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Author(s): Sara Pedron 1,2,3,*, Michael Hanselmann 1, Jacob Burns 1,4,5, Alexander Rich 2,4,5, Annette Peters 6, Margit Heier 6,7, Lars Schwettmann 2,8, Jacob H. Bor 9,10, Till Bärnighausen 11,12,13, Michael [...], Background Hypertension represents one of the major risk factors for cardiovascular morbidity and mortality globally. Early detection and treatment of this condition is vital to prevent complications. However, hypertension often goes undetected, and even if detected, not every patient receives adequate treatment. Identifying simple and effective interventions is therefore crucial to fight this problem and allow more patients to receive the treatment they need. Therefore, we aim at investigating the impact of a population-based blood pressure (BP) screening and the subsequent "low-threshold" information treatment on long-term cardiovascular disease (CVD) morbidity and mortality. Methods and findings We examined the impact of a BP screening embedded in a population-based cohort study in Germany and subsequent personalized "light touch" information treatment, including a hypertension diagnosis and a recommendation to seek medical attention. We pooled four waves of the KORA study, carried out between 1984 and 1996 (N = 14,592). Using a sharp multivariate regression discontinuity (RD) design, we estimated the impact of the information treatment on CVD mortality and morbidity over 16.9 years. Additionally, we investigated potential intermediate outcomes, such as hypertension awareness, BP, and behavior after 7 years. No evidence of effect of BP screening was observed on CVD mortality (hazard ratio (HR) = 1.172 [95% confidence interval (CI): 0.725, 1.896]) or on any (fatal or nonfatal) long-term CVD event (HR = 1.022 [0.636, 1.641]) for individuals just above (versus below) the threshold for hypertension. Stratification for previous self-reported diagnosis of hypertension at baseline did not reveal any differential effect. The intermediate outcomes, including awareness of hypertension, were also unaffected by the information treatment. However, these results should be interpreted with caution since the analysis might not be sufficiently powered to detect a potential intervention effect. Conclusions The study does not provide evidence of an effect of the assessed BP screening and subsequent information treatment on BP, health behavior, or long-term CVD mortality and morbidity. Future studies should consider larger datasets to detect possible effects and a shorter follow-up for the intermediate outcomes (i.e., BP and behavior) to detect short-, medium-, and long-term effects of the intervention along the causal pathway.

Time is a central aspect of exploratory choice. People must balance the immediate rewards of exploiting known alternatives against the long-term rewards of exploring uncertain alternatives. However, little research has investigated how this temporal aspect affects the exploratory decisions people make. In other intertemporal choices, people display a bias towards immediate rewards. We hypothesize that in exploratory choice, this present bias will cause them to under-explore. Across three experiments, including a preregistered design, we find no evidence that present bias influences exploratory choice, but also conclude that our stimuli may not effectively induce present bias. We discuss how present bias might better be investigated, and possible reasons that present bias may not affect exploratory choice. Keywords: exploration, decision making, intertemporal choice, present bias, Decision makers acting in uncertain environments frequently face the dilemma between choosing options known to be rewarding (i.e., exploitation) and choosing options that are unknown or uncertain (i.e., exploration). A [...]

The double helix of DNA, when composed of dinucleotide purine-pyrimidine repeats, can adopt a left-handed helical structure called Z-DNA. For reasons not entirely understood, such dinucleotide repeats in genomic sequences have been associated with genomic instability leading to cancer. Adoption of the left-handed conformation results in the formation of conformational junctions: A B-to-Z junction is formed at the boundaries of the helix, whereas a Z-to-Z junction is commonly formed in sequences where the dinucleotide repeat is interrupted by single base insertions or deletions that bring neighboring helices out of phase. B-Z junctions are shown to result in exposed nucleotides vulnerable to chemical or enzymatic modification. Here we describe the three-dimensional structure of a Z-Z junction stabilized by Z[alpha], the Z-DNA binding domain of the RNA editing enzyme ADAR1. We show that the junction structure consists of a single base pair and leads to partial or full disruption of the helical stacking. The junction region allows intercalating agents to insert themselves into the left-handed helix, which is otherwise resistant to intercalation. However, unlike a B-Z junction, in this structure the bases are not fully extruded, and the stacking between the two left-handed helices is not continuous. base stacking | DNA structure | extruded base | X-ray diffraction doi/ 10.1073/pnas.1003182107

