1 Station Biologique, UMR 7127, CNRS et Université Pierre et Marie Curie, BP 74, F-29682 Roscoff cedex, France
2 Carnegie Institution of Washington, Department of Plant Biology, 260 Panama Street, Stanford, CA 94305, USA
Correspondence
Isabelle Mary
mary{at}sb-roscoff.fr
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ABSTRACT |
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INTRODUCTION |
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Cyanobacteria are an ancient (over 3 billion years old), diverse and ecologically important group of eubacteria that evolved to compete in a wide range of habitats. As the only prokaryotes that perform oxygenic photosynthesis, they are major contributors to biomass accumulation on Earth. Since their apparatus for photosynthesis resembles that of plants, they have been used as model organisms by many scientists to investigate the structure and function of plant-type photosynthesis. Synechocystis sp. PCC 6803 is a freshwater, unicellular, non-nitrogen-fixing cyanobacterium for which the complete nucleotide sequence of the genome has been determined (Kaneko et al., 1996). This naturally transformable strain is one of the most popular organisms for genetic and physiological studies of photosynthesis and environmental gene regulation because of its ability to grow both photoautotrophically and photoheterotrophically. The cyanobacterium Prochlorococcus, the smallest known photosynthetic prokaryote, is a dominant primary producer throughout the oligotrophic oceans (Partensky et al., 1999
). It is ubiquitous within the 40°S to 40°N latitudinal band and is found both in surface waters, experiencing light intensities as high as 1500 µmol photons m2 s1, and at 150 m depth, where light intensities fall below 1 µmol photons m2 s1. The wide distribution of Prochlorococcus in the marine environment probably reflects the existence of several physiologically and genetically distinct ecotypes (Moore et al., 1998
). High-light-adapted genotypes occupy the upper, well-illuminated but nutrient-poor part of the water column whereas low-light-adapted genotypes are preferentially found at the bottom of the illuminated layer but in a nutrient-rich environment. These ecotypes differ in their optimal light intensity for growth (Moore et al., 1998
), pigment content (Moore et al., 1995
), light harvesting efficiency, sensitivity to trace metals (Mann, 2002
) and nitrogen utilization ability (Moore et al., 2002
). Full genome information is available for Prochlorococcus MED4, which experiences growth in high light (HL) waters of the open oceans, and Prochlorococcus MIT9313, which typically grows in low light (LL) environments and is found deeper down in the water column (Rocap et al., 2003
).
Cyanobacteria possess stress-associated genes, encoding chaperones such as DnaK, DnaJ, GroEL or ClpB, and proteases such as ClpP (Glatz et al., 1999). A number of these genes may be induced when cells are shifted to high temperatures (Eriksson et al., 2001
) or from LL to HL. In Synechocystis sp. PCC 6803, clpB, htpG, dnaK, groES, groEL, groEL2 and hsp17 transcripts all appeared to accumulate when cells were transferred from 20 to 300 µmol photons m2 s1 (Hihara et al., 2001
). These results are consistent with experiments performed under similar conditions by Huang et al. (2002)
. Stress-associated multigene family members appear to be differentially expressed under a variety of physiological conditions (Lindquist & Craig, 1988
). As an example, cyanobacteria possess four clpP genes (Porankiewicz et al., 1999
; Schelin et al., 2002
). clpP1 is monocistronic, while clpP2 and clpP3, similar to the situation in Escherichia coli, appear to be arranged in an operon with clpX (Schelin et al., 2002
; Clarke et al., 1998
) and clpP4 (also called clpR, encoding a protein that lacks the three active-site amino acids representative of ClpP proteases), respectively (Schelin et al., 2002
). Unlike most bacterial Clp proteins, and especially ClpP1 (Clarke et al., 1998
), the ClpP2, ClpP3, ClpP4 and ClpX polypeptides did not accumulate to high levels upon cold shock, oxidative stress or HL exposure of Synechococcus sp. PCC 7942 (Schelin et al., 2002
). Similarly, while the DnaK2 protein of Synechococcus sp. PCC 7942 exhibited a typical increased accumulation following heat shock, the levels of DnaK1 and DnaK3 did not change following the same treatment (Nimura et al., 2001
).
