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Toward the goal of identifying complete sets of transcription factor (TF)-binding sites in the genomes of several gamma proteobacteria, and hence describing their transcription regulatory networks, we present a phylogenetic footprinting method for identifying these sites. Probable transcription regulatory sites upstream of Escherichia coli genes were identified by cross-species comparison using an extended Gibbs sampling algorithm. Close examination of a study set of 184 genes with documented transcription regulatory sites revealed that when orthologous data were available from at least two other gamma proteobacterial species, 81% of our predictions corresponded with the documented sites, and 67% corresponded when data from only one other species were available. That the remaining predictions included bona fide TF-binding sites was proven by affinity purification of a putative transcription factor (YijC) bound to such a site upstream of the fabA gene. Predicted regulatory sites for 2097 E.coli genes are available at http://www.wadsworth.org/resnres/bioinfo/.

Abstract Background The procedural aspects of genome sequencing and assembly have become relatively inexpensive, yet the full, accurate structural annotation of these genomes remains a challenge. Next-generation sequencing transcriptomics (RNA-Seq), global microarrays, and tandem mass spectrometry (MS/MS)-based proteomics have demonstrated immense value to genome curators as individual sources of information, however, integrating these data types to validate and improve structural annotation remains a major challenge. Current visual and statistical analytic tools are focused on a single data type, or existing software tools are retrofitted to analyze new data forms. We present Visual Exploration and Statistics to Promote Annotation (VESPA) is a new interactive visual analysis software tool focused on assisting scientists with the annotation of prokaryotic genomes though the integration of proteomics and transcriptomics data with current genome location coordinates. Results VESPA is a desktop Java™ application that integrates high-throughput proteomics data (peptide-centric) and transcriptomics (probe or RNA-Seq) data into a genomic context, all of which can be visualized at three levels of genomic resolution. Data is interrogated via searches linked to the genome visualizations to find regions with high likelihood of mis-annotation. Search results are linked to exports for further validation outside of VESPA or potential coding-regions can be analyzed concurrently with the software through interaction with BLAST. VESPA is demonstrated on two use cases (Yersinia pestis Pestoides F and Synechococcus sp. PCC 7002) to demonstrate the rapid manner in which mis-annotations can be found and explored in VESPA using either proteomics data alone, or in combination with transcriptomic data. Conclusions VESPA is an interactive visual analytics tool that integrates high-throughput data into a genomic context to facilitate the discovery of structural mis-annotations in prokaryotic genomes. Data is evaluated via visual analysis across multiple levels of genomic resolution, linked searches and interaction with existing bioinformatics tools. We highlight the novel functionality of VESPA and core programming requirements for visualization of these large heterogeneous datasets for a client-side application. The software is freely available at https://www.biopilot.org/docs/Software/Vespa.php.

Abstract Background EtrA in Shewanella oneidensis MR-1, a model organism for study of adaptation to varied redox niches, shares 73.6% and 50.8% amino acid sequence identity with the oxygen-sensing regulators Fnr in E. coli and Anr in Pseudomonas aeruginosa, respectively; however, its regulatory role of anaerobic metabolism in Shewanella spp. is complex and not well understood. Results The expression of the nap genes, nrfA, cymA and hcp was significantly reduced in etrA deletion mutant EtrA7-1; however, limited anaerobic growth and nitrate reduction occurred, suggesting that multiple regulators control nitrate reduction in this strain. Dimethyl sulfoxide (DMSO) and fumarate reductase gene expression was down-regulated at least 2-fold in the mutant, which, showed lower or no reduction of these electron acceptors when compared to the wild type, suggesting both respiratory pathways are under EtrA control. Transcript analysis further suggested a role of EtrA in prophage activation and down-regulation of genes implicated in aerobic metabolism. Conclusion In contrast to previous studies that attributed a minor regulatory role to EtrA in Shewanella spp., this study demonstrates that EtrA acts as a global transcriptional regulator and, in conjunction with other regulators, fine-tunes the expression of genes involved in anaerobic metabolism in S. oneidensis strain MR-1. Transcriptomic and sequence analyses of the genes differentially expressed showed that those mostly affected by the mutation belonged to the "Energy metabolism" category, while stress-related genes were indirectly regulated in the mutant possibly as a result of a secondary perturbation (e.g. oxidative stress, starvation). We also conclude based on sequence, physiological and expression analyses that this regulator is more appropriately termed Fnr and recommend this descriptor be used in future publications.

