The innate immune system detects infection by employing germline-encoded receptors specific for conserved microbial molecules. Recognition of microbial ligands leads to the production of cytokines, such as type I interferons (IFN), that are essential for successful pathogen elimination. Cytosolic detection of pathogen-derived DNA is one major mechanism of IFN induction1,2, and requires signaling via Tank Binding Kinase 1 (TBK1), and its downstream transcription factor, Interferon Regulatory Factor 3 (IRF3). In addition, a transmembrane protein called STING (STimulator of INterferon Genes; also called MITA, ERIS, MPYS, TMEM173) functions as an essential signaling adaptor linking cytosolic detection of DNA to the TBK1/IRF3 signaling axis3–7. Recently, unique nucleic acids called cyclic dinucleotides, which function as conserved signaling molecules in bacteria8, were also shown to induce a STING-dependent type I interferon response9–12. However, a mammalian sensor of cyclic dinucleotides has not been identified. Here we report evidence that STING itself is an innate immune sensor of cyclic dinucleotides. We demonstrate that STING binds directly to radiolabelled cyclic diguanylate monophosphate (c-di-GMP) and that this binding is competed by unlabelled cyclic dinucleotides but not by other nucleotides or nucleic acids. Furthermore, we identify mutations in STING that selectively affect the response to cyclic dinucleotides without affecting the response to DNA. Thus, STING appears to function as a direct sensor of cyclic dinucleotides, in addition to its established role as a signaling adaptor in the interferon response to cytosolic DNA. Cyclic dinucleotides have shown promise as novel vaccine adjuvants and immunotherapeutics9,13. Our results provide insight into the mechanism by which cyclic dinucleotides are sensed by the innate immune system.

Telomerase adds telomeric repeats to chromosome ends using an internal RNA template and specialized telomerase reverse transcriptase (TERT), thereby maintaining genome integrity. Little is known about the physical relationships among protein and RNA subunits within a biologically functional holoenzyme. Here we describe the architecture of Tetrahymena thermophila telomerase holoenzyme determined by electron microscopy. Six of the 7 proteins and the TERT-binding regions of telomerase RNA (TER) have been localized by affinity labeling. Fitting with high-resolution structures reveals the organization of TERT, TER, and p65 in the RNP catalytic core. p50 has an unanticipated role as a hub between the RNP catalytic core, p75-p19-p45 subcomplex, and the DNA-binding Teb1. A complete in vitro holoenzyme reconstitution assigns function to these interactions in processive telomeric repeat synthesis. These studies provide the first view of the extensive network of subunit associations necessary for telomerase holoenzyme assembly and physiological function.

SlyD is a putative folding helper protein from the Escherichia coli cytosol, which consists of an N-terminal prolyl isomerase domain of the FKBP type and a presumably unstructured C-terminal tail. We produced truncated versions without this tail (SlyD) for SlyD from E. coli, as well as for the SlyD orthologues from Yersinia pestis, Treponema pallidum, Pasteurella multocida, and Vibrio cholerae. They are monomeric in solution and unfold reversibly. All SlyD variants catalyze the proline-limited refolding of ribonuclease T1 with very high efficiencies, and the specificity constants (kcat/KM) are equal to approximately 10(6) M(-1) s(-1). These large values originate from the high affinities of the SlyD orthologues for unfolded RCM-T1, which are reflected in low KM values of approximately 1 microM. SlyD also exhibits pronounced chaperone properties. Permanently unfolded proteins bind with high affinity to SlyD and thus inhibit its prolyl isomerase activity. The unfolded protein chains do not need to contain proline residues to be recognized and bound by SlyD. The conservation of prolyl isomerase activity and chaperone properties within the SlyD family suggests that these proteins might act as true folding helpers in the bacterial cytosol. The SlyD proteins are also well suited for biotechnological applications. As fusion partners they facilitate the refolding and increase the solubility of aggregation-prone proteins such as the gp41 ectodomain fragment of HIV-1.

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A new group of nitrogen-fixing Azospirillum sp. bacteria was isolated from the roots of the C 4 -gramineous plant Miscanthus. Polyphasic taxonomy was performed, including auxanography using API galleries, physiological tests and T ; reference strain Ma4 l DSM 13400). Its GMC content is 707 mol %.

Prolyl cis-trans isomerizations are intrinsically slow reactions and known to be rate-limiting in many protein folding reactions. Here we report that a proline is used as a molecular timer in the infection of Escherichia coli cells by the filamentous phage fd. The phage is activated for infection by the disassembly of the two N-terminal domains, N1 and N2, of its gene-3-protein, which is located at the phage tip. Pro213, in the hinge between N1 and N2, sets a timer for the infective state. The timer is switched on by cis-to-trans and switched off by the unusually slow trans-to-cis isomerization of the Gln212-Pro213 peptide bond. The switching rate and thus the infectivity of the phage are determined by the local sequence around Pro213, and can be tuned by mutagenesis.

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