Mammalian DAI (DNA-dependent activator of IFN-regulatory factors), an activator of the innate immune response, senses cytosolic DNA by using 2 N-terminal Z-DNA binding domains (ZBDs) and a third putative DNA binding domain located next to the second ZBD. Compared with other previously known ZBDs, the second ZBD of human DAI (hZ[[beta].sub.DAI]) shows significant variation in the sequence of the residues that are essential for DNA binding. In this article, the crystal structure of the hZ[[beta].sub.DAI]/Z-DNA complex reveals that hZ[[beta].sub.DAI] has a similar fold to that of other ZBDs, but adopts an unusual binding mode for recognition of Z-DNA. A residue in the first [beta]-strand rather than residues in the [beta]-loop contributes to DNA binding, and part of the ([alpha]3) recognition helix adopts a [3.sub.10] helix conformation. The role of each residue that makes contact with DNA was confirmed by mutational analysis. The 2 ZBDs of DAI can together bind to DNA and both are necessary for full B-to-Z conversion. It is possible that binding 2 DAIs to 1 dsDNA brings about dimerization of DAI that might facilitate DNA-mediated innate immune activation. circular dichroism | hydrogen bonding | interferon induction | X-ray crystallography | innate immunity

In this article, the effect of a d(CG) DNA dinucleotide repeat sequence on RNA polymerase II transcription is examined in yeast Saccharomyces cerevisiae. Our previous report shows that a d[(CG).sub.n] dinucleotide repeat sequence located proximally upstream of the TATA box enhances transcription from a minimal CYC1 promoter in a manner that depends on its surrounding negative supercoiling. Here, we demonstrate that the d[(CG).sub.g] repeat sequence stimulates gene activity by forming a Z-DNA secondary structure. Furthermore, the extent of transcriptional enhancement by Z-DNA is promoter-specific and determined by its separation distance relative to the TATA box. The stimulatory effect exerted by promoter proximal Z-DNA is not affected by helical phasing relative to the TATA box, suggesting that Z-DNA effects transcription without interacting with the general transcription machinery by looping-out the intervening DNA. A nucleosome-scanning assay reveals that the d[(CG).sub.g] repeat sequence in the Z conformation blocks nucleosome formation, and it is found in the linker DNA with two flanking nucleosomes. This result suggests that Z-DNA formation proximally upstream of a promoter is sufficient to demarcate the boundaries of its neighboring nucleosomes, which produces transcriptionally favorable locations for the TATA box near the nucleosomal DNA-entry site and at dyad positions on the nucleosome. These findings suggest that Z-DNA formation in chromatin is a part of the 'genomic code' for nucleosome positioning in vivo. chromatin remodeling | promoter | transcription | nucleosome positioning

Author(s): Sung Chul Ha [1]; Ky Lowenhaupt [3]; Alexander Rich [3]; Yang-Gyun Kim (corresponding author) [4]; Kyeong Kyu Kim (corresponding author) [1, 2] Left-handed Z-DNA is a higher-energy form of [...]

The vaccinia virus (VV) E3L protein is essential for virulence and has anti-apoptotic activity. In mice, Z-DNA-binding activity of the N-terminal domain of E3L (Z[alpha]) is necessary for viral lethality. Here, we report that inhibition of hygromycin-B-induced apoptosis in HeLa cells depends on Z-DNA binding of the E3L Z[alpha] domain. Z-DNA-binding domains of other proteins are equally effective in blocking apoptosis. Using a transient reporter assay, we demonstrate transactivation of human IL-6, nuclear factor of activated T cells (NF-AT), and p53 genes by E3L. This activation also requires Z-DNA binding of the N-terminal domain of E3L. Overall, this work suggests that the important role of E3L in VV pathogenesis involves modulating expression of host cellular genes at the transcriptional level and inhibiting apoptosis of host cells through Z-DNA binding. apoptosis | interleukin-6 | p53 | poxvirus

The double-stranded RNA (dsRNA)-dependent protein kinase (PKR) is induced as part of the IFN response in mammals and acts to shut down protein synthesis by the phosphorylation of eukaryotic initiation factor 2[alpha] (elF2[alpha]). In fish, a PKR-like kinase activity has been detected, but the enzyme responsible has eluded characterization. Here, we describe a PKR-like kinase from zebrafish. PhyIogenetic analysis shows that the C-terminal kinase domain is more closely related to the kinase domain of PKR than to any of the other three known elF2[alpha] kinases. Surprisingly, instead of the two dsRNA binding domains found at the N terminus of PKR, there are two Z[alpha] domains. Z[alpha] domains specifically bind dsDNA and RNA in the left-handed Z conformation, often with high affinity. They have been found previously in two other IFN-inducible proteins, the dsRNA editing enzyme, ADAR1, and Z-DNA binding protein 1 (ZBP1), as well as in the poxvirus virulence factor, E3L. This previously undescribed kinase, designated PKZ (protein kinase containing Z-DNA binding domains), is transcribed constitutively at low levels and is highly induced after injection of poly(inosinic)--poly(cytidylic) acid, which simulates viral infection. Binding of Z-DNA by the Z[alpha] domain of PKZ was demonstrated by circular dichroism. PKZ inhibits translation in transfected cells; site-directed mutagenesis indicates that this inhibition depends on its catalytic activity. Identification of a gene combining Z[alpha] domains with a PKR-like kinase domain strengthens the hypothesis that the ability to bind left-handed nucleic acid plays a role in the host response to viruses. E3L | interferon response | viral infection | Z-RNA | Z[alpha] domain