Abstract Background It is well established in E. coli and Vibrio cholerae that strains harboring mutations in the ferric uptake regulator gene (fur) are unable to utilize tricarboxylic acid (TCA) compounds, due to the down-regulation of key TCA cycle enzymes, such as AcnA and SdhABCD. This down-regulation is mediated by a Fur-regulated small regulatory RNA named RyhB. It is unclear in the γ-proteobacterium S. oneidensis whether TCA is also regulated by Fur and RyhB. Results In the present study, we showed that a fur deletion mutant of S. oneidensis could utilize TCA compounds. Consistently, expression of the TCA cycle genes acnA and sdhA was not down-regulated in the mutant. To explore this observation further, we identified a ryhB gene in Shewanella species and experimentally demonstrated the gene expression. Further experiments suggested that RyhB was up-regulated in fur mutant, but that AcnA and SdhA were not controlled by RyhB. Conclusions These cumulative results delineate an important difference of the Fur-RyhB regulatory cycle between S. oneidensis and other γ-proteobacteria. This work represents a step forward for understanding the unique regulation in S. oneidensis.

Abstract Background When transcription factor binding sites are known for a particular transcription factor, it is possible to construct a motif model that can be used to scan sequences for additional sites. However, few statistically significant sites are revealed when a transcription factor binding site motif model is used to scan a genome-scale database. Methods We have developed a scanning algorithm, PhyloScan, which combines evidence from matching sites found in orthologous data from several related species with evidence from multiple sites within an intergenic region, to better detect regulons. The orthologous sequence data may be multiply aligned, unaligned, or a combination of aligned and unaligned. In aligned data, PhyloScan statistically accounts for the phylogenetic dependence of the species contributing data to the alignment and, in unaligned data, the evidence for sites is combined assuming phylogenetic independence of the species. The statistical significance of the gene predictions is calculated directly, without employing training sets. Results In a test of our methodology on synthetic data modeled on seven Enterobacteriales, four Vibrionales, and three Pasteurellales species, PhyloScan produces better sensitivity and specificity than MONKEY, an advanced scanning approach that also searches a genome for transcription factor binding sites using phylogenetic information. The application of the algorithm to real sequence data from seven Enterobacteriales species identifies novel Crp and PurR transcription factor binding sites, thus providing several new potential sites for these transcription factors. These sites enable targeted experimental validation and thus further delineation of the Crp and PurR regulons in E. coli. Conclusion Better sensitivity and specificity can be achieved through a combination of (1) using mixed alignable and non-alignable sequence data and (2) combining evidence from multiple sites within an intergenic region.

Background: Mobile apps addressing a variety of workplace safety issues have proliferated over the last decade as mobile technology has advanced and smartphone ownership has increased. Workplace safety interventions are often designed for a specific work site. However, some of the most dangerous jobs are ones in which workers frequently change field locations, such as commercial fishing. Mobile apps may be particularly suitable for delivering safety interventions to these workers., Objective: We sought to gauge the potential for using mobile apps to deliver safety interventions to commercial fishing workers. The purpose of this paper is to describe how fishermen use their mobile devices during fishing operations and identify any mobile apps they already use for safety., Methods: Participants comprised commercial fishing captains who already owned an iOS or Android smartphone or tablet. They completed a questionnaire that asked about their current mobile device use and their use of safety-related mobile apps, in addition to questions about their fishing operations. We performed descriptive analyses of the data., Results: A total of 61 participants completed the questionnaire. The most common types of mobile devices participants reported owning were iPhones (n=36, 59%) and Android phones (n=24, 39%). Most participants (n=53, 87%) reported using their mobile device for both work and personal purposes, including while out at sea (n=52, 85%). Over half of the participants reported that they had either safety-related apps (n=17, 28%) or apps that help them with their work (n=35, 57%). The types of apps most frequently mentioned were apps for weather, wind, tides, and navigation., Conclusions: The results of this study indicate that some commercial fishing captains who own a mobile device are receptive to using safety-related apps for work. Apps that help avoid hazards by monitoring environmental conditions and apps optimized for use on smartphones may be most likely to be adopted and used. Overall, these results suggest that mobile apps are a promising avenue for improving safety among workers in commercial fishing and similar occupations., (©Maria T Bulzacchelli, Jenna M Bellantoni, Leigh McCue, Jerry Dzugan. Originally published in JMIR Formative Research (https://formative.jmir.org), 08.11.2022.)

