RESUMO
The family Turriviridae includes viruses with a dsDNA genome of 16-17 kbp. Virions are spherical with a diameter of approximately 75 nm and comprise a host-derived internal lipid membrane surrounded by a proteinaceous capsid shell. Members of the family Turriviridae infect extremophilic archaea of the genera Sulfolobus and Saccharolobus. Viral infection results in cell lysis for Sulfolobus turreted icosahedral virus 1 infection but other members of the family can be temperate. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Turriviridae, which is available at ictv.global/report/turriviridae.
Assuntos
Vírus de DNA , Genoma Viral , Vírion , Vírus de DNA/classificação , Vírus de DNA/genética , Vírus de DNA/ultraestrutura , Vírion/ultraestrutura , Vírus de Archaea/classificação , Vírus de Archaea/genética , Vírus de Archaea/ultraestrutura , Vírus de Archaea/fisiologia , Sulfolobus/virologia , Sulfolobus/genética , DNA Viral/genéticaRESUMO
Pseudouridine, one of the most abundant RNA modifications, is synthesized by stand-alone or RNA-guided pseudouridine synthases. Here, we comprehensively mapped pseudouridines in rRNAs, tRNAs and small RNAs in the archaeon Sulfolobus islandicus and identified Cbf5-associated H/ACA RNAs. Through genetic deletion and in vitro modification assays, we determined the responsible enzymes for these modifications. The pseudouridylation machinery in S. islandicus consists of the stand-alone enzymes aPus7 and aPus10, and six H/ACA RNA-guided enzymes that account for all identified pseudouridines. These H/ACA RNAs guide the modification of all eleven sites in rRNAs, two sites in tRNAs, and two sites in CRISPR RNAs. One H/ACA RNA shows exceptional versatility by targeting eight different sites. aPus7 and aPus10 are responsible for modifying positions 13, 54 and 55 in tRNAs. We identified four atypical H/ACA RNAs that lack the lower stem and the ACA motif and confirmed their function both in vivo and in vitro. Intriguingly, atypical H/ACA RNAs can be modified by Cbf5 in a guide-independent manner. Our data provide the first global view of pseudouridylation in archaea and reveal unexpected structures, substrates, and activities of archaeal H/ACA RNPs.
Assuntos
Pseudouridina , RNA Arqueal , RNA de Transferência , Sulfolobus , Pseudouridina/metabolismo , Sulfolobus/genética , Sulfolobus/metabolismo , RNA de Transferência/metabolismo , RNA de Transferência/genética , RNA Arqueal/genética , RNA Arqueal/metabolismo , RNA Arqueal/química , RNA Ribossômico/metabolismo , RNA Ribossômico/genética , Proteínas Arqueais/metabolismo , Proteínas Arqueais/genética , Processamento Pós-Transcricional do RNA , RNA Guia de Sistemas CRISPR-Cas/genética , RNA Guia de Sistemas CRISPR-Cas/metabolismo , Transferases Intramoleculares/genética , Transferases Intramoleculares/metabolismoRESUMO
Proteins in the structural maintenance of chromosomes (SMC) superfamily play key roles in chromosome organization and are ubiquitous across all domains of life. However, SMC proteins are notably absent in the Desulfurococcales of phylum Crenarchaeota. Intrigued by this observation, we performed chromosome conformation capture experiments in the model Desulfurococcales species Aeropyrum pernix. As in other archaea, we observe chromosomal interaction domains across the chromosome. The boundaries between chromosomal interaction domains show a dependence on transcription and translation for their definition. Importantly, however, we reveal an additional higher-order, bipartite organization of the chromosome-with a small high-gene-expression and self-interacting domain that is defined by transcriptional activity and loop structures. Viewing these data in the context of the distribution of SMC superfamily proteins in the Crenarchaeota, we suggest that the organization of the Aeropyrum genome represents an evolutionary antecedent of the compartmentalized architecture observed in the Sulfolobus lineage.