In this study we examined the responses of the groES, groEL, dnaK and clp transcripts in the freshwater cyanobacterium Synechocystis sp. PCC 6803, the marine HL-adapted strain Prochlorococcus MED4 and the marine LL-adapted strain Prochlorococcus MIT9313 following a shift from LL to HL. Using real-time, quantitative RT-PCR, the levels of transcripts were quantified over both the short term (30 min, 1 h) and relatively longer term (3, 6 and 12 h) following the shift to HL, allowing the determination of those heat-shock genes with elevated levels of expression following exposure to HL stress.
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METHODS |
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Flow-cytometric analysis.
A 1 ml aliquot of the Prochlorococcus culture was sampled at each RNA sampling time following HL exposure for the analysis of flow-cytometric cell parameters (i.e. cell number, chlorophyll fluorescence) as described by Jacquet et al. (2001). Aliquots were fixed for 10 min with glutaraldehyde (0·25 % final concentration), frozen in liquid nitrogen and stored at 80 °C for later analysis. Samples were analysed with a FACSort flow cytometer (Becton Dickinson) with 488 nm excitation. Cell chlorophyll fluorescence was normalized to that of 0·95 µm fluorescent beads (Polysciences).
RNA isolation.
RNA was isolated from pelleted cells that had been frozen at 80 °C, using a modification of the method of de Saizieu et al. (1998), as described by Bhaya et al. (2000)
. Briefly, 500 µl acidified phenol and 500 µl NAES (50 mM sodium acetate pH 5·1, 10 mM EDTA, 1 % SDS) were added to pelleted cells from 150 ml cultures; 100 mg glass beads (0·1 µm mean diameter, Bio-Rad) was placed in the suspension, which was then vortexed three times for 20 s each. This was followed by two phenol/chloroform (1 : 1, v/v) and one chloroform extraction. Nucleic acids were precipitated with 2 vols ethanol, resuspended in sterile, double-distilled H2O and then treated for 1 h at room temperature with DNase I (20 U FPLC purified, Amersham), according to the protocol recommended by the manufacturer. DNase-treated samples were extracted with phenol/chloroform (1 : 1), then with chloroform, and the RNA was precipitated from the aqueous phase upon addition of 2 vols ethanol. The final RNA pellet was dissolved in 50 µl 10 mM Tris/HCl pH 8·0, 1 mM EDTA and stored at 80 °C.
Real-time quantitative RT-PCR analysis of gene expression.
For the reverse transcriptase (RT) reaction, 200 ng RNA was incubated with a mixture of PCR reverse primers (4 pmol per 20 µl final vol) (Table 1) for 10 min at 70 °C prior to adding 100 U Superscript II RT (Gibco-BRL). The RT reaction was performed at 42 °C for 1 h and stopped by placing the reaction at 72 °C for 10 min. SYBR Green PCR Master mix (Applied Biosystems) or LightCycler DNA Master SYBR Green I (Roche Applied Science) were used as recommended by the manufacturers; each reaction of 25 µl contained 1 µl of a specific, diluted cDNA (diluted 1/10 relative to the cDNA reaction mixture) and 0·4 µmol l1 of the appropriate primers.
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Transcript levels of the Synechocystis sp. PCC 6803 hsp genes were analysed by real-time RT-PCR using the LightCycler System (Roche Applied Science). The amplification programme consisted of 1 cycle of 95 °C for 60 s followed by 40 cycles of 95 °C for 15 s, 55 °C for 20 s, 72 °C for 40 s. The relative levels of transcripts of hsp genes of the Prochlorococcus strains were analysed by real-time RT-PCR using the ABI Prism 5700 sequence detection system (Applied Biosystems). The amplification programme consisted of 1 cycle of 95 °C with a 60 s hold followed by 40 cycles of 95 °C with a 15 s hold and 60 °C with a 1 min hold. The rps (slr1984) and rnpB transcripts were used as external standards, for Synechocystis and Prochlorococcus respectively, since levels of these transcripts were not altered by HL conditions (Mary & Vaulot, 2003) (Fig. 1
).