A conserved feature of poxviruses is a protein, well characterized as E3L in vaccinia virus, that confers IFN resistance on the virus. This protein comprises two domains, an N-terminal Z-DNA-binding protein domain (Z[alpha]) and a C-terminal double-stranded RNA-binding domain. Both are required for pathogenicity of vaccinia virus in mice infected by intracranial injection. Here, we describe the crystal structure of the Z[alpha] domain from the E3L-like protein of Yaba-like disease virus, a Yatapoxvirus, in a complex with Z-DNA, solved at a 2.0-[Angstrom] resolution. The DNA contacting surface of Yaba-like disease virus [Z.sup.[alpha]E3L] closely resembles that of other structurally defined members of the Z[alpha] family, although some variability exists in the [beta]-hairpin region. In contrast to the Z-DNA-contacting surface, the nonbinding surface of members of the Z-DNA-binding family are unrelated; this surface may effect protein-specific interactions. The presence of the conserved and tailored Z-DNA-binding surface, which interacts specifically with the zigzag backbone and syn base diagnostic of the Z-form, reinforces the importance to poxvirus infection of the ability of this protein to recognize the Z-conformation.

The Noterminal domain of the vaccinia virus protein E3L ([Z.sub.[alpha]E3L]) is essential for full viral pathogenicity in mice. It has sequence similarity to the high-affinity human Z-DNA-binding domains [Z.sub.[alpha]ADAR1] and [Z.sub.[alpha]DLM1], Here, we report the solution structure of [Z.sub.[alpha]E3L] and the chemical shift map of its interaction surface with Z-DNA. The global structure and the Z-DNA interaction surface of [Z.sub.[alpha]E3L] are vary similar to the high-affinity Z-DNA-binding domains [Z.sub.[alpha]ADAR1] and [Z.sub.[alpha]DLM1]. However, the key Z-DNA contacting residue Y48 of [Z.sub.[alpha]E3L] adopts a different side chain conformation in unbound [Z.sub.[alpha]E3L]. which requires rearrangement for binding to Z-DNA. This difference suggests a molecular basis for the significantly lower in vitro affinity of [Z.sub.[alpha]E3L] to Z-DNA compared with its homologues.

The E3L gene product found in all poxviruses is required for the lethality of mice in vaccinia virus infection. Both the C-terminal region, consisting of a double-stranded RNA-binding motif, and the N-terminal region (VZ[E.sub.3L]), which is similar to the Z [alpha] family of Z-DNA-binding proteins, are required for infection. It has recently been demonstrated that the function of the N-terminal domain depends on its ability to bind Z-DNA; Z-DNA-binding domains from unrelated mammalian proteins fully complement an N-terminal deletion of E3L. Mutations that decrease affinity for Z-DNA have similar effects in decreasing pathogenicity. Compounds that block the Z-DNA-binding activity of E3L may also limit infection by the poxvirus. Here we show both an in vitro and an in vivo assay with the potential to be used in screening for such compounds. Using a conformation-specific yeast one-hybrid assay, we compared the results for Z-DNA binding of VZ[E.sub.3L] with those for human Z[Beta]ADAR1, a peptide that has similarity to the Z[alpha] motif but does not bind Z-DNA, and with a mutant of hZ[Beta]ADAR1, which binds Z-DNA. The results suggest that this system can be used for high-throughput screening.

The N-terminal domain of the E3L protein of vaccinia virus has sequence similarity to a family of Z-DNA binding proteins of defined three-dimensional structure and it is necessary for pathogenicity in mice. When other Z-DNA-binding domains are substituted for the similar E3L domain, the virus retains its lethality after intracranial inoculation. Mutations decreasing Z-DNA binding in the chimera correlate with decreases in viral pathogenicity, as do analogous mutations in wild-type E3L. A chimeric virus incorporating a related protein that does not bind Z-DNA is not pathogenic, but a mutation that creates Z-DNA binding makes a lethal virus. The ability to bind the Z conformation is thus essential to E3L activity. This finding may allow the design of a class of antiviral agents, including agents against variola (smallpox), which has an almost identical E3L.

