PARENT SESSION

Symposium S3D Genomic, proteomic and related technologies
Tuesday August 31st, 2004 10:20 AM-12:20 PM Room 510B
Chair: Arthur Grossman
Co-Chair: Jacques Joyard

The response of Synechocystis sp. PCC 6803 to nitrogen starvation: transcriptomics versus proteomics. Vladimir Krasikov^*,1, Henk Dekker², Hans Matthijs¹, ¹ Universiteit van Amsterdam, Amsterdam, the Netherlands² Universiteit van Amsterdam, Amsterdam, the Netherlands

ABSTRACT- Nitrogen is one of the nutrients that is playing an essential role in metabolism and ultimately is needed for the growth of cyanobacteria. To survive in nutrient limited conditions, cells should mobilise systems for high affinity uptake of limiting nutrients, adapt physiological processes to enable more economic usage of the nutrient concerned, and strive to be able to utilise non-traditional resources. Recently, an effective technique called DNA microarray has been developed and used to monitor gene expression in cyanobacteria. Classical proteomics including two-dimensional gel electrophoresis for separation and mass spectrometry for qualitative analysis of proteins in complex mixtures has been successfully applied to evaluate stress responses in cyanobacteria. However, overlaying of DNA array results with proved expression of proteins has not been reported so far. Protein expression profiles in Synechocystis sp. PCC 6803 cells, in response to relatively short nitrogen starvation, were determined using the cleavable isotope-coded affinity tag (cICAT) labeling strategy. The analysis included separation of the mixed protein samples by SDS-PAGE (total proteins isolated from cells growing 12 hours in nitrogen free medium were mixed in proportion 1:1 with proteins isolated from normally grown culture), followed by excision of regions from an entire gel lane. Proteins were subjected to in-gel digestion, biotin affinity chromatography, and analysis by nano-scale microcapillary liquid chromatography coupled to tandem mass spectrometry. The comparison of the global transcriptome analysis with qualitative and quantitative proteomics of cyanobacterial cells exposed to nitrogen starvation will be discussed.

KEY WORDS: microarray, cICAT, Nitrogen starvation, Synechocystis sp. PCC 6803