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RESULTS |
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Two genes encoding GroEL have been identified in the genome of the Prochlorococcus strains and Synechocystis sp. PCC 6803. The groEL2 gene is monocistronic and the groEL1 gene is part of a bicistronic operon with groES. In all of these strains, groEL1 and groES showed similar patterns of expression in terms of intensity and timing (Tables 35
). The strongest response was observed in Prochlorococcus MED4, while both Synechococystis PCC 6803 and Prochlorococcus MIT9313 exhibited moderate increases. Furthermore, the response occurred earlier (after 1 h) in Prochlorococcus MIT9313, in agreement with the cell response data. The groEL2 response was not as pronounced as that of groES/groEL1 and occurred later with respect to the transition time.
The second major chaperone system, the DnaK system, is encoded by multiple dnaK, dnaJ and grpE genes. In contrast to Synechocystis sp. PCC 6803, which has four dnaK genes (sll0086, sll0170, sll1932 and sll0058), the Prochlorococcus strains have only three. The polypeptides encoded by these genes in both Synechocystis sp. PCC 6803 and Prochlorococcus strains are very similar to those of analogous genes identified in Synechococcus sp. PCC 7942 (Nimura et al., 2001). The dnaK2 and dnaK3 genes are well conserved among the three strains. In contrast, the Prochlorococcus dnaK1 displayed low similarities to the analogous gene in Synechocystis sp. PCC 6803. Unlike Synechocystis sp. PCC 6803, which has four dnaJ genes (sll1666, sll1933, sll0093 and slr0897), Prochlorococcus MIT9313 and MED4 possess three and two dnaJ homologues, respectively. dnaJ3 shares strong identity (about 60 %) among the three strains, as does dnaJ2, which has 51 % identity between Synechocystis sp. PCC 6803 and Prochlorococcus MIT9313. In contrast, Prochlorococcus dnaJ1 is not well conserved (about 34 % identity) but may still be the paralogous gene to dnaJ1 of Synechocystis sp. PCC 6803 because it is located close to dnaK3 in both strains. Furthermore, grpE is in the same operon as dnaJ3 in the Prochlorococcus strains, whereas it is in an operon with a dnaK gene (sll0058) in Synechocystis sp. PCC 6803. Among the dnaK transcripts, that of dnaK2 increased strongly in all strains (Tables 3
5
). The dnaK1 and dnaK3 transcript levels increased moderately in Prochlorococcus MED4 and Synechocystis sp. PCC 6803. Some of these transcripts, including dnaK2 in Prochlorococcus MED4 and dnaJ3 in Prochlorococcus MIT9313, remained high even after 12 h of HL exposure. The dnaJ3 mRNA level increased dramatically in all strains, whereas dnaJ2 mRNA accumulated to a lesser extent in Prochlorococcus MIT9313. Furthermore, grpE mRNA accumulated only in Prochlorococcus MIT9313 (Tables 3
5
).
The clp family of genes in the three strains consists of single clpC and clpX genes and multiple clpB and clpP genes. The ClpP ATP-dependent protease is encoded by four distinct genes. The clpP1 gene appears to be monocistronic while clpP2 and clpP3 are each part of bicistronic operons with clpX and clpP4, respectively. Of the two clpB genes, only clpB1 mRNA levels rose following HL exposure of the three strains. As in Synechocystis sp. PCC 6803, the clpP1 transcript level markedly increased in Prochlorococcus MED4 (>10-fold), whereas clpP2, clpP3 and clpC mRNA accumulated to high levels in Prochlorococcus MED4 only. In contrast, many transcripts encoding Clp protease components, with the exception of clpB1 and clpB2, did not increase following HL exposure of Prochlorococcus MIT9313 (Tables 3 and 4).
