Modifications in the T arm of tRNA globally determine tRNA maturation, function, and cellular fitness.

Almost all elongator tRNAs (Transfer RNAs) harbor 5-methyluridine 54 and pseudouridine 55 in the T arm, generated by the enzymes TrmA and TruB, respectively, in Escherichia coli. TrmA and TruB both act as tRNA chaperones, and strains lacking trmA or truB are outcompeted by wild type. Here, we investigate how TrmA and TruB contribute to cellular fitness. Deletion of trmA and truB in E. coli causes a global decrease in aminoacylation and alters other tRNA modifications such as acp3U47. While overall protein synthesis is not affected in ΔtrmA and ΔtruB strains, the translation of a subset of codons is significantly impaired. As a consequence, we observe translationally reduced expression of many specific proteins, that are either encoded with a high frequency of these codons or that are large proteins. The resulting proteome changes are not related to a specific growth phenotype, but overall cellular fitness is impaired upon deleting trmA and truB in accordance with a general protein synthesis impact. In conclusion, we demonstrate that universal modifications of the tRNA T arm are critical for global tRNA function by enhancing tRNA maturation, tRNA aminoacylation, and translation, thereby improving cellular fitness irrespective of the growth conditions which explains the conservation of trmA and truB.

RNA modifying enzymes shape tRNA biogenesis and function.

Transfer RNAs (tRNAs) are the most highly modified cellular RNAs, both with respect to the proportion of nucleotides that are modified within the tRNA sequence and with respect to the extraordinary diversity in tRNA modification chemistry. However, the functions of many different tRNA modifications are only beginning to emerge. tRNAs have two general clusters of modifications. The first cluster is within the anticodon stem-loop including several modifications essential for protein translation. The second cluster of modifications is within the tRNA elbow, and roles for these modifications are less clear. In general, tRNA elbow modifications are typically not essential for cell growth, but nonetheless several tRNA elbow modifications have been highly conserved throughout all domains of life. In addition to forming modifications, many tRNA modifying enzymes have been demonstrated or hypothesized to additionally play an important role in folding tRNA acting as tRNA chaperones. In this review, we summarize the known functions of tRNA modifying enzymes throughout the lifecycle of a tRNA molecule, from transcription to degradation. Thereby, we describe how tRNA modification and folding by tRNA modifying enzymes enhance tRNA maturation, tRNA aminoacylation, and tRNA function during protein synthesis, ultimately impacting cellular phenotypes and disease.

Fluorescent labeling of tRNA for rapid kinetic interaction studies with tRNA-binding proteins.

Transfer RNA (tRNA) plays a critical role during translation and interacts with numerous proteins during its biogenesis, functional cycle and degradation. In particular, tRNA is extensively post-transcriptionally modified by various tRNA modifying enzymes which each target a specific nucleotide at different positions within tRNAs to introduce different chemical modifications. Fluorescent assays can be used to study the interaction between a protein and tRNA. Moreover, rapid mixing fluorescence stopped-flow assays provide insights into the kinetics of the tRNA-protein interaction in order to elucidate the tRNA binding mechanism for the given protein. A prerequisite for these studies is a fluorescently labeled molecule, such as fluorescent tRNA, wherein a change in fluorescence occurs upon protein binding. In this chapter, we discuss the utilization of tRNA modifications in order to introduce fluorophores at particular positions within tRNAs. Particularly, we focus on in vitro thiolation of a uridine at position 8 within tRNAs using the tRNA modification enzyme ThiI, followed by labeling of the thiol group with fluorescein. As such, this fluorescently labeled tRNA is primarily unmodified, with the exception of the thiolation modification to which the fluorophore is attached, and can be used as a substrate to study the binding of different tRNA-interacting factors. Herein, we discuss the example of studying the tRNA binding mechanism of the tRNA modifying enzymes TrmB and DusA using internally fluorescein-labeled tRNA.

Molecular mechanism of tRNA binding by the Escherichia coli N7 guanosine methyltransferase TrmB.