As prevention trials advance with autosomal dominant Alzheimer disease (ADAD) participants, understanding the similarities and differences between ADAD and "sporadic" late-onset AD (LOAD) is critical to determine generalizability of findings between these cohorts. Cognitive trajectories of ADAD mutation carriers (MCs) and autopsy-confirmed LOAD individuals were compared to address this question. Longitudinal rates of change on cognitive measures were compared in ADAD MCs (n = 310) and autopsy-confirmed LOAD participants (n = 163) before and after symptom onset (estimated/observed). LOAD participants declined more rapidly in the presymptomatic (preclinical) period and performed more poorly at symptom onset than ADAD participants on a cognitive composite. After symptom onset, however, the younger ADAD MCs declined more rapidly. The similar but not identical cognitive trajectories (declining but at different rates) for ADAD and LOAD suggest common AD pathologies but with some differences., (© 2021 the Alzheimer's Association.)

Introduction: Research addressing Alzheimer disease and related dementias must examine nonbiological factors influencing the risk for and expression of Alzheimer disease and related dementias. These factors address the interplay of cognition with lived experiences and social and structural determinants of health (SSDOH). However, coordinated measures of SSDOH are limited., Methods: The Knight Alzheimer Disease Research Center (ADRC) at Washington University in St. Louis developed and piloted a comprehensive battery to measure SSDOH. One hundred and twelve participants, very mildly cognitively impaired or unimpaired, enrolled in memory studies completed the electronic SSDOH battery. The Clinical Dementia Rating (CDR) determined the presence or absence of cognitive impairment., Results: Four domains demonstrated above acceptable intraclass correlation scores for test-retest reliability (≥0.70), including adverse childhood events, discrimination, social status, and early education. Twenty very mildly impaired participants completed the electronic pilot study., Conclusion: Our findings indicate that participants with early-stage symptomatic Alzheimer disease are able to participate in electronic SSDOH data collection. In collaboration with the University of Pennsylvania ADRC, we replaced/modified certain assessments to increase intraclass correlation. The resulting battery, Social and Structural Life-courses Influencing Aging and Dementia (SS-DIAD), can serve as a SSDOH collection tool and is currently utilized in cognitively impaired and unimpaired research participants at both ADRCs., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Objective: To evaluate for racial differences in triggering receptor expressed on myeloid cells 2 (TREM2), a key immune mediator in Alzheimer disease, the levels of CSF soluble TREM2 (sTREM2), and the frequency of associated genetic variants were compared in groups of individuals who self-reported their race as African American (AA) or non-Hispanic White (NHW)., Methods: Community-dwelling older research participants underwent measurement of CSF sTREM2 concentrations and genetic analyses., Results: The primary cohort included 91 AAs and 868 NHWs. CSF sTREM2 levels were lower in the AA compared with the NHW group (1,336 ± 470 vs 1,856 ± 624 pg/mL, p < 0.0001). AAs were more likely to carry TREM2 coding variants (15% vs 3%, p < 0.0001), which were associated with lower CSF sTREM2. AAs were less likely to carry the rs1582763 minor allele (8% vs 37%, p < 0.0001), located near MS4A4A , which was associated with higher CSF sTREM2. These findings were replicated in an independent cohort of 23 AAs and 917 NHWs: CSF sTREM2 levels were lower in the AA group ( p = 0.03), AAs were more likely to carry coding TREM2 variants (22% vs 4%, p = 0.002), and AAs were less likely to carry the rs1582763 minor allele (16% vs 37%, p = 0.003)., Conclusions: On average, AAs had lower CSF sTREM2 levels compared with NHWs, potentially because AAs are more likely to carry genetic variants associated with lower CSF sTREM2 levels. Importantly, CSF sTREM2 reflects TREM2-mediated microglial activity, a critical step in the immune response to amyloid plaques. These findings suggest that race may be associated with risk for genetic variants that influence Alzheimer disease-related inflammation., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.)