Finally, the htpG transcript accumulated in the three strains but to a lesser extent in Prochlorococcus MED4 and Synechocystis PCC 6803 (Tables 35
).
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DISCUSSION |
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The transcripts for dnaJ2 and dnaJ4 do not increase strongly in Synechocystis sp. PCC 6803 during HL exposure, suggesting that they may not play a crucial role in this acclimation process. In contrast, the dnaK4 transcript increased moderately and the hsp17 transcript showed a strong increase in response to HL stress in Synechocystis sp. PCC 6803. Furthermore, the CIRCE (controlling inverted repeat of chaperone expression) binding repressor, HrcA, which is thought to regulate expression of some chaperone genes (Glatz et al., 1997), is absent from the Prochlorococcus MED4 genome but present in the two other strains, which suggests that another mechanism may be involved in the regulation of hsp expression in this HL-adapted strain.
Besides these differences, the three cyanobacterial strains share many features of hsp gene organization and induction. In many cases, the hsp transcripts increase in abundance soon after exposure to HL and then decline during the later stages of acclimation. This probably reflects the capacity of the cells to reach a homeostatic condition in which the physiology of the cell has been modified with respect to the capture and utilization of excitation energy under HL conditions. Such a change is illustrated by the lower chlorophyll fluorescence observed after 12 h of HL exposure in Prochlorococcus (Table 2). Interestingly, when Prochlorococcus MIT9313 cells were shifted from 6 to 40 µmol photons m2 s1, there was no significant increase in the level of most hsp transcripts, and little decrease in chlorophyll fluorescence, suggesting that low-amplitude changes in light levels do not induce drastic responses, and that the cells are able to quickly acclimate without major structural rearrangements.
Interestingly, the hsp genes that exhibit the strongest responses to HL exposure [i.e. groEL and groES (Tables 46
)] also show the most sustained accumulation of transcripts (over the 12 h period following transfer to HL). This is especially apparent with Prochlorococcus MED4 (Table 3
). The polypeptides encoded by such transcripts may be very important (either singly or in combinations) for maintenance of cellular function under specific stress conditions, such as HL. Among the three strains examined, groEL1 and groES are arranged in an operon and encode the most-conserved Hsps. In contrast, the GroEL2 polypeptide is much less conserved. The GroEL/GroES system is a major chaperone system in all bacteria and the ways in which this system responds to stress conditions have been extensively studied in cyanobacteria (Hihara et al., 2001
; Clarke & Eriksson, 1996
; Apte et al., 1998
). For example, the transcripts groEL1 and groEL2 have been shown to accumulate in response to elevated temperatures (Kovacs et al., 2001
). In all three strains groES and groEL1 showed somewhat similar levels and kinetics of mRNA accumulation, while the level of the groEL2 transcript increased later with respect to the HL exposure (Tables 3
5
). This suggests that the two groEL genes may be regulated differently, as hypothesized previously for Synechocystis sp. PCC 6803 (Rajaram et al., 2001
).
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Among the dnaK genes, the dnaK2 transcripts accumulated to a greater extent than those of the other dnaK genes (Tables 35
). This agrees with previous results showing that, upon shifting Synechococcus sp. PCC 7942 cells to elevated temperatures, the level of DnaK1 and DnaK3 polypeptides remained relatively constant while that of DnaK2 increased (Nimura et al., 2001
). Hihara et al. (2001)
demonstrated that HL caused increased accumulation of dnaK2 (sll0170) transcripts in Synechocystis sp. PCC 6803. This agrees with the observation that in most eukaryotes, Hsp70s (the DnaK homologues) are represented by a multigene family in which the different family members are differentially expressed (Lindquist & Craig, 1988
; Queitsch et al., 2000
).