The role of Z-DNA-binding proteins in vivo is explored in yeast. A conformation-specific yeast one-hybrid system is made in which formation of Z-DNA is studied near a minimal promoter site where it can be stabilized by negative supercoiling in addition to protein binding. Experiments were carried out with a Z-DNA-binding protein domain from the editing enzyme, double-stranded RNA adenosine deaminase 1. In the one-hybrid system, the reporter gene is activated when a Z-DNA-specific binding domain is fused with an activation domain and expressed in vivo. Significantly, it was found that even in the absence of the activation domain there is substantial transcription of the reporter gene if the Z-DNA-binding protein is expressed in the cell. This result suggests that Z-DNA formation in the promoter region induced or stabilized by a Z-DNA-binding protein can act as a cis-element in gene regulation. Related results have been found recently when the human chromatin-remodeling system converts a segment of DNA in the promoter region of the human colony-stimulating factor 1 gene into the left-handed Z-conformation.

Many pathogenic viruses use programmed -1 ribosomal frameshifting to regulate translation of their structural and enzymatic proteins from polycistronic mRNAs. Frameshifting is commonly stimulated by a pseudoknot located downstream from a slippery sequence, the latter positioned at the ribosomal A and P sites. We report here the structures of two crystal forms of the frameshifting RNA pseudoknot from beet western yellow virus at resolutions of 1.25 and 2.85 [Angstrom]. Because of the very high resolution of 1.25 [Angstrom], ten mono- and divalent metal ions per asymmetric unit could be identified, giving insight into potential roles of metal ions in stabilizing the pseudoknot. A magnesium ion located at the junction of the two pseudoknot stems appears to play a crucial role in stabilizing the structure. Because the two crystal forms exhibit mostly unrelated packing interactions and local crystallographic disorder in the high-resolution form was resolvable, the two structures offer the most detailed view yet of the conformational preference and flexibility of an RNA pseudoknot.

In mammals, RNA editing by site-selective adenosine deamination regulates key functional properties of neurotransmitter receptors in the central nervous system. Glutamate receptor subunit B is nearly 100% edited at one position (the Q/R-site), which is essential for normal receptor function. Its significance is apparent from mouse models in which a slightly reduced rate of Q/R-site editing is associated with early onset epilepsy and premature death. Here we report that in tissues from malignant human brain tumors, this editing position of glutamate receptor subunit B is substantially underedited compared with control tissues. We also observe alterations in editing and alternative splicing of serotonin receptor 5-H[T.sub.2C] transcripts. These changes correlate with a decrease in enzymatic activity of the editing enzyme adenosine deaminase acting on RNA (ADAR) 2, as deduced from analysis of ADAR2 self-editing. Our results suggest a role for RNA editing in tumor progression and may provide a molecular model explaining the occurrence of epileptic seizures in association with malignant gliomas.

The mammalian adenosine deaminases acting on RNA (ADARs) constitute a family of sequence-related proteins involved in pre-mRNA editing of nuclear transcripts through site-specific adenosine modification. We report here the identification and characterization of a human ADAR protein, hADAT1, that specifically deaminates adenosine 37 to inosine in eukaryotic [tRNA.sup.Ala]. It represents the functional homologue of the recently identified yeast protein Tad1p [Gerber, A., Grosjean, H., Melcher, T. & Keller, W. (1998) EMBO J. 17, 4780-4789]. The hADAT1 cDNA predicts a protein of 502 aa whose sequence displays strongest overall homology to a Drosophila melanogaster ORF (50% similarity, 32% identity), and the catalytic domain is closely related to the other ADAR proteins. In vitro, the recombinantly expressed and purified hADAT1 protein efficiently and specifically deaminates [A.sub.37] in the anticodon loop of [tRNA.sup.Ala] from higher eukaryotes and with lower efficiency from lower eukaryotes. It does not modify adenosines residing in double-stranded RNA or in pre-mRNAs that serve as substrates for ADAR1 or ADAR2. The anticodon stem-loop of [tRNA.sup.Ala] alone is not a functional substrate for hADAT1. The enzyme is expressed ubiquitously in human tissues and is represented by a single gene. The identification and cloning of hADAT1 should help to elucidate the physiological significance of this unique modification in [tRNA.sup.Ala], which is conserved from yeast to man.

Zalpha, an N-terminal domain of human double-stranded RNA adenosine deaminase (ADAR1), was found to interact with Z-DNA. It was determined whether Zalpha can stabilize the left-handed Z conformation adapted by unaltered, linear alternating (dGdC) polymer including the methylated form. The influence of Zalpha on short, methylated and unmethylated deoxycytidine-deoxyguanosine oligonucleotides was also studied. Results showed that Zalpha recognizes and binds to unaltered poly(dGdC) and also short d(CG)n oligonucleotide sequences.

