Versatile Genetic Tool Box for the Crenarchaeote Sulfolobus acidocaldarius

Wagner, Michaela; Wolferen, Marleen van; Wagner, Alexander; Lassak, Kerstin; Meyer, Benjamin; Reimann, Julia; Albers, Sonja‐Verena

doi:10.3389/fmicb.2012.00214

Cited by 178 publications

(263 citation statements)

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“…Using the marker-less mutant method in the uracil auxotrophic S. acidocaldarius mutant MW001 as reference strain (Wagner et al, 2012), in-frame deletion mutants were obtained for saci0102, saci0446 and saci1242. As this strategy was not successful for the construction of saci0133, saci1223 and saci1219, these ORFs were deleted by a single homologous recombination step disrupting each gene via the insertion of the pyrEF selection cassette.…”

Section: Resultsmentioning

confidence: 99%

“…acidocaldarius strains and growth conditions S. acidocaldarius MW001 (Wagner et al, 2012) and all in-frame marker-less deletion mutants were aerobically grown at 76 1C in Brock media (Brock et al, 1972), pH 3 and supplemented with 0.1% (w/v) N-Z-amine and 10 mg ml À 1 uracil. Uracil was not added to the media for cultivation of S. acidocaldarius MW001 pyrEF þ and pyrEF disruption mutants.…”

Section: Methodsmentioning

confidence: 99%

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Lrs14 transcriptional regulators influence biofilm formation and cell motility of Crenarchaea

et al. 2013

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Like bacteria, archaea predominately exist as biofilms in nature. However, the environmental cues and the molecular mechanisms driving archaeal biofilm development are not characterized. Here we provide data suggesting that the transcriptional regulators belonging to the Lrs14-like protein family constitute a key regulatory factor during Sulfolobus biofilm development. Among the six lrs14-like genes encoded by Sulfolobus acidocaldarius, the deletion of three led to markedly altered biofilm phenotypes. Although Dsaci1223 and Dsaci1242 deletion mutants were impaired in biofilm formation, the Dsaci0446 deletion strain exhibited a highly increased extracellular polymeric substance (EPS) production, leading to a robust biofilm structure. Moreover, although the expression of the adhesive pili (aap) genes was upregulated, the genes of the motility structure, the archaellum (fla), were downregulated rendering the Dsaci0446 strain non-motile. Gel shift assays confirmed that Saci0446 bound to the promoter regions of fla and aap thus controlling the expression of both cell surface structures. In addition, genetic epistasis analysis using Dsaci0446 as background strain identified a gene cluster involved in the EPS biosynthetic pathway of S. acidocaldarius. These results provide insights into both the molecular mechanisms that govern biofilm formation in Crenarchaea and the functionality of the Lrs14-like proteins, an archaea-specific class of transcriptional regulators.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Lrs14 transcriptional regulators influence biofilm formation and cell motility of Crenarchaea

et al. 2013

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“…As was the case with halophiles and methanogens, the development of key genetic tools allowing the creation of markerless inframe deletion mutants (140) propelled analysis of the N-glycosylation process in the crenarchaeote S. acidocaldarius. As in other archaeal model systems, here too, the S-layer glycoprotein has proven to be an excellent reporter for studying N-glycosylation.…”

Section: Pathway Of N-linked Glycosylation In the Thermoacidophile Sumentioning

confidence: 99%

N-Linked Glycosylation in Archaea: a Structural, Functional, and Genetic Analysis

Jarrell

Ding

Meyer

et al. 2014

Microbiol Mol Biol Rev

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SUMMARY N-glycosylation of proteins is one of the most prevalent posttranslational modifications in nature. Accordingly, a pathway with shared commonalities is found in all three domains of life. While excellent model systems have been developed for studying N-glycosylation in both Eukarya and Bacteria , an understanding of this process in Archaea was hampered until recently by a lack of effective molecular tools. However, within the last decade, impressive advances in the study of the archaeal version of this important pathway have been made for halophiles, methanogens, and thermoacidophiles, combining glycan structural information obtained by mass spectrometry with bioinformatic, genetic, biochemical, and enzymatic data. These studies reveal both features shared with the eukaryal and bacterial domains and novel archaeon-specific aspects. Unique features of N-glycosylation in Archaea include the presence of unusual dolichol lipid carriers, the use of a variety of linking sugars that connect the glycan to proteins, the presence of novel sugars as glycan constituents, the presence of two very different N-linked glycans attached to the same protein, and the ability to vary the N-glycan composition under different growth conditions. These advances are the focus of this review, with an emphasis on N-glycosylation pathways in Haloferax , Methanococcus , and Sulfolobus .

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“…Sulfolobus acidocaldarius is mainly found in terrestrial solfatara springs and is an aerobic thermoacidophilic crenarchaeon that grows on organic substrates in pH~1.8 and temperatures ranging between 70-80 C. S. acidocaldarius DSM639 is used as a model organism for the Crenarchaeota phylum. In addition to its available genome sequence (Chen et al, 2005), considerable progress has been made in establishing molecular biology tools such as knockout techniques and shuttle vectors that allow genetic studies (Berkner & Lipps, 2008;Wagner et al, 2012).…”

Section: Introductionmentioning

confidence: 99%

Transcriptomic analysis reveals how a lack of potassium ions increases Sulfolobus acidocaldarius sensitivity to pH changes

et al. 2016

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Extremely acidophilic microorganisms (optimum growth pH of 3) maintain a near neutral cytoplasmic pH via several homeostatic mechanisms, including an inside positive membrane potential created by potassium ions. Transcriptomic responses to pH stress in the thermoacidophilic archaeon, Sulfolobus acidocaldarius were investigated by growing cells without added sodium and/or potassium ions at both optimal and sub-optimal pH. Culturing the cells in the absence of added sodium or potassium ions resulted in a reduced growth rate compared to full-salt conditions as well as 43 and 75 significantly different RNA transcript ratios, respectively. Differentially expressed RNA transcripts during growth in the absence of added sodium ions included genes coding for permeases, a sodium/proline transporter and electron transport proteins. In contrast, culturing without added potassium ions resulted in higher RNA transcripts for similar genes as a lack of sodium ions plus genes related to spermidine that has a general role in response to stress and a decarboxylase that potentially consumes protons. The greatest RNA transcript response occurred when S. acidocaldarius cells were grown in the absence of potassium and/or sodium at a sub-optimal pH. These adaptations included those listed above plus osmoregulated glucans and mechanosensitive channels that have previously been shown to respond to osmotic stress. In addition, data analyses revealed two co-expressed IclR family transcriptional regulator genes with a previously unknown role in the S. acidocaldarius pH stress response. Our study provides additional evidence towards the importance of potassium in acidophile growth at acidic pH.

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Versatile Genetic Tool Box for the Crenarchaeote Sulfolobus acidocaldarius

Cited by 178 publications

References 46 publications

Lrs14 transcriptional regulators influence biofilm formation and cell motility of Crenarchaea

Lrs14 transcriptional regulators influence biofilm formation and cell motility of Crenarchaea

N-Linked Glycosylation in Archaea: a Structural, Functional, and Genetic Analysis

Transcriptomic analysis reveals how a lack of potassium ions increases Sulfolobus acidocaldarius sensitivity to pH changes

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