In parallel to what was observed for the dnaK genes, dnaJ3 mRNA accumulated more than that of the other dnaJ transcripts following HL exposure in Prochlorococcus MED4 and MIT9313, and it was the only dnaJ that increased in HL in Synechocystis sp. PCC 6803. Another common feature of the three strains examined in this study is the contiguous genomic locations of dnaJ1 and dnaK3, with the former immediately downstream of the latter. Interestingly, while these transcripts show some increase in HL in the three cyanobacterial strains (with the least change in Synechocystis sp. PCC 6803), these increases are neither marked nor as sustained as observed for dnaK2 and dnaJ3 (Tables 35
). In Synechococcus sp. PCC 7942, the dnaJ1 homologue was shown to be essential for growth and was detected in thylakoid membranes (Nimura et al., 2001
), suggesting that DnaK3 and DnaJ1 might function cooperatively with respect to photosynthesis.
While the ClpPs and ClpC proteases also appear conserved in Synechocystis sp. PCC 6803 and the Prochlorococcus strains, the responses of these genes to HL is much more variable (Tables 35
). These proteases, one of the best-studied families in E. coli, are composed of regulatory, ATPase/chaperone polypeptides associated with protease subunits. The three cyanobacteria studied here have two different clpB genes, as in Synechococcus sp. PCC 7942 (Eriksson et al., 2001
). In the present study, clpB1 transcripts transiently increased while clpB2 mRNA levels did not change following the transfer to HL (Tables 3
5
). ClpB1 was shown to be essential for resistance to high- and low-temperature stress in Synechococcus elongatus (Eriksson & Clarke, 1996
; Porankiewicz & Clarke, 1997
). While ClpB2 has been suggested to be an essential constitutive protein with general chaperone activity (Eriksson et al., 2001
), our data indicate that ClpB1 might have a stress-related function. The clpP transcripts also accumulated in Synechocystis sp. PCC 6803 and Prochlorococcus MED4 after HL exposure, especially clpP1. Interestingly, all of the clp transcripts declined in Prochlorococcus MIT9313 following exposure to HL, except for clpP1. ClpP1 was previously shown to be induced strongly during acclimation of Synechococcus sp. PCC 7942 to either low temperature or UV-B, but was not induced by heat shock (Clarke et al., 1998
; Porankiewicz et al., 1998
). Similarly, neither ClpP2, ClpP3 nor ClpP4 (ClpR) was shown to increase upon heat or cold shock (Schelin et al., 2002
; Porankiewicz et al., 1998
), whereas all four Clp proteins were induced by HL intensities to different degrees, but again the most pronounced increase was for ClpP1 in Synechococcus sp. PCC 7942 (Clarke et al., 1998
). ClpX, ClpP3 and ClpR were found to be primarily constitutive proteins, critical for cell viability in this strain (Schelin et al., 2002
). These data suggest the complexity in the control and importance of the different clp genes in cyanobacteria in response to stress conditions. The arrangement of clp genes is conserved in the genomes of several cyanobacteria including Synechocystis sp. PCC 6803, Synechococcus sp. PCC 7942, Nostoc punctiforme and Prochlorococcus marinus (Schelin et al., 2002
). For example, the clpP2 gene is part of an operon with clpX and, therefore, ClpP2 may preferentially associate with ClpX and not with other Clp/Hsp100 proteins, such as ClpC (Porankiewicz et al., 1999
). In the present study, clpX mRNA did not accumulate in any of the three strains following HL exposure (Tables 3
5
), raising the possibility that ClpX in cyanobacteria may function as an independent chaperone that is not functionally linked to ClpP (Levchenko et al., 1995
).