Novel restriction enzymes can be created by fusing the nuclease domain of FokI endonuclease with defined DNA binding domains. Recently, we have characterized a domain (Z[Alpha]) from the N-terminal region of human double-stranded RNA adenosine deaminase (hADARI), which binds the Z-conformation with high specificity. Here we report creation of a conformation-specific endonuclease, Z[Alpha] nuclease, which is a chimera of Z[Alpha] and FokI nuclease. Purified Z[Alpha] nuclease cleaves negatively supercoiled plasmids only when they contain a Z-DNA forming insert, such as [(dC-dG).sub.(13)]. The precise location of the cleavage sites was determined by primer extension. Cutting has been mapped to the edge of the B-Z junction, suggesting that Z[Alpha] nuclease binds within the Z-DNA insert, but cleaves in the nearby B-DNA, by using a mechanism similar to type IIs restriction enzymes. These data show that Z[Alpha] binds Z-DNA in an environment similar to that in a cell. Z[Alpha] nuclease, a structure-specific restriction enzyme, may be a useful tool for further study of the biological role of Z-DNA.

Editing of RNA changes the read-out of information from DNA by altering the nucleotide sequence of a transcript. One type of RNA editing found in all metazoans uses double-stranded RNA (dsRNA) as a substrate and results in the deamination of adenosine to give inosine, which is translated as guanosine. Editing thus allows variant proteins to be produced from a single pre-mRNA. A mechanism by which dsRNA substrates form is through pairing of intronic and exonic sequences before the removal of noncoding sequences by splicing. Here we report that the RNA editing enzyme, human dsRNA adenosine deaminase (DRADA1, or ADAR1) contains a domain (Z[Alpha]) that binds specifically to the left-handed Z-DNA conformation with high affinity ([K.sub.D] = 4 nM). As formation of Z-DNA in vivo occurs 5[prime] to, or behind, a moving RNA polymerase during transcription, recognition of Z-DNA by DRADA1 provides a plausible mechanism by which DRADA1 can be targeted to a nascent RNA so that editing occurs before splicing. Analysis of sequences related to Z[Alpha] has allowed identification of motifs common to this class of nucleic acid binding domain.

A 16-amino acid oligopeptide forms a stable [Beta]-sheet structure in water. In physiological solutions it is able to self-assemble to form a macroscopic matrix that stains with Congo red. On raising the temperature of the aqueous solution above 70[degrees]C, an abrupt structural transition occurs in the CD spectra from a [Beta]-sheet to a stable [Alpha]-helix without a detectable random-coil intermediate. With cooling, it retained the [Alpha]-helical form and took several weeks at room temperature to partially return to the [Beta]-sheet form. Slow formation of the stable [Beta]-sheet structure thus shows kinetic irreversibility. Such a formation of very stable [Beta]-sheet structures is found in the amyloid of a number of neurological diseases. This oligopeptide could be a model system for studying the protein conformational changes that occurs in scrapie or Alzheimer disease. The abrupt and direct conversion from a [Beta]-sheet to an [Alpha]-helix may also be found in other processes, such as protein folding and protein-protein interaction. Furthermore, such drastic structure changes may also be exploited in biomaterials designed as sensors to detect environmental changes.

DNA fragments with stretches of cytosine residues can fold into four-stranded structures in which two parallel duplexes, held together by hemiprotonated cytosine[center dot][cytosine.sup.+] (C[center dot][C.sup.+]) base pairs, intercalate into each other with opposite polarity. The structural details of this intercalated DNA quadruplex have been assessed by solution NMR and single crystal x-ray diffraction studies of cytosine-rich sequences, including those present in metazoan telomeres. A conserved feature of these structures is the absence of stabilizing stacking interactions between the aromatic ring systems of adjacent C[center dot][C.sup.+] base pairs from intercalated duplexes. Effective stacking involves only the exocyclic keto groups and amino groups of the cytidine bases. The apparent absence of stability provided by stacking interactions between the bases in this intercalated DNA has prompted us to examine the available structures in detail, in particular with regard to unusual features that could compensate for the lack of base stacking. In addition to base-on-deoxyribose stacking and intra-cytidine C - H...O hydrogen bonds, this analysis reveals the presence of a hitherto unobserved, systematic intermolecular C - H...O hydrogen bonding network between the deoxyribose sugar moieties of antiparallel backbones in the four-stranded molecule.

The intron of the corticotropin-releasing hormone (corticoliberin; CRH) gene contains a sequence of over 100 bp of alternating purine/pyrimidine residues. We have used binding of a Z-DNA-specific antibody in metabolically active, permeabilized nuclei to study the formation of Z-DNA in this sequence at various levels of transcription. In the NPLC human primary liver carcinoma cell line, activation of cAMP-dependent pathways increased the level of transcription, while adding glucocorticoids inhibited transcription of the CRH gene. These cells respond in a manner similar to hypothalamic cells. Z-DNA formation in this sequence was detected at the basal level of transcription, as well as after stimulation with forskolin. Inhibition of transcription by dexamethasone abolished Z-DNA formation. Z-DNA formation in the MYC gene (c-myc) was affected differently in the same experiment. Thus, changes in Z-DNA formation in the CRH gene are gene specific and are linked to the transcription of the gene.