Among the large and diverse collection of tRNA modifications, 7-methylguanosine (m7G) is frequently found in the tRNA variable loop at position 46. This modification is introduced by the TrmB enzyme, which is conserved in bacteria and eukaryotes. However, the molecular determinants and the mechanism for tRNA recognition by TrmB are not well understood. Complementing the report of various phenotypes for different organisms lacking TrmB homologs, we report here hydrogen peroxide sensitivity for the Escherichia coli ΔtrmB knockout strain. To gain insight into the molecular mechanism of tRNA binding by E. coli TrmB in real time, we developed a new assay based on introducing a 4-thiouridine modification at position 8 of in vitro transcribed tRNAPhe enabling us to fluorescently label this unmodified tRNA. Using rapid kinetic stopped-flow measurements with this fluorescent tRNA, we examined the interaction of WT and single substitution variants of TrmB with tRNA. Our results reveal the role of S-adenosylmethionine for rapid and stable tRNA binding, the rate-limiting nature of m7G46 catalysis for tRNA release, and the importance of residues R26, T127, and R155 across the entire surface of TrmB for tRNA binding.

Multi-wavelength analytical ultracentrifugation of biopolymer mixtures and interactions.

Multi-wavelength analytical ultracentrifugation (MW-AUC) is a recent development made possible by new analytical ultracentrifuge optical systems. MW-AUC extends the basic hydrodynamic information content of AUC and provides access to a wide range of new applications for biopolymer characterization, and is poised to become an essential analytical tool to study macromolecular interactions. It adds an orthogonal spectral dimension to the traditional hydrodynamic characterization by exploiting unique chromophores in analyte mixtures that may or may not interact. Here we illustrate the utility of MW-AUC for experimental investigations where the benefit of the added spectral dimension provides critical information that is not accessible, and impossible to resolve with traditional AUC methods. We demonstrate the improvements in resolution and information content obtained by this technique compared to traditional single- or dual-wavelength approaches, and discuss experimental design considerations and limitations of the method. We further address the advantages and disadvantages of the two MW optical systems available today, and the differences in data analysis strategies between the two systems.

Retest Reliability for Complete Airway Interruption Systems of Aerodynamic Measurement.

INTRODUCTION: Measures of subglottal pressure (Ps), phonation threshold pressure (PTP), and laryngeal resistance (LR) can be used as indicators of vocal cord disorders. The gold standard non-invasive measurement uses labial interruption, which has been shown to have reliability inconsistencies. Mechanical interruption methods have demonstrated promise in measurement reliability. The goal of the present study is to compare retest reliability of labial and mechanical interruption methods. METHODS: 55 subjects aged 18-69 participated. Ten trials were performed for each method. For labial interruption, subjects produced five labial plosives at comfortable and quiet volumes. For mechanical interruption, subjects produced a sustained /α/ while a balloon valve interrupted phonation five times. Thirty subjects completed a second study visit identical to the first approximately two weeks (15 days ± 3.76) after the first visit. Ps, PTP, mean airflow rate, and LR were determined for each subject and retest reliability for each was analyzed. RESULTS: The percent difference in measurement results for test-retest of Ps were 12.88% ± 10.15 for mechanical interruption and 27.56% ± 17.14 for labial interruption (P = 0.0003). The percent difference for PTP measurements were 21.46% ± 16.01 for mechanical and 17.04% ± 14.62 (P = 0.3372) for labial. Intra-subject coefficients of variation of Ps were 0.086 ± 0.046 for mechanical and 0.161 ± 0.078 for labial (P < 0.0001). For PTP, the coefficients were 0.177 ± 0.083 for mechanical and 0.186 ± 0.091 for labial (P = 0.5402). Lastly, for LR (Ps divided by mean airflow rate) the percent differences were 14.33% ± 10.06 for mechanical and 53.87% ± 43.19 for labial (P < 0.0001) with intra-subject variability of 0.115 ± 0.050 for mechanical and 0.287 ± 0.222 for labial (P < 0.0001). CONCLUSIONS: Ps and LR measured using mechanical interruption showed more consistency for both retesting across separate study visits and intra-subject variability. PTP was similar in retesting and intra-subject variability. Continued work to improve mechanical interruption techniques is warranted as these methods offer higher reliability and consistency than the labial interruption methods.

