Structural analysis of the Sulfolobus solfataricus TF55ß chaperonin by cryo-electron microscopy.

Chaperonins are biomolecular complexes that assist in protein folding. Thermophilic factor 55 (TF55) is a group II chaperonin found in the archaeal genus Sulfolobus that has α, ß and γ subunits. Using cryo-electron microscopy, structures of the ß-only complex of S. solfataricus TF55 (TF55ß) were determined to 3.6-4.2 Šresolution. The structures of the TF55ß complexes formed in the presence of ADP or ATP highlighted an open state in which nucleotide exchange can occur before progressing in the refolding cycle.

Electron microscopy studies of Type III CRISPR machines in Sulfolobus solfataricus.

The CRISPR (clustered regularly interspaced short palindromic repeats) system is an adaptive immune system that targets viruses and other mobile genetic elements in bacteria and archaea. Cells store information of past infections in their genome in repeat-spacer arrays. After transcription, these arrays are processed into unit-length crRNA (CRISPR RNA) that is loaded into effector complexes encoded by Cas (CRISPR-associated) genes. CRISPR-Cas complexes target invading nucleic acid for degradation. CRISPR effector complexes have been classified into three main types (I-III). Type III effector complexes share the Cas10 subunit. In the present paper, we discuss the structures of the two Type III effector complexes from Sulfolobus solfataricus, SsoCSM (subtype III-A) and SsoCMR (subtype III-B), obtained by electron microscopy and single particle analysis. We also compare these structures with Cascade (CRISPR-associated complex for antiviral defence) and with the RecA nucleoprotein.

Sulfolobus turreted icosahedral virus c92 protein responsible for the formation of pyramid-like cellular lysis structures.

Host cells infected by Sulfolobus turreted icosahedral virus (STIV) have been shown to produce unusual pyramid-like structures on the cell surface. These structures represent a virus-induced lysis mechanism that is present in Archaea and appears to be distinct from the holin/endolysin system described for DNA bacteriophages. This study investigated the STIV gene products required for pyramid formation in its host Sulfolobus solfataricus. Overexpression of STIV open reading frame (ORF) c92 in S. solfataricus alone is sufficient to produce the pyramid-like lysis structures in cells. Gene disruption of c92 within STIV demonstrates that c92 is an essential protein for virus replication. Immunolocalization of c92 shows that the protein is localized to the cellular membranes forming the pyramid-like structures.

Appendage-mediated surface adherence of Sulfolobus solfataricus.

Attachment of microorganisms to surfaces is a prerequisite for colonization and biofilm formation. The hyperthermophilic crenarchaeote Sulfolobus solfataricus was able to attach to a variety of surfaces, such as glass, mica, pyrite, and carbon-coated gold grids. Deletion mutant analysis showed that for initial attachment the presence of flagella and pili is essential. Attached cells produced extracellular polysaccharides containing mannose, galactose, and N-acetylglucosamine. Genes possibly involved in the production of the extracellular polysaccharides were identified.

Particle assembly and ultrastructural features associated with replication of the lytic archaeal virus sulfolobus turreted icosahedral virus.

Little is known about the replication cycle of archaeal viruses. We have investigated the ultrastructural changes of Sulfolobus solfataricus P2 associated with infection by Sulfolobus turreted icosahedral virus (STIV). A time course of a near synchronous STIV infection was analyzed using both scanning and transmission electron microscopy. Assembly of STIV particles, including particles lacking DNA, was observed within cells, and fully assembled STIV particles were visible by 30 h postinfection (hpi). STIV was determined to be a lytic virus, causing cell disruption beginning at 30 hpi. Prior to cell lysis, virus infection resulted in the formation of pyramid-like projections from the cell surface. These projections, which have not been documented in any other host-virus system, appeared to be caused by the protrusion of the cell membrane beyond the bordering S-layer. These structures are thought to be sites at which progeny virus particles are released from infected cells. Based on these observations of lysis, a plaque assay was developed for STIV. From these studies we propose an overall assembly model for STIV.

Transcriptome analysis of infection of the archaeon Sulfolobus solfataricus with Sulfolobus turreted icosahedral virus.

Microarray analysis of infection by Sulfolobus turreted icosahedral virus (STIV) revealed insights into the timing and extent of virus transcription, as well as differential regulation of host genes. Using a microarray containing genes from both the host and the virus, the infection cycle of STIV was studied. Following infection of Sulfolobus solfataricus strain 2-2-12 with STIV, transcription of virus genes was first detected at 8 h postinfection (p.i.), with a peak at 24 h p.i. Lysis of cells was first detected at 32 h p.i. There was little temporal control of the transcription of virus genes, although the three open reading frames on the noncoding strand were transcribed later in the infection process. During the infection, 177 host genes were determined to be differentially expressed, with 124 genes up-regulated and 53 genes down-regulated. The up-regulated genes were dominated by genes associated with DNA replication and repair and those of unknown function, while the down-regulated genes, mostly detected at 32 h p.i., were associated with energy production and metabolism. Examination of infected cells by transmission electron microscopy revealed alterations in cell ultrastructure consistent with the microarray analysis. The observed patterns of transcription suggest that up-regulated genes are likely used by the virus to reprogram the cell for virus replication, while the down-regulated genes reflect the imminent lysis of the cells.

Flagellar motility and structure in the hyperthermoacidophilic archaeon Sulfolobus solfataricus.

Flagellation in archaea is widespread and is involved in swimming motility. Here, we demonstrate that the structural flagellin gene from the crenarchaeaon Sulfolobus solfataricus is highly expressed in stationary-phase-grown cells and under unfavorable nutritional conditions. A mutant in a flagellar auxiliary gene, flaJ, was found to be nonmotile. Electron microscopic imaging of the flagellum indicates that the filaments are composed of right-handed helices.