Assuntos
Crenarchaeota , Sulfolobus , Archaea/genética , Crenarchaeota/genética , Expressão Gênica , Sulfolobus/genética , CromossomosRESUMO
Archaeal lemon-shaped viruses have unique helical capsids composed of highly hydrophobic protein strands which can slide past each other resulting in remarkable morphological reorganization. Here, using atomic force microscopy, we explore the biomechanical properties of the lemon-shaped virions of Sulfolobus monocaudavirus 1 (SMV1), a double-stranded DNA virus which infects hyperthermophilic (~80 °C) and acidophilic (pH ~ 2) archaea. Our results reveal that SMV1 virions are extremely soft and withstand repeated extensive deformations, reaching remarkable strains of 80% during multiple cycles of consecutive mechanical assaults, yet showing scarce traces of disruption. SMV1 virions can reversibly collapse wall-to-wall, reducing their volume by ~90%. Beyond revealing the exceptional malleability of the SMV1 protein shell, our data also suggest a fluid-like nucleoprotein cargo which can flow inside the capsid, resisting and accommodating mechanical deformations without further alteration. Our experiments suggest a packing fraction of the virus core to be as low as 11%, with the amount of the accessory proteins almost four times exceeding that of the viral genome. Our findings indicate that SMV1 protein capsid displays biomechanical properties of lipid membranes, which is not found among protein capsids of other viruses. The remarkable malleability and fluidity of the SMV1 virions are likely necessary for the structural transformations during the infection and adaptation to extreme environmental conditions.
Assuntos
Vírus de Archaea , Sulfolobus , Vírus de Archaea/genética , Vírus de Archaea/química , Capsídeo/metabolismo , Nucleoproteínas/genética , Proteínas do Capsídeo/genética , Genoma Viral , TomografiaRESUMO
Aldehyde dehydrogenase (ALDH) is widely distributed in nature and its characteristics have been examined. ALDH plays an important role in aldehyde detoxification. Sources of aldehydes include incomplete combustion and emissions from paints, linoleum and varnishes in the living environment. Acetaldehyde is also considered to be carcinogenic and toxic. Thermostable ALDH from the hyperthermophilic archaeon Sulfolobus tokodaii exhibits high activity towards acetaldehyde and has potential applications as a biosensor for acetaldehyde. Thermostable ALDH displays a unique and wide adaptability. Therefore, its crystal structure can provide new insights into the catalytic mechanism and potential applications of ALDHs. However, a crystal structure of a thermostable ALDH exhibiting high activity towards acetaldehyde has not been reported to date. In this study, crystals of recombinant thermostable ALDH from S. tokodaii were prepared and the crystal structure of its holo form was determined. A crystal of the enzyme was prepared and its structure in complex with NADP was determined at a resolution of 2.2â Å. This structural analysis may facilitate further studies on catalytic mechanisms and applications.
Assuntos
Sulfolobus , Archaea , Cristalografia por Raios X , AcetaldeídoRESUMO
Endonuclease V (EndoV), which is widespread in bacteria, eukarya and Archaea, can cleave hypoxanthine (Hx)-containing DNA or RNA strand, and play an essential role in Hx repair. However, our understanding on archaeal EndoV's function remains incomplete. The model archaeon Sulfolobus islandicus REY15A encodes a putative EndoV protein (Sis-EndoV). Herein, we probed the biochemical characteristics of Sis-EndoV and dissected the roles of its seven conserved residues. Our biochemical data demonstrate that Sis-EndoV displays maximum cleavage efficiency at above 60 °C and at pH 7.0-9.0, and the enzyme activity is dependent on a divalent metal ion, among which Mg2+ is optimal. Importantly, we first measured the activation energy for cleaving Hx-containing ssDNA by Sis-EndoV to be 9.6 ± 0.8 kcal/mol by kinetic analyses, suggesting that chemical catalysis might be a rate-limiting step for catalysis. Mutational analyses show that residue D38 in Sis-EndoV is essential for catalysis, but has no role in DNA binding. Furthermore, we first revealed that residues Y41 and D189 in Sis-EndoV are involved in both DNA cleavage and DNA binding, but residues F77, H103, K156 and F161 are only responsible for DNA binding.