Background: Anticholinergic burden (ACB) is associated with an increased risk of delirium in the older population outside of the acute hospital setting. In acute settings, delirium is associated with increased mortality, and this association is greater with full syndromal delirium (FSD) than with subsyndromal delirium (SSD). Little is known about the impact of ACB on delirium prevalence or subtype in hospitalized older adults or the impact on mortality in this population., Objectives: Our objectives were to determine whether ACB moderates associations between the subtype of delirium experienced by hospitalized older adults and to explore factors (including ACB) that might moderate consequent associations between delirium and mortality in hospital inpatients., Methods: We conducted a retrospective analysis of a cohort of 784 older adults with unplanned admission to a North London acute medical unit between June and December 2007. Univariate regression analyses were performed to explore associations between ACB, as represented by the Anticholinergic Burden Scale (ACBS), delirium subtype (FSD vs. SSD), and mortality., Results: The mean age of the sample was 83 ± standard deviation (SD) 7.4 years, and the majority of patients were female (59%), lived in their own homes (71%), were without dementia (75%), and died between hospital admission and the end of the 2-year follow-up period (59%). Mean length of admission was 13.2 ± 14.4 days. Prescription data revealed an ACBS score of 1 in 26% of the cohort, of 2 in 12%, and of ≥ 3 in 16%. The mean total ACBS score for the cohort was 1.1 ± 1.4 (range 0-9). Patients with high ACB on admission were more likely to have severe dementia, to have multiple comorbidities, and to live in residential care. Higher ACB was not associated with delirium of either subtype in hospitalized older adults. Delirium itself was associated with increased mortality, and greater associations were seen in FSD (hazard ratio [HR] 2.27; 95% confidence interval [CI] 1.70-3.01) than in SSD (HR 1.58; 95% CI 1.2-2.09); however, ACB had no impact on this relationship., Conclusions: ACB was not found to be associated with increased delirium of either subtype or to have a demonstrable impact on mortality in delirium. Prior suggestions of links between ACB and mortality in similar populations may be mediated by higher levels of functional dependence, greater levels of residential home residence, or an increased prevalence of dementia in this population.