Agarose-encapsulated, metabolically active, permeabilized nuclei for human hematopoietic cell lines were analyzed for Z-DNA formation in the beta-globin gene cluster. Biotinylated monoclonal antibodies against Z-DNA were diffused into the nuclei and cross-linked to DNA. After digestion, the fragments that had formed Z-DNA were isolated. Five of the seventeen regions with Z-DNA motifs in the 73-kb region were found in the Z formation.

A [M.sub.r] 140,000 protein has been purified from chicken lungs to apparent homogeneity. The protein binds with high affinity to a non-BNA conformation, which is most likely to be Z-DNA. The protein also has a binding site for double-stranded RNA (dsRNA). Peptide sequences from this protein show similarity to dsRNA adenosine deaminase, an enzyme that deaminates adenosine in dsRNA to form inosine. Assays for this enzyme confirm that dsRNA adenosine deaminase activity and Z-DNA binding are properties of the same molecule. The coupling of these two activities in a single molecule may indicate a distinctive mechanism of gene regulation that is, in part, dependent on DNA topology. As such, DNA topology, through its effects on the efficiency and extent of RNA editing may be important in the generation of new phenotypes during evolution.

The three-dimensional (3D) structures of many DNA-binding proteins have been solved, and the mechanism of sequence-specific DNA recognition is largely understood. In contrast, little is known about the recognition systems [...], The translational regulator protein regA is encoded by the T4 bacteriophage and binds to a region of messenger RNA (mRNA) that includes the initiator codon. RegA is unusual in that it represses the translation of about 35 early T4 mRNAs but does not affect nearly 200 other mRNAs. The crystal structure of regA was determined at 1.9 A resolution; the protein was shown to have an a-helical core and two regions with antiparallel β sheets. One of these β sheets has four antiparallel strands and has some sequence homology to RNP-1 and RNP-2, which are believed to be RNA-binding motifs and are found in a number of known RNA-binding proteins. Structurally guided mutants may help to uncover the basis for this variety of RNA interaction.

The results of an experiment revealed a stable loop of DNA is not confined to a DNA duplex and associates with four-stranded cytosine structures to reinforce the possibility of conformations for the cytosine-rich telomere strands. This was established by using crystal structure of the telomeric cytosine-rich strand repeating sequence d(TAACCC)that forms a four-stranded cytosine-intercalated structure and utilizes the sequence TAA to form a loop where 5' end of one molecule is very close to 3' end of another.

A structural analysis of the duplex form of the DNA fragment containing 7,8-dihydro-8-oxoguanine (G(O)), d(CCAG(O)CGCTGG), indicates the absence of steric interaction between the phospho-deoxyribose backbone and 8-oxygen of G(O). G(O) is present in the anti conformation and pairs with the opposite C to form Watson-Crick base pairs. The major groove of the fragment contains an unusual pattern of hydrogen bond donors/acceptors that help recognize the elements that enable substrate identification by the DNA repair protein MutM.

X-ray crystallographic analysis of the four-stranded intercalated DNA d(C4) at a resolution of 2.3 angstrom units shows that d(C4) consists of two intercalated complexes linked by hemiprotonated cytosine-cytosine base pairs. The duplexes have parallel strands and two of them exhibit an opposite orientation. The d(C4) molecule also contains a 12.4 angstrom unit, slow, right-handed twist between the cytosine-cytosine base pairs and a 3.1 angstrom unit base stacking distance.

Analysis of the crystal structure of d(C3T) at 1.4 angstrom resolution reveals the presence of a four-stranded intercalated complex containing two intercalated parallel-stranded duplexes that are connected by cytosine-protonated cytosine base pairs. The molecule is asymmetric, having a flat, lath-like appearance, with the covalently bonded cytosine having a right-handed twist of 17.1 degrees. The phosphate groups on one of the flat side are rotated far from the molecule's center and bridging water molecules that bind the NH of cytosine and a phosphate group of the opposite chain retain the orientation of these phosphate groups. The structure exhibits microheterogeneism with the cytosine hemiprotonating at pH 7.

Genetic-mutational studies and structural investigation helped dissect the domain of HAP2 that mediates subunit association in the heteromeric CCAAT-binding complex. Protein-protein association is mediated by an extremely short site in HAP2. An alpha-helix configuration is likely for this domain.

A study of the left-handed Z-DNA duplex pure-spermine form X-ray crystal structure reveals that a hitherto unknown second intrahelix spermine molecule and two hydrated sodium ions are present at a temperature of -110 degrees Celsius. The second spermine molecule is mainly found within the duplexes' minor grooves of the crystal lattice. The intrahelix spermine molecule reduces cross-groove electrostatic repulsion within the Z-DNA helix and enhances stability.