Assuntos
Desoxirribonuclease (Dímero de Pirimidina) , Sulfolobus , Desoxirribonuclease (Dímero de Pirimidina)/química , Desoxirribonuclease (Dímero de Pirimidina)/genética , Desoxirribonuclease (Dímero de Pirimidina)/metabolismo , Sulfolobus/genética , Sulfolobus/metabolismo , Reparo do DNA , Dano ao DNA , DNARESUMO
Analysis of the profile of the tRNA modifications in several Archaea allowed us to observe a novel modified uridine in the V-loop of several tRNAs from two species: Pyrococcus furiosus and Sulfolobus acidocaldarius Recently, Ohira and colleagues characterized 2'-phosphouridine (Up) at position 47 in tRNAs of thermophilic Sulfurisphaera tokodaii, as well as in several other archaea and thermophilic bacteria. From the presence of the gene arkI corresponding to the RNA kinase responsible for Up47 formation, they also concluded that Up47 should be present in tRNAs of other thermophilic Archaea Reanalysis of our earlier data confirms that the unidentified residue in tRNAs of both P. furiosus and S. acidocaldarius is indeed 2'-phosphouridine followed by m5C48. Moreover, we find this modification in several tRNAs of other Archaea and of the hyperthermophilic bacterium Aquifex aeolicus.
Assuntos
Archaea , Sulfolobus , Archaea/genética , Bactérias/genética , Sulfolobus/genéticaRESUMO
Uracil DNA glycosylase (UDG) can remove uracil from DNA, thus playing an essential role in maintaining genomic stability. Family IV UDG members are mostly widespread in hyperthermophilic Archaea and bacteria. In this work, we characterized the family IV UDG from the hyperthermophilic crenarchaeon Sulfolobus islandicus REY15A (Sis-UDGIV) biochemically, and dissected the roles of nine conserved residues in uracil excision by mutational analyses. Biochemical data demonstrate that Sis-UDGIV displays maximum efficiency for uracil excision at 50 °C ~ 70 °C and at pH 7.0-9.0. Additionally, the enzyme has displays a weak activity without a divalent metal ion, but maximum activity with Mg2+. Our mutational analyses show that residues E48 and F55 in Sis-UDGIV are essential for uracil removal, and residues E48, F55, R87, R92 and K146 are responsible for binding DNA. Importantly, we systemically revealed the roles of four conserved cysteine residues C14, C17, C86 and C102 in Sis-UDGIV that are required for being ligands of FeS cluster in maintaining the overall protein conformation and stability by circular dichroism analyses. Overall, our work has provided insights into biochemical function and DNA-binding specificity of archaeal family IV UDGs.
Assuntos
Sulfolobus , Uracila-DNA Glicosidase , Uracila-DNA Glicosidase/química , Sulfolobus/genética , Sequência de Aminoácidos , DNA/metabolismo , Reparo do DNA , Archaea/metabolismo , Uracila/metabolismoRESUMO
Cell cycle regulation is of paramount importance for all forms of life. Here, we report that a conserved and essential cell cycle-specific transcription factor (designated as aCcr1) and its viral homologs control cell division in Sulfolobales. We show that the transcription level of accr1 reaches peak during active cell division (D-phase) subsequent to the expression of CdvA, an archaea-specific cell division protein. Cells over-expressing the 58-aa-long RHH (ribbon-helix-helix) family cellular transcription factor as well as the homologs encoded by large spindle-shaped viruses Acidianus two-tailed virus (ATV) and Sulfolobus monocaudavirus 3 (SMV3) display significant growth retardation and cell division failure, manifesting as enlarged cells with multiple chromosomes. aCcr1 over-expression results in downregulation of 17 genes (>4-fold), including cdvA. A conserved motif, aCcr1-box, located between the TATA-binding box and the translation initiation site of 13 out of the 17 highly repressed genes, is critical for aCcr1 binding. The aCcr1-box is present in the promoters and 5' UTRs of cdvA genes across Sulfolobales, suggesting that aCcr1-mediated cdvA repression is an evolutionarily conserved mechanism by which archaeal cells dictate cytokinesis progression, whereas their viruses take advantage of this mechanism to manipulate the host cell cycle.
Assuntos
Sulfolobus , Fatores de Transcrição , Fatores de Transcrição/genética , Archaea , Divisão Celular , Sulfolobus/genética , Regulação da Expressão GênicaRESUMO
Anti-Clustered regularly interspaced small palindromic repeat (CRISPR) (Acr) phages cooperate to establish a successful infection in CRISPR-containing host. We report here the selective advantage provided by a replication initiator, Rep, toward cooperative host immunosuppression by viruses encoding Acrs. A rep knockout mutant (Δgp16) of Sulfolobus islandicus rod-shaped virus 2 produced around fourfold less virus in a CRISPR-null host, suggesting that Rep is the major replication initiator. In addition to Rep-dependent replication initiation from the viral genomic termini, we detected Rep-independent replication initiation from nonterminal sites. Intriguingly, despite the presence of Acrs, lack of Rep showed a profound effect on virus propagation in a host carrying CRISPR-Cas immunity. Accordingly, the co-infecting parental virus (rep-containing) outcompeted the Δgp16 mutant much more quickly in the CRISPR-containing host than in CRISPR-null host. Despite the nonessentiality, rep is carried by all known members of Rudiviridae, which is likely an evolutionary outcome driven by the ubiquitous presence of CRISPR-Cas in Sulfolobales.
