Stringency of antisense regulation varies based on volatility of mRNA target region

Bacteria can regulate gene expression by transcribing antisense RNA to interfere with protein translation. Antisense has been shown to control a wide variety of prokaryotic proteins, including membrane proteins, protein toxins, and proteins involved in transport and metabolism. This type of regulation can be used in the production of biologics to optimize the health of the culture and maximize production of the desired product. We examined naturally occurring antisense to enhance design principles for product optimization. We found that the level of secondary structure fluctuation of the antisense binding site varied depending on the function of the target. We hypothesized that stringency of regulation by naturally evolved antisense was driven by the impact of the target molecule on cellular survival. Specifically, high stringency was important for toxin-antitoxin systems where survival depended on high levels of control. Toxin-antitoxin systems rely on effective antisense to prevent the translation of self-damaging proteins. Antisense-based systems regulating transport and metabolism potentially benefited from less stringent antisense control. Basal levels of antisense-regulated proteins involved in metabolic processes could allow for quick adaptation to changing nutrient conditions. More than fifty naturally occurring sense/antisense pairs were analyzed to demonstrate that antisense binding sites correlate to the level of stringency needed in regulating the target protein. Please click Additional Files below to see the full abstract

Production of Secretory and Extracellular N-Linked Glycoproteins in Escherichia coli▿ †

The Campylobacter jejuni pgl gene cluster encodes a complete N-linked protein glycosylation pathway that can be functionally transferred into Escherichia coli. In this system, we analyzed the interplay between N-linked glycosylation, membrane translocation and folding of acceptor proteins in bacteria. We developed a recombinant N-glycan acceptor peptide tag that permits N-linked glycosylation of diverse recombinant proteins expressed in the periplasm of glycosylation-competent E. coli cells. With this “glycosylation tag,” a clear difference was observed in the glycosylation patterns found on periplasmic proteins depending on their mode of inner membrane translocation (i.e., Sec, signal recognition particle [SRP], or twin-arginine translocation [Tat] export), indicating that the mode of protein export can influence N-glycosylation efficiency. We also established that engineered substrate proteins targeted to environments beyond the periplasm, such as the outer membrane, the membrane vesicles, and the extracellular medium, could serve as substrates for N-linked glycosylation. Taken together, our results demonstrate that the C. jejuni N-glycosylation machinery is compatible with distinct secretory mechanisms in E. coli, effectively expanding the N-linked glycome of recombinant E. coli. Moreover, this simple glycosylation tag strategy expands the glycoengineering toolbox and opens the door to bacterial synthesis of a wide array of recombinant glycoprotein conjugates

Psychiatric Advance Directives and Reduction of Coercive Crisis Interventions

Background: Psychiatric advance directives are intended to enable self-determined treatment for patients who lose decisional capacity, and thus reduce the need for coercive interventions such as police transport, involuntary commitment, seclusion and restraints, and involuntary medications during mental health crises; whether PADs can help prevent the use of these interventions in practice is unknown. Aims: This study examined whether completion of a Facilitated Psychiatric Advance Directive (F-PAD) was associated with reduced frequency of coercive crisis interventions. Method: The study prospectively compared a sample of PAD completers (n = 147) to non-completers (n = 92) on the frequency of any coercive interventions, with follow-up assessments at 6, 12, and 24 months. Repeated-measures multiple regression analysis was used to estimate the effect of PADs. Models controlled for relevant covariates including a propensity score for initial selection to PADs, baseline history of coercive interventions, concurrent global functioning and crisis episodes with decisional incapacity. Results: F-PAD completion was associated with lower odds of coercive interventions (adjusted OR = 0.50; 95% CI = 0.26–0.96; p \u3c 0.05). Conclusions: PADs may be an effective tool for reducing coercive interventions around incapacitating mental health crises. Less coercion should lead to greater autonomy and self-determination for people with severe mental illness

Serotonin transporter 5HTTLPR polymorphism and affective disorders: no evidence of association in a large European multicenter study.

The available data from preclinical and pharmacological studies on the role of the serotonin transporter (5-HTT) support the hypothesis that a dysfunction in brain serotonergic system activity contributes to the vulnerability to affective disorders (AD). 5-HTT is the major site of serotonin reuptake into the presynaptic neuron, and it has been shown that the polymorphic repeat polymorphism in the 5-HTT promotor region (5-HTTLPR) may affect gene-transcription activity. 5-HTT maps to chromosome 17 at position 17q11.17-q12, and the 5-HTTLPR polymorphisms have been extensively investigated in AD with conflicting results. The present study tested the genetic contribution of the 5-HTTLPR polymorphism in a large European multicenter case-control sample, including 539 unipolar (UPAD), 572 bipolar patients (BPAD), and 821 controls (C). Our European collaboration has led to efforts to optimize a methodology that attenuates some of the major limitations of the case-control association approach. No association was found with primary psychiatric diagnosis (UPAD and BPAD) and with phenotypic traits (family history of AD, suicidal attempt, and presence of psychotic features). Our negative findings are not attributable to the lack of statistical power, and may contribute to clarify the role of 5-HTTLPR polymorphism in AD.Comparative StudyJournal ArticleMulticenter StudyResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

Genome Scan Meta-Analysis of Schizophrenia and Bipolar Disorder, Part III: Bipolar Disorder

Genome scans of bipolar disorder (BPD) have not produced consistent evidence for linkage. The rank-based genome scan meta-analysis (GSMA) method was applied to 18 BPD genome scan data sets in an effort to identify regions with significant support for linkage in the combined data. The two primary analyses considered available linkage data for “very narrow” (i.e., BP-I and schizoaffective disorder–BP) and “narrow” (i.e., adding BP-II disorder) disease models, with the ranks weighted for sample size. A “broad” model (i.e., adding recurrent major depression) and unweighted analyses were also performed. No region achieved genomewide statistical significance by several simulation-based criteria. The most significant P values (<.01) were observed on chromosomes 9p22.3-21.1 (very narrow), 10q11.21-22.1 (very narrow), and 14q24.1-32.12 (narrow). Nominally significant P values were observed in adjacent bins on chromosomes 9p and 18p-q, across all three disease models on chromosomes 14q and 18p-q, and across two models on chromosome 8q. Relatively few BPD pedigrees have been studied under narrow disease models relative to the schizophrenia GSMA data set, which produced more significant results. There was no overlap of the highest-ranked regions for the two disorders. The present results for the very narrow model are promising but suggest that more and larger data sets are needed. Alternatively, linkage might be detected in certain populations or subsets of pedigrees. The narrow and broad data sets had considerable power, according to simulation studies, but did not produce more highly significant evidence for linkage. We note that meta-analysis can sometimes provide support for linkage but cannot disprove linkage in any candidate region