CRISPR-Cas type I-A Cascade complex couples viral infection surveillance to host transcriptional regulation in the dependence of Csa3b

CRISPR-Cas (clustered regularly interspaced short palindromic repeats and the associated genes) constitute adaptive immune systems in bacteria and archaea and they provide sequence specific immunity against foreign nucleic acids. CRISPR-Cas systems are activated by viral infection. However, little is known about how CRISPR-Cas systems are activated in response to viral infection or how their expression is controlled in the absence of viral infection. Here, we demonstrate that both the transcriptional regulator Csa3b, and the type I-A interference complex Cascade, are required to transcriptionally repress the interference gene cassette in the archaeon Sulfolobus. Csa3b binds to two palindromic repeat sites in the promoter region of the cassette and facilitates binding of the Cascade to the promoter region. Upon viral infection, loading of Cascade complexes onto crRNA-matching protospacers leads to relief of the transcriptional repression. Our data demonstrate a mechanism coupling CRISPR-Cas surveillance of protospacers to transcriptional regulation of the interference gene cassette thereby allowing a fast response to viral infection

An archaeal CRISPR type III-B system exhibiting distinctive RNA targeting features and mediating dual RNA and DNA interference

CRISPR-Cas systems provide a small RNA-based mechanism to defend against invasive genetic elements in archaea and bacteria. To investigate the in vivo mechanism of RNA interference by two type III-B systems (Cmr-α and Cmr-β) in Sulfolobus islandicus, a genetic assay was developed using plasmids carrying an artificial mini-CRISPR (AC) locus with a single spacer. After pAC plasmids were introduced into different strains, Northern analyses confirmed that mature crRNAs were produced from the plasmid-borne CRISPR loci, which then guided gene silencing to target gene expression. Spacer mutagenesis identified a trinucleotide sequence in the 3′-region of crRNA that was crucial for RNA interference. Studying mutants lacking Cmr-α or Cmr-β system showed that each Cmr complex exhibited RNA interference. Strikingly, these analyses further revealed that the two Cmr systems displayed distinctive interference features. Whereas Cmr-β complexes targeted transcripts and could be recycled in RNA cleavage, Cmr-α complexes probably targeted nascent RNA transcripts and remained associated with the substrate. Moreover, Cmr-β exhibited much stronger RNA cleavage activity than Cmr-α. Since we previously showed that S. islandicus Cmr-α mediated transcription-dependent DNA interference, the Cmr-α constitutes the first CRISPR system exhibiting dual targeting of RNA and DNA

Causes of Infection after Earthquake, China, 2008

To determine which organisms most commonly cause infection after natural disasters, we cultured specimens from injured earthquake survivors in Wenchuan, China, 2008. Of 123 cultures, 46 (59%) grew only 1 type of pathogenic bacteria. Smear was more effective than culture for early diagnosis of gas gangrene. Early diagnosis and treatment of wounds are crucial

Nonlinear optical properties in a quantum well with the hyperbolic confinement potential

We have performed theoretical calculation of the nonlinear optical properties in a quantum well (QW) with the hyperbolic confinement potential. Calculation results reveal that the transition energy, oscillator strength, second-order nonlinear optical rectification (OR), geometric factor and nonlinear optical absorption (OA) are strongly affected by the parameters ($\alpha, \sigma$) of the hyperbolic confinement potential. And an increment of the parameter $\alpha$ reduces all these physical quantities, while an increment of the parameter $\sigma$ enhances them, but not for geometric factor. In addition, it is found that one can control the optical properties of QW by tuning these parameters.Comment: 16pages,6 figures,Accepted for publication in Physica

A type III-B CRISPR–Cas effector complex mediating massive target DNA destruction

Funding: Biotechnology and Biological Sciences Research Council [BB/M000400/1 to MFW].The CRISPR (clustered regularly interspaced short palindromic repeats) system protects archaea and bacteria by eliminating nucleic acid invaders in a crRNA-guided manner. The Sulfolobus islandicus type III-B effector Cmr-α complex targets invading nucleic acid at both RNA and DNA levels and DNA targeting relies on the directional transcription of the protospacer in vivo. To gain further insight into the involved mechanism, we purified a native effector complex of III-B Cmr-α from S. islandicus and characterized it in vitro. Cmr-α cleaved RNAs complementary to crRNA present in the complex and its ssDNA destruction activity was activated by target RNA. ssDNA cleavage required mismatches between the 5’-tag of crRNA and the 3’-flanking region of target RNA. An invader plasmid assay showed that mutation either in the HD domain (a quadruple mutation) or in the GGDD motif of theCmr-2α protein resulted in attenuation of the DNA interference in vivo. However, double mutation of the HD motif only abolished the DNase activity in vitro. Furthermore, the activated Cmr-α binary complex functioned as a highly active DNase to destroy a large excess of substrate, which could provide a powerful means to rapidly degrade replicating viral DNA.Publisher PDFPeer reviewe