Assuntos
Bacteriófagos , Sulfolobus , Vírus , Sistemas CRISPR-Cas/genética , Edição de Genes , Sulfolobus/genética , Vírus/genética , Bacteriófagos/genéticaRESUMO
All living organisms have evolved DNA damage response (DDR) strategies in coping with threats to the integrity of their genome. In response to DNA damage, Sulfolobus islandicus activates its DDR network in which Orc1-2, an ortholog of the archaeal Orc1/Cdc6 superfamily proteins, plays a central regulatory role. Here, we show that pretreatment with UV irradiation reduced virus genome replication in S. islandicus infected with the fusellovirus SSV2. Like treatment with UV or the DNA-damaging agent 4-nitroquinoline-1-oxide (NQO), infection with SSV2 facilitated the expression of orc1-2 and significantly raised the cellular level of Orc1-2. The inhibitory effect of UV irradiation on the virus DNA level was no longer apparent in the infected culture of an S. islandicus orc1-2 deletion mutant strain. On the other hand, the overexpression of orc1-2 decreased virus genomic DNA by ~102-fold compared to that in the parent strain. Furthermore, as part of the Orc1-2-mediated DDR response genes for homologous recombination repair (HRR), cell aggregation and intercellular DNA transfer were upregulated, whereas genes for cell division were downregulated. However, the HRR pathway remained functional in host inhibition of SSV2 genome replication in the absence of UpsA, a subunit of pili essential for intercellular DNA transfer. In agreement with this finding, lack of the general transcriptional activator TFB3, which controls the expression of the ups genes, only moderately affected SSV2 genome replication. Our results demonstrate that infection of S. islandicus by SSV2 triggers the host DDR pathway that, in return, suppresses virus genome replication. IMPORTANCE Extremophiles thrive in harsh habitats and thus often face a daunting challenge to the integrity of their genome. How these organisms respond to virus infection when their genome is damaged remains unclear. We found that the thermophilic archaeon Sulfolobus islandicus became more inhibitory to genome replication of the virus SSV2 after preinfection UV irradiation than without the pretreatment. On the other hand, like treatment with UV or other DNA-damaging agents, infection of S. islandicus by SSV2 triggers the activation of Orc1-2-mediated DNA damage response, including the activation of homologous recombination repair, cell aggregation and DNA import, and the repression of cell division. The inhibitory effect of pretreatment with UV irradiation on SSV2 genome replication was no longer observed in an S. islandicus mutant lacking Orc1-2. Our results suggest that DNA damage response is employed by S. islandicus as a strategy to defend against virus infection.
Assuntos
Fuselloviridae , Sulfolobus , Dano ao DNA/genética , Reparo do DNA/genética , Fuselloviridae/genética , Sulfolobus/genética , Sulfolobus/efeitos da radiação , Sulfolobus/virologia , Replicação Viral , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos da radiação , Raios Ultravioleta , 4-Nitroquinolina-1-Óxido/farmacologia , Complexo de Reconhecimento de Origem/genética , Complexo de Reconhecimento de Origem/metabolismoRESUMO
Uracil DNA glycosylase (UDG) can excise uracil from DNA, thus playing an essential role in counteracting mutations. The genome of the hyperthermophilic crenarchaeon Sulfolobus islandicus REY15A encodes one putative Family V UDG (Sis-UDGV). Herein, we provide evidence that Sis-UDGV is a bi-functional glycosylase that can not only excise uracil from DNA, but cleave the generated apurinic/apyrimidinic (AP) site, which differs from other reported mono-functional Family V UDG homologs. Intriguingly, the enzyme can cleave DNA containing an AP site, thus suggesting that it might be involved in AP site repair. Biochemical data demonstrate that Sis-UDGV displays maximum activity for uracil removal at 45 °C â¼ 65 oC and at pH 8.0 â¼ 9.0. Furthermore, Sis-UDGV displays a substrate preference for uracil-containing ssDNA over uracil-containing dsDNA, but has no activity and weak activity for excising hypoxanthine from ssDNA and dsDNA, respectively. Importantly, we dissected the roles of seven conserved residues in Sis-UDGV by mutational analyses, demonstrating that residues D91, E117, E128, H167 and R192 are essential for catalysis. To our knowledge, it is the first report on the novel Family V UDG from Archaea with bi-functionality that harbors glycosylase/AP lyase activity.
