Revealing the sequence of interactions of PuroA peptide with Candida albicans cells by live-cell imaging
| Autor: | Mrinal Bhave, Nadin Shagaghi, Andrew H. A. Clayton, Enzo A. Palombo |
|---|---|
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Fluorescence-lifetime imaging microscopy Time Factors 030106 microbiology Antimicrobial peptides Peptide Time-Lapse Imaging Article Microbiology 03 medical and health sciences chemistry.chemical_compound Live cell imaging Candida albicans Fluorescence microscope Propidium iodide chemistry.chemical_classification Microbial Viability Multidisciplinary biology Chemistry biology.organism_classification 030104 developmental biology Microscopy Fluorescence Biophysics Intracellular Antimicrobial Cationic Peptides |
| Zdroj: | Scientific Reports |
| ISSN: | 2045-2322 |
| DOI: | 10.1038/srep43542 |
| Popis: | To determine the mechanism(s) of action of antimicrobial peptides (AMPs) it is desirable to provide details of their interaction kinetics with cellular, sub-cellular and molecular targets. The synthetic peptide, PuroA, displays potent antimicrobial activities which have been attributed to peptide-induced membrane destabilization, or intracellular mechanisms of action (DNA-binding) or both. We used time-lapse fluorescence microscopy and fluorescence lifetime imaging microscopy (FLIM) to directly monitor the localization and interaction kinetics of a FITC- PuroA peptide on single Candida albicans cells in real time. Our results reveal the sequence of events leading to cell death. Within 1 minute, FITC-PuroA was observed to interact with SYTO-labelled nucleic acids, resulting in a noticeable quenching in the fluorescence lifetime of the peptide label at the nucleus of yeast cells, and cell-cycle arrest. A propidium iodide (PI) influx assay confirmed that peptide translocation itself did not disrupt the cell membrane integrity; however, PI entry occurred 25–45 minutes later, which correlated with an increase in fractional fluorescence of pores and an overall loss of cell size. Our results clarify that membrane disruption appears to be the mechanism by which the C. albicans cells are killed and this occurs after FITC-PuroA translocation and binding to intracellular targets. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|