Partially modified tRNAs for the study of tRNA maturation and function.

Transfer RNA (tRNA) is the most highly and diversely modified class of RNA in all domains of life. However, we still have only a limited understanding of the concerted action of the many enzymes that modify tRNA during tRNA maturation and the synergistic functions of tRNA modifications for protein synthesis. Here, we describe the preparation of in vitro transcribed tRNAs with a partial set of defined modifications and the use of partially modified tRNAs in biochemical assays. By comparing the affinity and activity of tRNA modification enzymes for partially modified and unmodified tRNAs, we gain insight into the preferred pathways of tRNA maturation. Additionally, partially modified tRNAs will be highly useful to investigate the importance of tRNA modifications for tRNA function during translation including the interaction with aminoacyl-tRNA synthases, translation factors and the ribosome. Thereby, the methods described here lay the foundation for understanding the mechanistic function of tRNA modifications.

Assaying the Molecular Determinants and Kinetics of RNA Pseudouridylation by H/ACA snoRNPs and Stand-Alone Pseudouridine Synthases.

Posttranscriptional modifications of RNA play an important role in promoting the maturation and functional diversity of many RNA species. Accordingly, understanding the enzymes and mechanisms that underlie RNA modifications is an important aspect in advancing our knowledge of the continually expanding RNA modification field. However, of the more than 160 currently identified RNA modifications, a large portion remains without quantitative detection assays for their biochemical characterization. Here, we describe the tritium release assay as a convenient tool allowing for the quantitative assessment of in vitro RNA pseudouridylation by stand-alone or box H/ACA RNA-guided pseudouridine synthases. This assay enables quantification of RNA pseudouridylation over a time course to effectively compare pseudouridylation activity between different substrates and/or different recombinant enzymes as well as to determine kinetic parameters. With the help of a quench-flow apparatus, the tritium release assay can be adapted for rapid kinetic measurements under single-turnover conditions to dissect reaction mechanisms. As examples, we show the formation of pseudouridines by a reconstituted Saccharomyces cerevisiae H/ACA small ribonucleoprotein (snoRNP) and an Escherichia coli stand-alone pseudouridine synthase.

Identification and characterization of a G-quadruplex structure in the pre-core promoter region of hepatitis B virus covalently closed circular DNA.

Approximately 250 million people worldwide are chronically infected with the hepatitis B virus (HBV) and are at increased risk of developing cirrhosis and hepatocellular carcinoma. The HBV genome persists as covalently closed circular DNA (cccDNA), which serves as the template for all HBV mRNA transcripts. Current nucleos(t)ide analogs used to treat HBV do not directly target the HBV cccDNA genome and thus cannot eradicate HBV infection. Here, we report the discovery of a unique G-quadruplex structure in the pre-core promoter region of the HBV genome that is conserved among nearly all genotypes. This region is central to critical steps in the viral life cycle, including the generation of pregenomic RNA, synthesis of core and polymerase proteins, and genome encapsidation; thus, an increased understanding of the HBV pre-core region may lead to the identification of novel anti-HBV cccDNA targets. We utilized biophysical methods (circular dichroism and small-angle X-ray scattering) to characterize the HBV G-quadruplex and the effect of three distinct G to A mutants. We also used microscale thermophoresis to quantify the binding affinity of G-quadruplex and its mutants with a known quadruplex-binding protein (DHX36). To investigate the physiological relevance of HBV G-quadruplex, we employed assays using DHX36 to pull-down cccDNA and compared HBV infection in HepG2 cells transfected with wild-type and mutant HBV plasmids by monitoring the levels of genomic DNA, pregenomic RNA, and antigens. Further evaluation of this critical host-protein interaction site in the HBV cccDNA genome may facilitate the development of novel anti-HBV therapeutics against the resilient cccDNA template.

