The most recent genome-editing system called CRISPR-Cas9 (clustered regularly interspaced short palindromic repeat system with associated protein 9-nuclease) was employed to delete four non-essential genes (i.e., Caeco1, Caidh1, Carom2, and Cataf10) individually to establish their gene functionality annotations in pathogen Candida albicans. The biological roles of these genes were investigated with respect to the cell wall integrity and biogenesis, calcium/calcineurin pathways, susceptibility of mutants towards temperature, drugs and salts. All the mutants showed increased vulnerability compared to the wild-type background strain towards the cell wall-perturbing agents, (antifungal) drugs and salts. All the mutants also exhibited repressed and defective hyphal growth and smaller colony size than control CA14. The cell cycle of all the mutants decreased enormously except for those with Carom2 deletion. The budding index and budding size also increased for all mutants with altered bud shape. The disposition of the mutants towards cell wall-perturbing enzymes disclosed lower survival and more rapid cell wall lysis events than in wild types. The pathogenicity and virulence of the mutants was checked by adhesion assay, and strains lacking rom2 and eco1 were found to possess the least adhesion capacity, which is synonymous to their decreased pathogenicity and virulence.
- MeSH
- acetyltransferasy nedostatek genetika fyziologie MeSH
- antifungální látky farmakologie MeSH
- buněčná adheze MeSH
- buněčná stěna účinky léků MeSH
- buněčný cyklus MeSH
- Candida albicans účinky léků genetika patogenita fyziologie MeSH
- chitinasy farmakologie MeSH
- CRISPR-Cas systémy MeSH
- delece genu MeSH
- endo-1,3-beta-glukanasa farmakologie MeSH
- faktory asociované s proteinem vázajícím TATA box nedostatek genetika fyziologie MeSH
- fungální proteiny genetika fyziologie MeSH
- geny hub * MeSH
- hyfy růst a vývoj MeSH
- isocitrátdehydrogenasa nedostatek genetika fyziologie MeSH
- kationty farmakologie MeSH
- nepohlavní rozmnožování MeSH
- otevřené čtecí rámce MeSH
- poškození DNA MeSH
- vápník fyziologie MeSH
- virulence genetika MeSH
- Publikační typ
- časopisecké články MeSH
Severe progressive neurological paediatric disease mucopolysaccharidosis III type C is caused by mutations in the HGSNAT gene leading to deficiency of acetyl-CoA: α-glucosaminide N-acetyltransferase involved in the lysosomal catabolism of heparan sulphate. To understand the pathophysiology of the disease we generated a mouse model of mucopolysaccharidosis III type C by germline inactivation of the Hgsnat gene. At 6-8 months mice showed hyperactivity, and reduced anxiety. Cognitive memory decline was detected at 10 months and at 12-13 months mice showed signs of unbalanced hesitant walk and urinary retention. Lysosomal accumulation of heparan sulphate was observed in hepatocytes, splenic sinus endothelium, cerebral microglia, liver Kupffer cells, fibroblasts and pericytes. Starting from 5 months, brain neurons showed enlarged, structurally abnormal mitochondria, impaired mitochondrial energy metabolism, and storage of densely packed autofluorescent material, gangliosides, lysozyme, phosphorylated tau, and amyloid-β. Taken together, our data demonstrate for the first time that deficiency of acetyl-CoA: α-glucosaminide N-acetyltransferase causes lysosomal accumulation of heparan sulphate in microglial cells followed by their activation and cytokine release. They also show mitochondrial dysfunction in the neurons and neuronal loss explaining why mucopolysaccharidosis III type C manifests primarily as a neurodegenerative disease.
- MeSH
- acetyltransferasy nedostatek genetika MeSH
- chování zvířat MeSH
- energetický metabolismus fyziologie MeSH
- gangliosidy metabolismus MeSH
- glykosaminoglykany metabolismus MeSH
- mitochondriální nemoci etiologie patologie MeSH
- mukopolysacharidóza III komplikace patologie psychologie MeSH
- myši inbrední C57BL MeSH
- myši MeSH
- neuritida etiologie patologie MeSH
- neurodegenerativní nemoci etiologie patologie psychologie MeSH
- neurologické vyšetření MeSH
- poruchy proteostázy patologie MeSH
- proteiny asociované s mikrotubuly metabolismus MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