In Alzheimer's disease (AD), deposition of insoluble amyloid-β (Aβ) is followed by intracellular aggregation of tau in the neocortex and subsequent neuronal cell loss, synaptic loss, brain atrophy, and cognitive impairment. By the time even the earliest clinical symptoms are detectable, Aβ accumulation is close to reaching its peak and neocortical tau pathology is frequently already present. The period in which AD pathology is accumulating in the absence of cognitive symptoms represents a clinically relevant time window for therapeutic intervention. Sleep is increasingly recognized as a potential marker for AD pathology and future risk of cognitive impairment. Previous studies in animal models and humans have associated decreased non-rapid eye movement (NREM) sleep slow wave activity (SWA) with Aβ deposition. In this study, we analyzed cognitive performance, brain imaging, and cerebrospinal fluid (CSF) AD biomarkers in participants enrolled in longitudinal studies of aging. In addition, we monitored their sleep using a single-channel electroencephalography (EEG) device worn on the forehead. After adjusting for multiple covariates such as age and sex, we found that NREM SWA showed an inverse relationship with AD pathology, particularly tauopathy, and that this association was most evident at the lowest frequencies of NREM SWA. Given that our study participants were predominantly cognitively normal, this suggested that changes in NREM SWA, especially at 1 to 2 Hz, might be able to discriminate tau pathology and cognitive impairment either before or at the earliest stages of symptomatic AD., (Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Introduction: Individuals in early stages of Alzheimer's disease are a targeted population for secondary prevention trials aimed at preserving normal cognition. Understanding within-person biomarker(s) change over time is critical for trial enrollment and design., Methods: Longitudinal cerebrospinal fluid samples from the Alzheimer's Disease Neuroimaging Initiative were assayed for novel markers of neuronal/synaptic injury (visinin-like protein 1, Ng, and SNAP-25) and neuroinflammation (YKL-40) and compared with β amyloid 42, tau, and phospho-tau181. General linear mixed models were used to compare within-person rates of change in three clinical groups (cognitively normal, mild cognitive impairment, and Alzheimer's disease) further defined by β amyloid status., Results: Levels of injury markers were highly positively correlated. Despite elevated baseline levels as a function of clinical status and amyloid-positivity, within-person decreases in these measures were observed in the early symptomatic, amyloid-positive Alzheimer's disease group., Discussion: Knowledge of within-person biomarker change will impact interpretation of biomarker outcomes in clinical trials that are dependent on disease stage., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

APOE4 is the strongest genetic risk factor for late-onset Alzheimer disease. ApoE4 increases brain amyloid-β pathology relative to other ApoE isoforms. However, whether APOE independently influences tau pathology, the other major proteinopathy of Alzheimer disease and other tauopathies, or tau-mediated neurodegeneration, is not clear. By generating P301S tau transgenic mice on either a human ApoE knock-in (KI) or ApoE knockout (KO) background, here we show that P301S/E4 mice have significantly higher tau levels in the brain and a greater extent of somatodendritic tau redistribution by three months of age compared with P301S/E2, P301S/E3, and P301S/EKO mice. By nine months of age, P301S mice with different ApoE genotypes display distinct phosphorylated tau protein (p-tau) staining patterns. P301S/E4 mice develop markedly more brain atrophy and neuroinflammation than P301S/E2 and P301S/E3 mice, whereas P301S/EKO mice are largely protected from these changes. In vitro, E4-expressing microglia exhibit higher innate immune reactivity after lipopolysaccharide treatment. Co-culturing P301S tau-expressing neurons with E4-expressing mixed glia results in a significantly higher level of tumour-necrosis factor-α (TNF-α) secretion and markedly reduced neuronal viability compared with neuron/E2 and neuron/E3 co-cultures. Neurons co-cultured with EKO glia showed the greatest viability with the lowest level of secreted TNF-α. Treatment of P301S neurons with recombinant ApoE (E2, E3, E4) also leads to some neuronal damage and death compared with the absence of ApoE, with ApoE4 exacerbating the effect. In individuals with a sporadic primary tauopathy, the presence of an ε4 allele is associated with more severe regional neurodegeneration. In individuals who are positive for amyloid-β pathology with symptomatic Alzheimer disease who usually have tau pathology, ε4-carriers demonstrate greater rates of disease progression. Our results demonstrate that ApoE affects tau pathogenesis, neuroinflammation, and tau-mediated neurodegeneration independently of amyloid-β pathology. ApoE4 exerts a 'toxic' gain of function whereas the absence of ApoE is protective.