tRNA elbow modifications affect the tRNA pseudouridine synthase TruB and the methyltransferase TrmA.

tRNAs constitute the most highly modified class of RNA. Every tRNA contains a unique set of modifications, and Ψ55, m5U54, and m7G46 are frequently found within the elbow of the tRNA structure. Despite the abundance of tRNA modifications, we are only beginning to understand the orchestration of modification enzymes during tRNA maturation. Here, we investigated whether pre-existing modifications impact the binding affinity or catalysis by tRNA elbow modification enzymes. Specifically, we focused on the Escherichia coli enzymes TruB, TrmA, and TrmB which generate Ψ55, m5U54, and m7G46, respectively. tRNAs containing a single modification were prepared, and the binding and activity preferences of purified E. coli TrmA, TruB, and TrmB were examined in vitro. TruB preferentially binds and modifies unmodified tRNA. TrmA prefers to modify unmodified tRNA, but binds most tightly to tRNA that already contains Ψ55. In contrast, binding and modification by TrmB is insensitive to the tRNA modification status. Our results suggest that TrmA and TruB are likely to act on mostly unmodified tRNA precursors during the early stages of tRNA maturation whereas TrmB presumably acts on later tRNA intermediates that are already partially modified. In conclusion, we uncover the mechanistic basis for the preferred modification order in the E. coli tRNA elbow region.

Endometriosis: A Malignant Fingerprint.

BACKGROUND: Endometriosis is complex, but identifying the novel biomarkers, inflammatory molecules, and genetic links holds the key to the enhanced detection, prediction and treatment of both endometriosis and endometriosis related malignant neoplasia. Here we review the literature relating to the specific molecular mechanism(s) mediating tumorigenesis arising within endometriosis. METHODS: Guidance (e.g. Cochrane) and published studies were identified. The Published studies were identified through PubMed using the systematic review methods filter, and the authors' topic knowledge. These data were reviewed to identify key and relevant articles to create a comprehensive review article to explore the molecular fingerprint associated with in endometriosis-driven tumorigenesis. RESULTS: An important focus is the link between C3aR1, PGR, ER1, SOX-17 and other relevant gene expression profiles and endometriosis-driven tumorigenesis. Further studies should also focus on the combined use of CA-125 with HE-4, and the role for OVA1/MIA as clinically relevant diagnostic biomarkers in the prediction of endometriosis-driven tumorigenesis. CONCLUSIONS: Elucidating endometriosis' molecular fingerprint is to understand the molecular mechanisms that drive the endometriosis-associated malignant phenotype. A better understanding of the predictive roles of these genes and the value of the biomarker proteins will allow for the derivation of unique molecular treatment algorithms to better serve our patients.

A randomized controlled trial of online symptom searching to inform patient generated differential diagnoses.

Patient online health searching is now commonplace, however, the accuracy of patient generated differentials for new symptoms and potential for patient anxiety are concerns. We aimed primarily to determine the accuracy of patient generated differentials for new symptoms with and without online searching, and secondarily, to evaluate the impact of searching on anxiety levels. In the waiting room prior to seeing a clinician, 300 patients with new symptoms were randomly assigned 1:1:1 to Google searching with health related features including a symptom search tool vs Google searching with health related features disabled vs no searching. Participants were 18 years or older and presenting to the emergency department of an urban academic medical center with new low-acuity symptoms that were not due to exacerbation of a chronic condition. Search groups received access on a tablet/smartphone to Google searching with or without health related features. Both search groups could access any websites; health related features led the patient to common diagnoses and physician-validated information. The primary outcome was accuracy of the patient generated differential assessed by matching at least two of the top three diagnoses on the clinician's differential. A secondary outcome was anxiety by a visual analogue scale. Patients were a median of 33.1 (IQI 26.2-45.9) years old, 60% women, 63% black, 82% had a high school education or less, and 45.7% reported having performed an online search prior to presentation. Search group patients spent a median of 3.82 (2.53-5.72) minutes searching online. Similar proportions of patients in each group matched at least two of three clinician diagnoses: 27.0% and 28.3% for Google searching with and without health related features vs 23.8% in the no search group. Patients in the search groups had a similar odds of matching ≥2/3 diagnoses as the no search group [OR (95% CI): 1.23 (0.70-2.13), p = 0.47]. Anxiety was unchanged with online searching. In conclusion, brief online searching in the waiting room did not improve accuracy of patient generated differential diagnoses for new symptoms. The absence of an increase in patient anxiety provides reassurance for subsequent work to refine and investigate online symptom search tools.