The htpG transcript accumulated in all strains following exposure to HL (Tables 35
), which is consistent with recent microarray studies (Hihara et al., 2001
; Huang et al., 2002
). Moreover, it has been shown that HtpG is essential for survival during heat (Tanaka & Nakamoto, 1999
) and cold stress (Hossain & Nakamoto, 2002
), and is also involved in acclimation of Synechococcus sp. PCC 7942 to oxidative stress (Hossain & Nakamoto, 2003
).
Overall, hsp genes were induced to higher levels for longer periods after the light shift for Prochlorococcus MED4 than for either Prochlorococcus MIT9313 or Synechocystis sp. PCC 6803. For example, while in Synechocystis sp. PCC 6803 only the clpP1 transcript level rose substantially following exposure to HL and no clpP transcript accumulated in Prochlorococcus MIT9313, transcripts for all four of the clpPs strongly increased in Prochlorococcus MED4. Furthermore, while transcripts encoding ClpC (Kaneko et al., 1996; Clarke & Eriksson, 1996
) declined following HL exposure of Synechocystis sp. PCC 6803 (Eriksson et al., 2001
) and Prochlorococcus MIT9313, they showed a sustained increase in Prochlorococcus MED4. Prochlorococcus MED4 is found in ocean surface waters, where light intensities can reach up to 1500 µmol photons m2 s1 at midday, nutrient levels can be extremely low (Partensky et al., 1999
), and the cells probably experience oxidative stress. Our results for Prochlorococcus MED4 suggest that considerable protein misfolding and aggregation could take place in surface oceanic waters and that hsp genes could be critical for survival in such an extreme environment. Another example of adaptation of Prochlorococcus MED4 to a HL environment is the high number (22) of hli genes encoded by its genome relative to the genomes of the LL-adapted strains (Bhaya et al., 2002
); this gene family is critical for cyanobacterial survival during HL exposure (He et al., 2001
),
In conclusion, based on expression patterns, at least seven genes, groEL1/ES, groEL2, dnaK2, dnaJ3, clpB1 and clpP1, are suspected to play a key role in the acclimation of cyanobacteria to HL (Table 6). Interestingly, these genes are among the most conserved in the three strains considered. They should be targeted for more detailed analyses including protein expression and genetic studies.
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ACKNOWLEDGEMENTS |
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REFERENCES |
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Asada, K. (1994). Production and action of active oxygen species in photosynthetic tissues. In Causes of Photooxidative Stress and Amelioration of Defence Systems in Plants, pp. 77104. Edited by C. H. Foyer & P. M. Mullineaux. Boca Raton, FL: CRC Press.
Bhaya, D., Vaulot, D., Amin, P., Takahashi, A. & Grossman, A. (2000). Isolation of regulated genes of the cyanobacterium Synechocystis sp. strain PCC 6803 by differential display. J Bacteriol 182, 56925699.
Bhaya, D., Dufresne, A., Vaulot, D. & Grossman, A. (2002). Analysis of the hli gene family in marine and freshwater cyanobacteria. FEMS Microbiol Lett 215, 209219.[CrossRef][Medline]
Bukau, B. & Horwich, A. L. (1998). The Hsp70 and Hsp60 chaperone machines. Cell 92, 351366.[Medline]
Clarke, A. K. & Eriksson, M. J. (1996). The cyanobacterium Synechococcus sp. PCC 7942 possesses a close homologue to the chloroplast ClpC protein of higher plants. Plant Mol Biol 31, 721730.[Medline]
Clarke, A. K., Schelin, J. & Porankiewicz, J. (1998). Inactivation of the clpP1 gene for the proteolytic subunit of the ATP-dependent Clp protease in the cyanobacterium Synechococcus limits growth and light acclimation. Plant Mol Biol 37, 791801.[CrossRef][Medline]
De Saizieu, A., Certa, U., Warrington, J., Gray, C., Keck, W. & Mous, J. (1998). Bacterial transcript imaging by hybridization of total RNA to oligonucleotide arrays. Nat Biotechnol 16, 4548.[CrossRef][Medline]
Eriksson, M. J. & Clarke, A. K. (1996). The heat shock protein ClpB mediates the development of thermotolerance in the cyanobacterium Synechococcus sp. strain PCC 7942. J Bacteriol 178, 48394846.[Abstract]
Eriksson, M. J., Schelin, J., Miskiewicz, E. & Clarke, A. K. (2001). Novel form of ClpB/HSP100 protein in the cyanobacterium Synechococcus. J Bacteriol 183, 73927396.