Objective: Deposition of amyloid β (Aβ)-containing plaques as evidenced by amyloid imaging and cerebrospinal fluid (CSF) Aβ1-42 (Aβ42) is an early indicator of preclinical Alzheimer disease (AD). To better understand their relationship during the earliest preclinical stages, we investigated baseline CSF markers in cognitively normal individuals at different stages of amyloid deposition defined by longitudinal amyloid imaging with Pittsburgh compound B (PIB): (1) PIB-negative at baseline and follow-up (PIB(-) ; normal), (2) PIB-negative at baseline but PIB-positive at follow-up (PIB converters; early preclinical AD), and (3) PIB-positive at baseline and follow-up (PIB(+) ; preclinical AD)., Methods: Cognitively normal individuals (n = 164) who had undergone baseline PIB scan and CSF collection within 1 year of each other and at least 1 additional PIB follow-up were included. Amyloid status was defined dichotomously using an a priori mean cortical cutoff., Results: PIB converters (n = 20) at baseline exhibited significantly lower CSF Aβ42 compared to those who remained PIB-negative (n = 123), but higher compared to the PIB(+) group (n = 21). A robust negative correlation (r = -0.879, p = 0.0001) between CSF Aβ42 and absolute (but subthreshold) PIB binding was observed during this early preclinical stage. The negative correlation was not as strong once individuals were PIB-positive (r = -0.456, p = 0.038), and there was no correlation in the stable PIB(-) group (p = 0.905) or in the group (n = 10) with early symptomatic AD (p = 0.537)., Interpretation: CSF Aβ42 levels are tightly coupled with cortical amyloid load in the earliest stages of preclinical AD, and begin to decrease dramatically prior to the point when an abnormal threshold of cortical accumulation is detected with amyloid imaging. Ann Neurol 2016;80:379-387., Competing Interests: Potential conflicts of interests AGV, LM, MSJ, BG, YS, CX, DMH, TLSB, JCM, and AMF have no conflict to report., (© 2016 American Neurological Association.)

Objective: Limited clinical trials have validated the hypothesis of controlling graft-versus-host disease (GVHD) arising from stem cell transplant utilizing suicidal T-lymphocytes that have been transduced to express the HSV-TK gene. However, clinical utility has been limited by diminished T-cell function arising from the production process. To evaluate strategies for harnessing the graft-versus-leukemia (GVL) effect while improving the safety and function of suicidal lymphocytes, we have developed techniques to produce fully functional, retrovirally transduced, HSV-TK-positive murine T cells (TK+TC)., Methods: Utilizing a murine major histocompatibility complex-matched transplant model, we evaluated the ability of TK+TC to generate a GVL effect and the ability to control GVHD in experiments where we varied the dose of TK+TC, ganciclovir (GCV) dose, the start of GCV administration (day 4, 7, 10, 13, 15, or 19) posttransplantation, and the GCV administration route (osmotic pump versus intraperitoneal)., Results: At TK+TC doses in excess of the standard lethal dose (SLD) of unmanipulated T-cells, GCV administration completely (2 x SLD) and partially (4 x SLD) controlled GVHD. Additionally, GVHD remained reversible despite delaying administration of GCV for a week after GVHD developed. Importantly, GVHD was controlled with a 1-log but not 2-log reduction in GCV dose, and this "partial suicide" preserved more circulating TK+TC compared with standard-dose GCV. Survival of leukemia-positive mice receiving TK+TC and GCV was significantly increased compared with control cohorts not receiving GCV or transplanted with unmanipulated T cells, thereby demonstrating a GVL effect., Conclusion: Retrovirally transduced suicidal lymphocytes generate a potent GVL effect while simultaneously enabling control of GVHD, which results in improved leukemia and GVHD-free survival.

To identify sporulation-specific proteins that might serve as targets of developmental regulatory factors in Streptomyces, we examined total proteins of Streptomyces griseus by two-dimensional gel electrophoresis. Among five proteins that were present at high levels during sporulation but absent from vegetative cells, two of the proteins, P3 and P4, were absent from developmental mutants that undergo aberrant morphogenesis. The deduced amino acid sequence of the gene that encodes P3 (EshA) showed extensive similarity to proteins from mycobacteria and a cyanobacterium, Synechococcus, that are abundant during nutritional stress but whose functions are unknown. Uniquely among these proteins, EshA contains a cyclic nucleotide-binding domain, suggesting that the activity of EshA may be modulated by a cyclic nucleotide. The eshA gene was strongly expressed from a single transcription start site only during sporulation, and accumulation of the eshA transcript depended on a developmental gene, bldA. During submerged sporulation, a null mutant strain that produced no EshA could not extend sporogenic hyphae from new branch points but instead accelerated septation and spore maturation at the preexisting vegetative filaments. These results indicated that EshA is required for the growth of sporogenic hyphae and localization of septation and spore maturation but not for spore viability.

