Proteomics of Shewanella oneidensis MR-1 Subjected to Solar UV radiation Stress
EMSL Project ID
10294
Abstract
Solar ultraviolet radiation (UVR) is lethal and potentially mutagenic to all organisms at species-specific levels. The stratospheric ozone layer absorbs UVC (<290 nm) effectively, however, both UVA (320 to 400 nm) and UVB (290 to 320nm) wavelengths penetrate to the earth’s surface. Terrestrial UVR contains about 5% UVB and 95% of UVA. UVB can cause direct DNA damage by inducing the formation of DNA photoproducts and single or double DNA strands breaks, which can be lethal through the blockage of DNA replication and transcription. UVA can cause photodamage to a variety of molecules as well as physiological processes directly or indirectly by inducing the production of reactive oxygen species (ROS). Shewanella oneidensis MR-1 is uniformly sensitive to UVC, UVB, UVA, solar UVR as well as ionizing radiation. Microarray-based gene expression profiles have been carried out for each component of solar UV as well as natural solar light. The largest cellular response was observed in MR-1 after exposing to solar light. More than one thousand genes showed significant differentiation (P<0.01) with more than 2 fold change in gene expression level, of which about 553 genes were induced and 884 genes were repressed during a 1 h recovery period after exposure. The differentially expressed genes were distributed in 19 functional categories. Both conserved hypothetical proteins (258 in total) and hypothetical proteins (195 in total) were dominant groups among either up- or down-regulated genes. In addition, more genes involved in transcription, biosynthesis, metabolism and membrane transport were repressed than induced. This unique transcriptional profile indicated a unique global cellular response to natural solar UVR compared to UVB or UVA. Proteomic work will enhance our understanding of this complicated biological process in MR-1 in response to solar UV stress.
MR-1 will be grown in Davis medium with 15 mM lactate as carbon source until OD600 reaches 0.25. The culture will split into two parts in a volume of 250 ml, one is used as a dark control and the other one will be exposed to solar light with a dose of 558 J m-2 of solar UVB. Both control and solar UV treated samples will be returned to a shaker at 30 ?C and incubated for 20 min. The cells will be collected by centrifugation and washed three times with PBS. During the last wash, the cell suspension will be resuspended in 5 ml of PBS and aliquoted in 1 ml. The cell pellets will be stored at -80?C and shipped to PNNL on dry ice for proteomic analysis.
We would would like to access is the 9 tesla FTICR (3 weeks), the 11 telsa FTICR (3 weeks) and the LCQ instruments (7 weeks)
Project Details
Project type
Exploratory Research
Start Date
2004-08-10
End Date
2006-08-18
Status
Closed
Released Data Link
Team
Principal Investigator