Assuntos
Sulfolobus , Uracila-DNA Glicosidase , Uracila-DNA Glicosidase/metabolismo , Sulfolobus/genética , Sulfolobus/metabolismo , Uracila , Reparo do DNA , DNARESUMO
CRISPR-Cas systems empower prokaryotes with adaptive immunity against invasive mobile genetic elements. At the first step of CRISPR immunity adaptation, short DNA fragments from the invaders are integrated into CRISPR arrays at the leader-proximal end. To date, the mechanism of recognition of the leader-proximal end remains largely unknown. Here, in the Sulfolobus islandicus subtype I-A system, we show that mutations destroying the proximal region reduce CRISPR adaptation in vivo. We identify that a stem-loop structure is present on the leader-proximal end, and we demonstrate that Cas1 preferentially binds the stem-loop structure in vitro. Moreover, we demonstrate that the integrase activity of Cas1 is modulated by interacting with a CRISPR-associated factor Csa3a. When translocated to the CRISPR array, the Csa3a-Cas1 complex is separated by Csa3a binding to the leader-distal motif and Cas1 binding to the leader-proximal end. Mutation at the leader-distal motif reduces CRISPR adaptation efficiency, further confirming the in vivo function of leader-distal motif. Together, our results suggest a general model for binding of Cas1 protein to a leader motif and modulation of integrase activity by an accessory factor.
Assuntos
Proteínas Associadas a CRISPR , Sulfolobus , Proteínas Associadas a CRISPR/metabolismo , Sistemas CRISPR-Cas , Integrases/metabolismo , Motivos de Nucleotídeos , Sulfolobus/genética , Sulfolobus/metabolismoRESUMO
Homologous recombination-based gene targeting is a powerful and classic reverse genetics approach to precisely elucidate in vivo gene functions in the organisms across all three domains of life. Gene function studies in Archaea, particularly for those flourishing in inhospitable natural environments that are anaerobic, usually hot, and acidic, have been a great challenge; however, this situation was recently overturned with the increasing availability of genetic manipulation systems in several cultivable archaeal species. In the present chapter, we describe a detailed procedure to rapidly generate gene disruption mutants in the hyperthermophilic crenarchaeon Sulfolobus islandicus via a recently developed Microhomology-Mediated Gene Inactivation (MMGI) approach. We highlight crucial experimental details required to be carefully considered when using the MMGI approach for genetic manipulations. We hope this highly reproducible procedure can supplement existing genetic tools for studying the biology of archaeal order Sulfolobales.
Assuntos
Sulfolobus , Archaea/genética , Inativação Gênica , Marcação de Genes , Técnicas Genéticas , Sulfolobus/genéticaRESUMO
Transposon insertion mutagenesis is a forward genetic approach that has been widely utilized for genetic characterization of bacteria and single-celled eukaryotes, and its applications are being rapidly expanded into a few archaeal model organisms for gene function analysis. Previously, we developed a Tn5-based in vivo transposon insertion mutagenesis system in the hyperthermophilic crenarchaeon S. islandicucs M.16.4 and defined the essential gene set under laboratory growth conditions. In this chapter, we will mainly focus on presenting details regarding the generation of a near-saturating transposon insertion mutant library in this crenarchaeal model. We envision that the traditional transposon-based forward mutagenesis screening paired with next generation sequencing will greatly speed up the exploration of archaeal genomic features.
