Life in the Cold: a Proteomic Study of Cold-Repressed Proteins in the Antarctic Bacterium Pseudoalteromonas haloplanktis TAC125

Abstract

The proteomes expressed at 4°C and 18°C by the psychrophilic Antarctic bacterium Pseudoalteromonas haloplanktis were compared using two-dimensional differential in-gel electrophoresis with special reference to proteins repressed by low temperatures. Remarkably, the major cold-repressed proteins, almost undetectable at 4°C, were heat shock proteins involved in folding assistance.The Gram-negative bacterium Pseudoalteromonas haloplanktis is a typical representative of the Gammaproteobacteria found in cold marine environments, and strain TAC125 has been isolated from seawater sampled along the Antarctic ice shelf. Such strains thrive permanently in seawater at about Ϫ2°C to ϩ4°C but are also anticipated to endure long-term frozen conditions when entrapped in the winter ice pack. The genome of P. haloplanktis TAC125 has been fully sequenced (11). This work has further allowed a proteomic study of its cold acclimation proteins (CAPs), i.e., proteins that are continuously overexpressed at a high level during growth at low temperatures (14). This has demonstrated that protein synthesis and protein folding are the main upregulated functions, suggesting that both cellular processes are limiting factors for bacterial development in cold environments. Here we report a proteomic survey of cold-repressed proteins at 4°C in order to complete the metabolic pattern of the bacterium's growth at low temperature.Temperature dependence of growth. The Antarctic bacterium maintains a doubling time of ϳ4 h at 4°C in a marine broth, with an extrapolated generation time of 5 h 15 min at 0°C (Fig. 1a). When the culture temperature is raised to 20°C, the generation time decreases moderately (e.g., 1 h 40 min at 18°C) with a concomitant increase in the biomass produced at the stationary phase (Fig. 1b). At temperatures higher than 20°C, a drastic reduction in cell density at the stationary phase is recorded (Fig. 1b), indicating heat-induced stress on the cell. P. haloplanktis fails to grow above 29°C. According to this growth behavior, the temperatures of 4°C and 18°C were selected here for the differential comparison of the proteomes, as 18°C does not induce excessive stress as far as growth rate and biomass are concerned.Cold-induced versus cold-repressed proteins. The proteomes expressed by the Antarctic bacterium at 4°C and 18°C during the logarithmic phase of growth were compared by two-dimensional (2D) differential in-gel electrophoresis as described previously (14). In a typical single 2D gel (see Fig. S1 in the supplemental material), 142 protein spots were more abundant at 4°C (CAPs), whereas 309 protein spots were less intense at 4°C than at 18°C. This unexpectedly large number of cold-repressed proteins already indicates that numerous cellular functions are downregulated during growth at low temperature. The repression factors (or induction factors for CAPs), given by relative spot abundance between 4°C and 18°C, are illustrated in Fig. 2. This distribution shows that 21% of coldrepressed proteins display a downregulation facto...

Is there a cold shock response in the Antarctic psychrophile Pseudoalteromonas haloplanktis?

“…For instance, the Antarctic bacterium P. haloplanktis grown at low temperature overexpresses enzymes involved in protein synthesis and folding and regulates its cytoplasmic redox balance as a result of improved oxygen solubility. Furthermore, the psychrophilic bacterium strongly represses the synthesis of heat shock chaperones, depresses its general metabolism and avoids pathways involving metal ions that are prone to reactive oxygen species production (Piette et al 2010(Piette et al , 2011a. However, for all psychrophiles studied so far, proteomes were compared using cells growing sustainably at a low and a high temperature.…”

Life in the Cold: Proteomics of the Antarctic Bacterium Pseudoalteromonas haloplanktis

“…Proteomic studies performed on psychrophilic microorganisms. References: 1, Seo et al, 2004;2, Goodchild et al, 2004a;3, Goodchild et al, 2004b;4, Goodchild et al, 2005;5, Saunders et al, 2005;6, Saunders et al, 2006;7, Qiu et al, 2006;8, Kawamoto et al, 2007;9, Bakermans et al, 2007;10, Zheng et al, 2007;11, Tunsjo et al, 2007;12, Bergholz et al, 2009;13, Ting et al, 2010;14, Williams et al, 2010;15, Garnier et al, 2010;16, Piette et al, 2010;Piette et al, 2011;17, Williams et al, 2011;18, Mykytczuk et al, 2011. identification of enzymes belonging to the pentose phosphate pathway suggest that regulation of the cytoplasmic redox buffering capacity via glutathione is a key component.…”

“…However, about 20% of these differentially expressed proteins display up-or down-regulation factors higher than 5, revealing that some key cellular functions are strongly regulated. Amongst all these differentially expressed proteins, 40 CAPs and 83 CRPs were retained, which satisfied both statistical biological variation analysis and mass spectrometry identification scores, as detailed in the original publications (Piette et al, 2010;Piette et al, 2011). Accordingly, the identified proteins should be analyzed as markers of a pathway or of a general function, rather than for their specific function as they represent 27% of the differentially expressed proteins at 4°C.…”

“…Spot views on 2D-gels (circled) and three-dimensional images. Adapted with permission from Piette et al, 2010;Piette et al, 2011. © 2010Wiley and © 2011 American Society for Microbiology.…”

“…This has demonstrated that protein synthesis and protein folding are the main up-regulated functions, suggesting that both cellular processes are limiting factors for bacterial development in cold environments. Furthermore, a proteomic survey of cold-repressed proteins at 4 °C has revealed a strong repression of most heat shock proteins (Piette et al, 2011). This chapter describes the various proteomic features analyzed in the context of adaptation to life at low temperature.…”