X-ray diffraction analysis was used to examine the molecular structure of the complex found between the minor groove binding drug netropsin and dodecamer d(CGCAAATTTGCG). The results reveal a refinement of 19.8% of the R-factor and a revolution of 2.2-angstroms. Five out of six of the A.T base pairs from A5. T20 to T9.A16 were covered by netropsin from its structural position in the narrow minor groove of the B-DNA fragment. Netropsin is permitted to bind in a position with maximum contact with the DNA by the long six A.T base pair. Hydrogen bonds with O2 of thymines and N3 of adenines are created by the binding.

The properties of self-complementary EAK16 peptide ((Ala-Glu-Ala-Glu-Ala-Lys-Ala-Lys)2) were reported. Circular dichroism studies of EAK16 indicate a spectrum of beta-sheet formation. It spontaneously associates to form an insoluble macroscopic membrane, that is stable and resistant to digestion with several proteases. This peptide structure has implications for biology and for biomaterial research.

The presence and characterization of a Z-DNA binding nuclear activity from chicken blood nuclear extracts were described. The protein (Zalpha), with a minimal molecular mass of 39 kDa, appears to be highly specific for the left-handed Z-DNA conformer. The binding activity of Zalpha is competitively blocked specifically both by brominated poly(dC-dG) in the Z-DNA form and supercoiled plasmid containing a Z-DNA insert.

X-ray crystallography was used to determine the three-dimensional structures of three DNA-RNA chimeric duplexes at a resolution of 2-25 angstroms. The conformations of the self-complementary duplexes (r(G)d(CGTATACGC))2 and (d(GCGT)r(A)d(TACGC))2 and the Okazaki fragment d(GGGTATACGC)r(GCG)d(TATACCC) were found to be similar to those of A-type duplexes. However, significant differences were found in the local geometries of the three structures. Intramolecualr interactions between the ribose 2'-hydroxyl groups were found to contribute to the stability of the A-conformation. It was concluded that a sole RNA residue is enough to induce conformational equilibrium toward such a conformation in DNA-RNA chimeric duplexes.

The Z-DNA-forming sequences in the SV40 enhancer were investigated using DNA minicircles which were synthesized as relaxed circles and then supercoiled by incremental treatment with topoisomerase I in the presence of ethidium bromide. The resulting topoisomers were then resolved by gel electrophoresis. The results showed that enhancer-containing minicircles have a superhelix density of -0.05.

Gender differences in the psychosocial correlates of suicidal ideation were studied. A sample of 613 high school students (ages 14-19) completed measures of suicidal ideation, depression, hopelessness, life stress, loneliness, alcohol and drug use, and reasons for living. The results of a discriminant function analysis indicated that males reported higher loneliness and substance abuse scores than females whereas females reported greater suicidal ideation, depression, and reasons for living. The results of multiple regression analyses found that, although the same four variables, depression, hopelessness, substance abuse, and few reasons for living emerged as significant predictors of suicidal ideation in both samples, the predictive equation accounted for more of the variance in ideation scores in females (57%) than in males (46%). In a final analysis a discriminant function analysis of the subscales of the reasons for living inventory revealed that females have a greater fear of death and injury whereas males have a greater fear of social disapproval over having suicidal thoughts. This may account for the greater rate of suicide completing among males. Fear of social disapproval, more anger and impulsivity, and less help-seeking behavior among males are offered as potential variables to explain the observed gender differences.

The influence of a short RNA segment on double-stranded DNA conformation was determined using X-ray crystallography analysis on synthetic Okazaki fragment. The synthetic Okazaki fragment was formed by combination of DNA and its chimeric RNA-DNA oligonucleotides. The RNA trimer was found to adopt an A-type conformation in order to lock the complete fragment.

A stress-vulnerability model of suicidal behavior among college students was proposed by Bonner and Rich [1, 2]. The purpose of the current study was to evaluate and extend this model using a younger adolescent (i.e., high school) sample. A total of 613 students (328 females and 285 males) completed the self-report measures of life stress, depression, hopelessness, reasons for living, loneliness, alcohol and drug use, and suicidal ideation. These factors served as predictor variables in stepwise multiple regressions with suicidal ideation serving as the criterion variable. Four variables emerged as significant predictors of suicidal ideation: depression, hopelessness, few reasons for living, and problem substance use. The linear combination of these variables accounted for 52 percent of the variance in suicide ideation scores. The substance abuse variable accounted for variance in ideation scores independent of the other factors.