Glatz, A., Horvath, I., Varvasovszki, V., Kovacs, E., Torok, Z. & Vigh, L. (1997). Chaperonin genes of the Synechocystis PCC 6803 are differentially regulated under light-dark transition during heat stress. Biochem Biophys Res Commun 239, 291297.[CrossRef][Medline]
Glatz, A., Vass, I., Los, D. A. & Vigh, L. (1999). The Synechocystis model of stress: from molecular chaperones to membranes. Plant Physiol Biochem 37, 112.[CrossRef]
Glover, J. R. & Lindquist, S. (1998). Hsp104, Hsp70, and Hsp40: a novel chaperone system that rescues previously aggregated proteins. Cell 94, 7382.[Medline]
Gottesman, S. (1996). Proteases and their targets in Escherichia coli. Annu Rev Genet 30, 465506.[CrossRef][Medline]
Gottesman, S., Wickner, S. & Maurizi, M. R. (1997). Protein quality control: triage by chaperones and proteases. Genes Dev 11, 815823.[CrossRef][Medline]
Gottesman, S., Roche, E., Zhou, Y. & Sauer, R. T. (1998). The ClpXP and ClpAP proteases degrade proteins with carboxy-terminal peptide tails added by the SsrA-tagging system. Genes Dev 12, 13381347.
He, Q., Dolganov, N., Bjorkman, O. & Grossman, A. R. (2001). The high light-inducible polypeptides in Synechocystis PCC6803. Expression and function in high light. J Biol Chem 276, 306314.
Herman, C. & D'Ari, R. (1998). Proteolysis and chaperones: the destruction/reconstruction dilemma. Curr Opin Microbiol 1, 204209.[CrossRef][Medline]
Hihara, Y., Kamei, A., Kanehisa, M., Kaplan, A. & Ikeuchi, M. (2001). DNA microarray analysis of cyanobacterial gene expression during acclimation to high light. Plant Cell 13, 793806.
Hossain, M. M. & Nakamoto, H. (2002). HtpG plays a role in cold acclimation in cyanobacteria. Curr Microbiol 44, 291296.[CrossRef][Medline]
Hossain, M. M. & Nakamoto, H. (2003). Role for the cyanobacterial HtpG in protection from oxidative stress. Curr Microbiol 46, 7076.[CrossRef][Medline]
Huang, L., McCluskey, M. P., Ni, H. & LaRossa, R. A. (2002). Global gene expression profiles of the cyanobacterium Synechocystis sp. strain PCC 6803 in response to irradiation with UV-B and white light. J Bacteriol 184, 68456858.
Jacquet, S., Partensky, F., Marie, D., Casotti, R. & Vaulot, D. (2001). Cell cycle regulation by light in Prochlorococcus strains. Appl Environ Microb 67, 782790.
Kaneko, T., Sato, S., Kotani, H. & 21 other authors (1996). Sequence analysis of the genome of the unicellular cyanobacterium Synechocystis sp. strain PCC6803. II. Sequence determination of the entire genome and assignment of potential protein-coding regions. DNA Res 3, 109136.[Medline]
Kovacs, E., van der Vies, S. M., Glatz, A., Torok, Z., Varvasovszki, V., Horvath, I. & Vigh, L. (2001). The chaperonins of Synechocystis PCC 6803 differ in heat inducibility and chaperone activity. Biochem Biophys Res Commun 289, 908915.[CrossRef][Medline]
Levchenko, I., Luo, L. & Baker, T. (1995). Disassembly of the Mu transposase tetramer by the ClpX chaperone. Genes Dev 9, 23992408.[Abstract]
Lindquist, S. & Craig, E. A. (1988). The heat-shock proteins. Annu Rev Genet 22, 631677.[CrossRef][Medline]
Mann, N. H. (2002). Phages of the marine cyanobacterial picophytoplankton. FEMS Microbiol Lett 27, 1734.
