DEEP RNA SEQUENCING ANALYSIS OF THE ANTARCTIC BACTERIUM PSEUDOMONAS EXTREMAUSTRALIS IN RESPONSE TO LOW TEMPERATURE.

TRIBELLI, P.M.; RICARDI,M.M.; MOLIN, S.; LÓPEZ, N.I.

Congreso; X congreso de Microbiología General (SAMIGE); 2014

In cold environments low temperatures and ice presence exerts severe constraints on living organisms and those able to survive present several adaptations to cope with these unfavorable conditions. Environmental adaptability involves different physiological and genetic strategies including the response of individual genes or operons and complex regulatory networks that coordinate the control of several genes. RNA-deep sequence is a powerful tool in bacterial species to analyze gene expression, discover previously unannotated genes and detect small regulatory RNA. We used RNA-seq technology in Pseudomonas extremaustralis, an Antarctic bacterium able to grow under low temperatures and survive to freezing. Total RNA was extracted from cultures grown at 30oC and 5oC and the rRNA was depleted from the samples to allow a better coverage. Directional libraries were prepared with ScriptSeq v2RNA-Seq Library Preparation Kit (Epicentre) and were sequenced using the Illumina X HiSeq2000 platform with a paired-end protocol and read lengths of 100 nt. For each condition duplicated independent RNA extraction and libraries were used. Bioinformatics analysis of around 6000 transcripts expression levels in both conditions using Rockhoper software showed that the majority of transcripts did not present a statistical significant change in the expression level. Down-regulated genes were around 750 and included cytochromes and enzymes belonging to different catabolic pathways. Interestingly, 76 genes were up-regulated under cold conditions including several lipoproteins, transcriptional regulators with unknown function, alginate biosynthesis activator, osmotic response elements and other cellular functions. We also detected several unnanotated transcripts in both conditions. Additionally, we have found around 170 putative non-coding small RNAs. Some of them presented altered expression levels under low temperatures with unknown function. The results showed a novel variety of transcripts and regulatory elements that could explain growth and survival under low temperatures in this psycrotolerant bacterium.