Assuntos
Sulfolobus , Elementos de DNA Transponíveis/genética , Biblioteca Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Mutagênese Insercional , Sulfolobus/genéticaRESUMO
Type III CRISPR-Cas systems show the target (tg)RNA-activated indiscriminate DNA cleavage and synthesis of oligoadenylates (cOA) and a secondary signal that activates downstream nuclease effectors to exert indiscriminate RNA/DNA cleavage, and both activities are regulated in a spatiotemporal fashion. In III-B Cmr systems, cognate tgRNAs activate the two Cmr2-based activities, which are then inactivated via tgRNA cleavage by Cmr4, but how Cmr4 nuclease regulates the Cmr immunization remains to be experimentally characterized. Here, we conducted mutagenesis of Cmr4 conserved amino acids in Saccharolobus islandicus, and this revealed that Cmr4α RNase-dead (dCmr4α) mutation yields cell dormancy/death. We also found that plasmid-borne expression of dCmr4α in the wild-type strain strongly reduced plasmid transformation efficiency, and deletion of CRISPR arrays in the host genome reversed the dCmr4α inhibition. Expression of dCmr4α also strongly inhibited plasmid transformation with Cmr2αHD and Cmr2αPalm mutants, but the inhibition was diminished in Cmr2αHD,Palm. Since dCmr4α-containing effectors lack spatiotemporal regulation, this allows an everlasting interaction between crRNA and cellular RNAs to occur. As a result, some cellular RNAs, which are not effective in mediating immunity due to the presence of spatiotemporal regulation, trigger autoimmunity of the Cmr-α system in the S. islandicus cells expressing dCmr4α. Together, these results pinpoint the crucial importance of tgRNA cleavage in autoimmunity avoidance and in the regulation of immunization of type III systems.
Assuntos
Proteínas Associadas a CRISPR , Sulfolobus , Autoimunidade/genética , Proteínas Associadas a CRISPR/genética , Sistemas CRISPR-Cas/genética , RNA/genética , Clivagem do RNA , Sulfolobus/genéticaRESUMO
A small subset of acidic hot springs sampled in Yellowstone National Park yielded rod-shaped viruses which lysed liquid host cultures and formed clear plaques on lawns of host cells. Three isolates chosen for detailed analysis were found to be genetically related to previously described isolates of the Sulfolobus islandicus rod-shaped virus (SIRV), but distinct from them and from each other. Functional stability of the new isolates was assessed in a series of inactivation experiments. UV-C radiation inactivated one of the isolates somewhat faster than bacteriophage λ, suggesting that encapsidation in the SIRV-like virion did not confer unusual protection of the DNA from UV damage. With respect to high temperature, the new isolates were extremely, but not equally, stable. Several chemical treatments were found to inactivate the virions and, in some cases, to reveal apparent differences in virion stability among the isolates. Screening a larger set of isolates identified greater variation of these stability properties but found few correlations among the resulting profiles. The majority of host cells infected by the new isolates were killed, but survivors exhibited heritable resistance, which could not be attributed to CRISPR spacer acquisition or the loss of the pilus-related genes identified by earlier studies. Virus-resistant host variants arose at high frequency and most were resistant to multiple viral strains; conversely, resistant host clones generated virus-sensitive variants, also at high frequency. Virus-resistant cells lacked the ability of virus-sensitive cells to bind virions in liquid suspensions. Rapid interconversion of sensitive and resistant forms of a host strain suggests the operation of a yet-unidentified mechanism that acts to allow both the lytic virus and its host to propagate in highly localized natural populations, whereas variation of virion-stability phenotypes among the new viral isolates suggests that multiple molecular features contribute to the biological durability of these viruses.
Assuntos
Vírus de Archaea , Fontes Termais , Sulfolobus , Vírus , Vírus de Archaea/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , América do Norte , Vírus/genéticaRESUMO
Archaeal viruses with a spindle-shaped virion are abundant and widespread in extremely diverse environments. However, efforts to obtain the high-resolution structure of a spindle-shaped virus have been unsuccessful. Here, we present the structure of SSV19, a spindle-shaped virus infecting the hyperthermophilic archaeon Sulfolobus sp. E11-6. Our near-atomic structure reveals an unusual sevenfold symmetrical virus tail consisting of the tailspike, nozzle, and adaptor proteins. The spindle-shaped capsid shell is formed by seven left-handed helical strands, constructed of the hydrophobic major capsid protein, emanating from the highly glycosylated tail assembly. Sliding between adjacent strands is responsible for the variation of a virion in size. Ultrathin sections of the SSV19-infected cells show that SSV19 virions adsorb to the host cell membrane through the tail after penetrating the S-layer. The tailspike harbors a putative endo-mannanase domain, which shares structural similarity to a Bacteroides thetaiotaomicro endo-mannanase. Molecules of glycerol dibiphytanyl glycerol tetraether lipid were observed in hydrophobic clefts between the tail and the capsid shell. The nozzle protein resembles the stem and clip domains of the portals of herpesviruses and bacteriophages, implying an evolutionary relationship among the archaeal, bacterial, and eukaryotic viruses.