Several of the viral proteins required for human cytomegalovirus (HCMV) DNA replication have been difficult to study due to their low abundance in infected cells and low solubility in bacterial or insect-cell expression systems. Therefore we used the Semliki Forest virus expression system to express these proteins in mammalian cells. All of the recombinant proteins were soluble, on the basis of ultracentrifugation properties and their ability to be immunoprecipitated from solution with specific antibodies. Pulse-chase analysis of the 86-kDa major immediate-early protein (IE86) revealed two expressed forms-a precursor and a product-indicating that this recombinant protein, like the native HCMV protein, is posttranslationally processed. The recombinant proteins (polymerase core and accessory as well as the IE86 and pUL84) formed stable complexes similar to those known to form in HCMV-infected cells. The recombinant DNA polymerase holoenzyme also exhibited enzyme activity that was phosphonoformic acid sensitive, as is the infected-cell DNA polymerase activity. This expression system offers many advantages for the expression and study of the HCMV replication proteins, including the expression of soluble, active proteins that are able to interact to form complexes. Additionally, the relative ease with which SFV recombinants can be made lends itself to the construction and evaluation of mutants., (Copyright 1998 Academic Press.)

A Bayesian procedure for the simultaneous alignment and classification of sequences into subclasses is described. This Gibbs sampling algorithm iterates between an alignment step and a classification step. It employs Bayesian inference for the identification of the number of conserved columns, the number of motifs in each class, their size, and the size of the classes. Using Bayesian prediction, inter-class differences in all these variables are brought to bare on the classification. Application to a superfamily of cyclic nucleotide-binding proteins identifies both similarities and differences in the sequence characteristics of the five subclasses identified by the procedure: 1) cNMP-dependent kinases, 2) prokaryotic cAMP-dependent regulatory proteins, CRP-type, 3) prokaryotic regulatory proteins, FNR-type, 4) cAMP gated ion channel proteins of animals, and 5) cAMP gated ion channels of plants.

Although the overall picture of HCMV DNA synthesis appears typical of the herpesviruses, some novel features are emerging. Six herpesvirus-group-common genes encode proteins that likely constitute the replication fork machinery, including a two-subunit DNA polymerase, a helicas-primase complex and a single-stranded DNA-binding protein. No homolog of the herpes simplex virus origin-binding helicase is evident, but at least one additional HCMV protein of unknown function, pUL84, appears to be required for initiation. Replication initiates within or near the large and structurally complex lytic-phase replicator, ori-Lyt, near the center of UL. Recent findings suggest that ori-Lyt-mediated initiation of DNA synthesis occurs through a mechanism distinct from that employed by herpes simplex virus.

Previous studies had indicated that the form II or B cluster of CO2 fixation structural genes is part of a large operon in Rhodobacter sphaeroides (Gibson, J. L., Chen, J.-H., Tower, P. A., and Tabita, F. R. (1990) Biochemistry 29, 8085-8093). In this investigation, we have sequenced the DNA between the prkB and rbpL genes and provide evidence for three distinct open reading frames which encode additional structural genes of the Calvin reductive pentose phosphate pathway; these genes encode the enzymes transketolase, glyceraldehyde phosphate dehydrogenase, and aldolase. Noteworthy is transketolase, which may be expressed to high levels in Escherichia coli. This study thus represents the initial description of the primary structure of bacterial transketolase, a key enzyme of the reductive and the oxidative pentose phosphate pathways. Each of the genes are separated by short stretches of intergenic sequence, consistent with earlier evidence which suggested that these genes are cotranscribed and part of a large operon controlled by sequences upstream from fbpB.