The purpose of this present investigation was to cross-validate a stress--psychosocial vulnerability model of suicidal ideation and behavior in a jail population. Measures of social alienation, cognitive distortions, adaptive resources, situational (jail environment) stress, depression, hopelessness, and suicide ideation were administered to 146 male inmates at a county jail facility. It was predicted that each of the vulnerability factors, in addition to jail stress, would be significantly related to suicide ideation among inmates. Furthermore, it was hypothesized that psychosocial vulnerability among inmates would interact with jail stress to best explain suicide intent. The result of a multiple-regression analysis, stepwise forward-inclusion algorithm, indicated that 51% of the variation in suicide ideation could be accounted for by the linear combination of low reasons for living, irrational beliefs, jail stress, and loneliness. In addition, when the variables were entered into a hierarchical multiple-regression model, interactions between select psychosocial vulnerability factors and jail stress were found to best explain suicide intent. Results of the study are discussed in terms of the proposed model, and implications for future research and prevention efforts in jail suicide are noted.

Author(s): Alexander Rich [1]; Charles F. Stevens [2] [illus. 1] Alexander Rich writes: Francis Crick, who died on 28 July at the age of 88, was trained as a physicist [...]

The crystal structure of a high-efficiency frameshifting mutant of the pseudoknot from potato leaf roll virus (PLRV) is determined to understand the role of structural elements of RNA pseudoknots in controlling the extent of -1-type ribosomal frameshifting. The resulting structure defines a minimal RNA pseudoknot motif composed of 22 nucleotides capable of stimulating-1-type ribosomal frameshifts.

An autobiographical account on the life and experiences of the Alexander Rich, and his contributions in the field of molecular biology is presented. He expresses his great excitement on discoveries made, and relates his success to the motivation provided by his brilliant teachers, colleagues and students.

H-pai and lone pair-pai (l.p.-pai) interactions between water molecules as well as functionally important unstacked residues in the 1.25 Angstrom crystal structure of the ribosomal frameshifting RNA pseudoknot from beet western yellow virus are reported. The virus of the traditional hydrogen bonds N-H...O and O-H...O, include key roles in macromolecular structure and function.

Many sequences in eukaryotic genomes have the potential to adopt a left-handed Z-DNA conformation. We used a previously described assay based on the binding of a mAb to Z-DNA to inquire whether Z-DNA is formed in the rat nucleolin (Ncl) gene in metabolically active, permeabilized nuclei. Using real-time PCR to measure Z-DNA formation, the potential Z-DNA sequence element Z1 [[(CA).sub.10][(CG).sub.8]] in the promoter region was found to be enriched 571- to 4,040-fold in different cell lines, whereas Z2 [AC[(GC).sub.5]CCGT[(CG).sub.2]] in the first intron was enriched 12- to 34-fold. Ncl promoter activity was 1.5- to 16-fold stronger than that of the simian virus 40 promoter and enhancer. This activity was further increased 36-54% when Z1 was deleted. The inhibitory effect of Z1 on Ncl promoter activity was independent of location and orientation. The Ncl Z1 element is identical to the genetic marker D9Arb5. Five allelic variants of Z1 were identified by sequence analysis of genomic DNA from various rats. The two most common alleles differed significantly (up to 27%) in their capacity to inhibit Ncl promoter activity. This finding suggests that differences in Z-DNA formation by polymorphic dinucleotide repeats may be one of the factors contributing to genetic variation. gene regulation | polymorphism | microsatellites | left-handed DNA

The Z[Alpha] domain of human double-stranded RNA adenosine deaminase 1 binds specifically to left-handed Z-DNA and stabilizes the Z-conformation. Here we report spectroscopic and analytical results that demonstrate that Z[Alpha] can also stabilize the left-handed Z-conformation in double-stranded RNA. Z[Alpha] induces a slow transition from the right-handed A-conformation to the Z-form in duplex r[(CG).sub.6], with an activation energy of 38 kcal [mol.sup.-1]. We conclude that Z-RNA as well as Z-DNA can be accommodated in the tailored binding site of Z[Alpha]. The specific binding of Z-RNA by Z[Alpha] may be involved in targeting double-stranded RNA adenosine deaminase 1 for a role in hypermutation of RNA viruses.

A new type of self-assembling peptide (sapeptide) scaffolds that serve as substrates for neurite outgrowth and synapse formation is described. These peptide-based scaffolds are amenable to molecular design by using chemical or biotechnological syntheses. They can be tailored to a variety of applications. The sapeptide scaffolds are formed through the spontaneous assembly of ionic self-complementary [Beta]-sheet oligopeptides under physiological conditions, producing a hydrogel material. The scaffolds can support neuronal cell attachment and differentiation as well as extensive neurite outgrowth. Furthermore, they are permissive substrates for functional synapse formation between the attached neurons. That primary rat neurons form active synapses on such scaffold surfaces in situ suggests these scaffolds could be useful for tissue engineering applications. The buoyant sapeptide scaffolds with attached cells in culture can be transported readily from one environment to another. Furthermore, these peptides did not elicit a measurable immune response or tissue inflammation when introduced into animals. These biological materials created through molecular design and self assembly may be developed as a biologically compatible scaffold for tissue repair and tissue engineering. biological materials | cell attachment | molecular material design | primary neurons | PC12 cells