Assuntos
Fuselloviridae , Sulfolobus , Proteínas do Capsídeo/química , Fuselloviridae/química , Fuselloviridae/genética , Fuselloviridae/isolamento & purificação , Genoma Viral , Glicerol , Sulfolobus/virologia , Vírion/química , Vírion/genética , Vírion/isolamento & purificaçãoRESUMO
Sulfolobus islandicus is thermophilic archaea that live in an extreme environment of 75 °C-80 °C and pH 2-3. Currently, the molecular mechanism of archaeal adaptation to high temperatures and the stability of proteins at high temperatures are still unclear. This study utilizes proteomics to analyze the differential expression of S. islandicus proteins at different temperatures. We found that ribosomes, glycolysis, nucleotide metabolism, RNA metabolism, transport system, and sulfur metabolism are all affected by temperature. Methylation modification of some proteins changed with temperature. Thermal proteome profiling (TPP) was used to analyze the thermal stability of proteins under 65 °C-85 °C growth conditions. It is suggested that the Tm values of proteins are mainly distributed around the optimum growth temperature (OGT). The proteins in the glycolysis pathway had high thermal stability. Meanwhile, proteins related to DNA replication and translation showed low thermal stability. The protein thermal stability of S. islandicus cultured under 65 °C and 85 °C was higher than that of 75 °C. Our study reveals that S. islandicus may adapt to temperature changes by regulating protein synthesis and carbon metabolism pathways, changing post-translational modifications, and improving protein stability at the same time. SIGNIFICANCE: The molecular mechanism of archaeal adaptation to high temperatures and the stability of proteins at high temperatures are still unclear. Our proteomics study identified 477 differentially expressed proteins of S. islandicus at different temperatures, suggesting that ribosomes, glycolysis, nucleotide metabolism, RNA metabolism, transport system, and sulfur metabolism are affected by temperature. Meanwhile, we found that methylation modification of some proteins changed with temperature. To evaluate the thermal stability of the proteome, we performed thermal proteome profiling to analyze the Tm of proteins under 65 °C-85 °C growth conditions. Tm values of proteins are mainly distributed around the optimum growth temperature. The proteins in the glycolysis pathway had high thermal stability. Meanwhile, proteins related to DNA replication and translation showed low thermal stability. Our study reveals that S. islandicus may adapt to temperature changes by regulating protein synthesis and carbon metabolism pathways, changing post-translational modifications, and improving protein stability at the same time.
Assuntos
Proteínas Arqueais , Sulfolobus , Proteínas Arqueais/genética , Carbono/metabolismo , Nucleotídeos/metabolismo , Proteoma/metabolismo , RNA , Sulfolobus/química , Sulfolobus/genética , Sulfolobus/metabolismo , Enxofre/metabolismo , TemperaturaRESUMO
In all organisms, the DNA sequence and the structural organization of chromosomes affect gene expression. The extremely thermophilic crenarchaeon Sulfolobus has one circular chromosome with three origins of replication. We previously revealed that this chromosome has defined A and B compartments that have high and low gene expression, respectively. As well as higher levels of gene expression, the A compartment contains the origins of replication. To evaluate the impact of three-dimensional organization on genome evolution, we characterized the effect of replication origins and compartmentalization on primary sequence evolution in eleven Sulfolobus species. Using single-nucleotide polymorphism analyses, we found that distance from an origin of replication was associated with increased mutation rates in the B but not in the A compartment. The enhanced polymorphisms distal to replication origins suggest that replication termination may have a causal role in their generation. Further mutational analyses revealed that the sequences in the A compartment are less likely to be mutated, and that there is stronger purifying selection than in the B compartment. Finally, we applied the Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) to show that the B compartment is less accessible than the A compartment. Taken together, our data suggest that compartmentalization of chromosomal DNA can influence chromosome evolution in Sulfolobus. We propose that the A compartment serves as a haven for stable maintenance of gene sequences, while sequences in the B compartment can be diversified.