Mary, I. & Vaulot, D. (2003). Two-component systems in Prochlorococcus MED4: genomic analysis and differential expression under stress. FEMS Microbiol Lett 226, 135144.[CrossRef][Medline]
Moore, L. R., Goericke, R. & Chisholm, S. W. (1995). Comparative physiology of Synechococcus and Prochlorococcus: influence of light and temperature on growth, pigments, fluorescence and absorptive properties. Mar Ecol Prog Ser 116, 259275.
Moore, L. R., Rocap, G. & Chisholm, S. W. (1998). Physiology and molecular phylogeny of coexisting Prochlorococcus ecotypes. Nature 393, 464467.[CrossRef][Medline]
Moore, L. R., Post, A. F., Rocap, G. & Chisholm, S. W. (2002). Utilization of different nitrogen sources by the marine cyanobacteria Prochlorococcus and Synechococcus. Limnol Oceanogr 47, 98996.
Nimura, K., Takahashi, H. & Yoshikawa, H. (2001). Characterization of the dnaK multigene family in the cyanobacterium Synechococcus sp. strain PCC7942. J Bacteriol 183, 13201328.
Partensky, F., Hess, W. R. & Vaulot, D. (1999). Prochlorococcus, a marine photosynthetic prokaryote of global significance. Microbiol Mol Biol Rev 63, 106127.
Porankiewicz, J. & Clarke, A. K. (1997). Induction of the heat shock protein ClpB affects cold acclimation in the cyanobacterium Synechococcus sp. strain PCC 7942. J Bacteriol 179, 51115117.[Abstract]
Porankiewicz, J., Schelin, J. & Clarke, A. K. (1998). The ATP-dependent Clp protease is essential for acclimation to UV-B and low temperature in the cyanobacterium Synechococcus. Mol Microbiol 29, 275283.[CrossRef][Medline]
Porankiewicz, J., Wang, J. & Clarke, A. (1999). New insights into the ATP-dependent Clp protease: Escherichia coli and beyond. Mol Microbiol 32, 449458.[CrossRef][Medline]
Queitsch, C., Hong, S. W., Vierling, E. & Lindquist, S. (2000). Heat shock protein 101 plays a crucial role in thermotolerance in Arabidopsis. Plant Cell 12, 479492.
Rajaram, H., Ballal, A. D., Apte, S. K., Wiegert, T. & Schumann, W. (2001). Cloning and characterization of the major groESL operon from a nitrogen-fixing cyanobacterium, Anabaena sp. strain L-31. Biochim Biophys Acta 1519, 143146.[Medline]
Rippka, R., Deruelles, J., Waterbury, J. B., Herdman, M. & Stanier, R. Y. (1979). Generic assignments, strain histories and properties of pure cultures of cyanobacteria. J Gen Microbiol 111, 161.
Rocap, G., Larimer, F. W., Lamerdin, J. & 21 other authors (2003). Genome divergence in two Prochlorococcus ecotypes reflects oceanic niche differentiation. Nature 424, 10011002.[CrossRef][Medline]
Schelin, J., Lindmark, F. & Clarke, A. K. (2002). The clpP multigene family for the ATP-dependent Clp protease in the cyanobacterium Synechococcus. Microbiology 148, 22552265.
Tanaka, N. & Nakamoto, H. (1999). HtpG is essential for the thermal stress management in cyanobacteria. FEBS Lett 458, 117123.[CrossRef][Medline]
Received 8 January 2004;
accepted 13 January 2004.