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The majority of Crenarchaeota utilize the cell division system (Cdv) to divide. This system consists of three highly conserved genes, cdvA, cdvB and cdvC that are organized in an operon. CdvC is homologous to the AAA-type ATPase Vps4, involved in multivesicular body biogenesis in eukaryotes. CdvA is a unique archaeal protein that interacts with the membrane, while CdvB is homologous to the eukaryal Vps24 and forms helical filaments. Most Crenarcheota contain additional CdvB paralogs. In Sulfolobus acidocaldarius these are termed CdvB1–3. We have used a gene inactivation approach to determine the impact of these additional cdvB genes on cell division. Independent deletion mutants of these genes were analyzed for growth and protein localization. One of the deletion strains (DcdvB3) showed a severe growth defect on plates and delayed growth on liquid medium. It showed the formation of enlarged cells and a defect in DNA segregation. Since these defects are accompanied with an aberrant localization of CdvA and CdvB, we conclude that CdvB3 fulfills an important accessory role in cell division.

Within the archaea, the thermoacidophilic crenarchaeote Sulfolobus solfataricus has become an important model organism for physiology and biochemistry, comparative and functional genomics, as well as, more recently also for systems biology approaches. Within the Sulfolobus Systems Biology (“SulfoSYS”)-project the effect of changing growth temperatures on a metabolic network is investigated at the systems level by integrating genomic, transcriptomic, proteomic, metabolomic and enzymatic information for production of a silicon cell-model. The network under investigation is the central carbohydrate metabolism. The generation of high-quality quantitative data, which is critical for the investigation of biological systems and the successful integration of the different datasets, derived for example from high-throughput approaches (e.g., transcriptome or proteome analyses), requires the application and compliance of uniform standard protocols, e.g., for growth and handling of the organism as well as the “–omics” approaches. Here, we report on the establishment and implementation of standard operating procedures for the different wet-lab and in silico techniques that are applied within the SulfoSYS-project and that we believe can be useful for future projects on Sulfolobus or (hyper)thermophiles in general. Beside established techniques, it includes new methodologies like strain surveillance, the improved identification of membrane proteins and the application of crenarchaeal metabolomics. Electronic supplementary material The online version of this article (doi:10.1007/s00792-009-0280-0) contains supplementary material, which is available to authorized users.

The crenarchaea Sulfolobus acidocaldarius, S. solfataricus and S. tokodaii, release membrane vesicles into the medium. These membrane vesicles consist of tetraether lipids and are coated with an S-layer. A proteomic analysis reveals the presence of proteins homologous to subunits of the eukaryotic endosomal sorting complex required for transport (ESCRT). Immunodetection of one of these homologs suggest a cell surface localization in intact cells. These data suggest that the membrane vesicles in Sulfolobus sp. emerge from a specific budding process with similarity to the endosomal sorting pathway.

Sugar uptake in Sulfolobus solfataricus, a thermoacidophilic archaeon, occurs through high-affinity binding of protein-dependent ABC transporters. We have investigated the expression patterns of two sugar transport operons, that is, the glucose and arabinose transporters. Analysis of the araS promoter activity, and the mRNA and protein levels in S. solfataricus cells grown on different carbon sources showed that expression of the arabinose transporter gene cluster is highly regulated and dependent on the presence of arabinose in the medium. Glucose in the growth medium repressed the expression of the arabinose transport genes. By means of primer extension, the transcriptional start site for the arabinose operon was mapped. Interestingly, expression of the arabinose transporter is down-regulated by addition of a selective set of amino acids to the medium. Expression of the glucose transporter genes appeared constitutive. These data confirm the earlier observation of a catabolite repression-like system in S. solfataricus.

In extreme environments, mainly Archaea are encountered. The archaeal cytoplasmic membrane contains unique ether lipids that cannot easily be degraded, are temperature- and mechanically resistant, and highly salt tolerant. Moreover, thermophilic and extreme acidophilic Archaea possess membrane-spanning tetraether lipids that form a rigid monolayer membrane which is nearly impermeable to ions and protons. These properties make the archaeal lipid membranes more suitable for life and survival in extreme environments than the ester-type bilayer lipids of Bacteria or Eukarya.

Sugar binding proteins of the thermoacidophile Sulfolobus solfataricus function together with ABC transporters in the uptake of sugars. They are synthesized as precursors with a class III signal peptide that are normally found in archaeal flagellins and bacterial type IV pilins. The functional expression of sugar binding proteins at the cell surface is dependent on the bindosome assembly system (Bas) that is homologous to bacterial type IV pilin assembly systems. The Bas system consists of an assembly ATPase, BasE; a membrane anchoring protein, BasF; and three small class III signal peptide containing proteins BasABC. Expression of BasEF in a S. solfataricus Delta basEF strain restored the uptake of glucose, while an ATPase mutant of BasE was unable to complement. BasEF was detergent-extracted from S. solfataricus membranes as a stable protein complex. Solute binding proteins can be extracted from the cell surface as two high molecular mass complexes of 600 and 400 kDa, wherein the largest complex also contains the main S-layer protein SlaA. Electron microscopic analysis of the cell surface of the wild-type and Delta basEF strain indicates that the absence of the BasEF complex causes an alteration in cell morphology and the corrugation of the S-layer pattern that is reversed by complementation with the BasEF complex. These results suggest an interaction between the S-layer and the sugar binding proteins that contribute to cell shape.