously observed in S. cerevisiae, was genetically demonstrated in C. albicans using conditional TOR1 alleles. A small molecule inhibitor of Pho84, an FDA-approved drug, inhibits TORC1 signaling and potentiates the activity of the antifungals amphotericin B and micafungin. Anabolic TORC1-dependent processes require significant amounts of phosphate. Our study demonstrates that phosphate availability is monitored and also controlled by TORC1, and that TORC1 can be indirectly targeted by inhibiting Pho84.\n\nSignificanceThe human fungal pathogen Candida albicans uses the TOR signaling pathway to contend with varying host environments and thereby regulate cell growth. Seeking novel components of the C. albicans TOR pathway we identified a cell-surface phosphate importer, Pho84, and its molecular link to TOR complex 1 (TORC1). Since phosphorus is a critical element for anabolic processes like DNA replication, ribosome biogenesis, translation and membrane biosynthesis, TORC1 monitors its availability in regulating these processes. By depleting the central kinase in the TORC1 pathway, we showed that TORC1 signaling modulates regulation of phosphate acquisition. An FDA-approved small-molecule inhibitor of Pho84 inhibits TORC1 signaling and potentiates the activity of the gold-standard antifungal amphotericin B and the echinocandin micafungin.

microbiology↗