Abstract
Prokaryotic and eukaryotic cells exhibit an intrinsic natural fluorescence due to the presence of fluorescent cellular structural components and metabolites. Therefore, cellular autofluorescence (AF) is expected to vary with the metabolic states of cells. We examined how exposure to the different stressors changes the AF of Escherichia coli cells. We observed that bactericidal treatments increased green cellular AF, and that de novo protein synthesis was required for the observed AF increase. Excitation and emission spectra and increased expression of the genes from the flavin biosynthesis pathway, strongly suggested that flavins are major contributors to the increased AF. An increased expression of genes encoding diverse flavoproteins which are involved in energy production and ROS detoxification, indicates a cellular strategy to cope with severe stresses. An observed increase in AF under stress is an evolutionary conserved phenomenon as it occurs not only in cells from different bacterial species, but also in yeast and human cells.
Highlights
All prokaryotic and eukaryotic cells exhibit an intrinsic natural fluorescence due to the presence of different fluorescent cellular structural components and metabolites, such as flavins, nicotinamide-adenine dinucleotide (NAD), aromatic amino acids, lipofuscins, advanced glycation end products, and collagen[1,2]
We found that treatment of E. coli with the ß-lactam antibiotic ampicillin or with sodium hypochlorite, significantly increased cellular AF, while no significant AF increase was observed in protein synthesis inhibitor-treated cells
We found that bactericidal stressors, such as ß-lactam ampicillin, significantly increased cellular AF and that de novo protein synthesis is required for the AF increase
Summary
All prokaryotic and eukaryotic cells exhibit an intrinsic natural fluorescence (autofluorescence; AF) due to the presence of different fluorescent cellular structural components and metabolites, such as flavins, nicotinamide-adenine dinucleotide (NAD), aromatic amino acids, lipofuscins, advanced glycation end products, and collagen[1,2]. Flavins, NAD, and lipofuscin emit green, blue, and orange light respectively when excited at appropriate wavelengths. For this reason, AF frequently overlaps with the spectrum of exogenous fluorophores used for research purposes, and interferes with the fluorescent microscopy and cytometric analyses. AF precludes the detection of weak signals from the fluorescent reporters for low-abundance proteins Correction of this “contaminating” AF is problematic because it is frequently unevenly distributed within and between cells. Flavins and NAD are extensively studied because they are responsible for most of the cytoplasmic AF and because of their prominent role in cell metabolism. We demonstrated that the increase in green cellular AF subjected to life-threating treatments is an evolutionary conserved phenomenon as it occurs in cells from different bacterial species and in yeast and human cells
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