Molecular Determinants for 23S rRNA Recognition and Modification by the E. coli Pseudouridine Synthase RluE.

The isomerization of uridine to pseudouridine is the most common type of RNA modification found in RNAs across all domains of life and is performed by RNA-dependent and RNA-independent enzymes. The Escherichia coli pseudouridine synthase RluE acts as a stand-alone, highly specific enzyme forming the universally conserved pseudouridine at position 2457, located in helix 89 (H89) of the 23S rRNA in the peptidyltransferase center. Here, we conduct a detailed structure-function analysis to determine the structural elements both in RluE and in 23S rRNA required for RNA-protein interaction and pseudouridine formation. We determined that RluE recognizes a large part of 23S rRNA comprising both H89 and the single-stranded flanking regions which explains the high substrate specificity of RluE. Within RluE, the target RNA is recognized through sequence-specific contacts with loop L7-8 as well as interactions with loop L1-2 and the flexible N-terminal region. We demonstrate that RluE is a faster pseudouridine synthase than other enzymes which likely enables it to act in the early stages of ribosome formation. In summary, our biochemical characterization of RluE provides detailed insight into the molecular mechanism of RluE forming a highly conserved pseudouridine during ribosome biogenesis.

One-year human experience with a novel endoluminal, endoscopic gastric bypass sleeve for morbid obesity.

INTRODUCTION: Here, we report the first series of patients with 1-year implantation of a novel, endoluminal, endoscopically delivered and retrieved gastro-duodeno-jejunal bypass sleeve (GJBS) (ValenTx, Inc. Carpinteria, CA, USA). In this report, we present the safety, feasibility of the device, weight loss, and changes in comorbidities. METHODS AND PROCEDURES: A prospective, single-center, 12-month trial was designed. The patients are morbidly obese individuals who meet the NIH criteria for bariatric surgery. The GJBS is a 120-cm sleeve secured at the esophago-gastric junction with endoscopic and laparoscopic techniques that is designed to create an endoluminal gastro-duodeno-jejunal bypass. The device was implanted and, at the completion of the trial, retrieved with an endoscopic technique. The primary endpoints were safety and incidence of adverse events. The secondary outcomes included the percentage of excess weight loss (EWL) and changes in comorbidities, specifically glucose control, use of antihyperglycemics, and changes in hemoglobin A1C levels. RESULTS: From July 2009 until October 2009, 13 patients were prospectively enrolled for the 1-year trial. The study included five men and eight women with a mean preoperative BMI of 42 kg/m(2). One patient was excluded, at the time of endoscopic evaluation, due to inflammation at the GE junction. Two additional patients required early explantation of the device, within the first 4 weeks, due to patient intolerance. Upon explant of the device, both patients' symptoms improved. In the remaining ten patients, the device was implanted, left in situ for 12 months, and then retrieved endoscopically. Safe delivery of the cuff at the gastro-esophageal junction was seen in all ten patients whom had device implants, without complication. No esophageal leak was seen immediately post-procedure or during follow-up. The sleeve device was well tolerated within the bowel lumen during the 12-month study, specifically, no bowel erosions, ulceration, or pancreatitis was observed. All ten patients reached the 1-year mark. Of the ten, six had fully attached and functional devices throughout the follow-up, verified by endoscopy. The mean percentage EWL, at 1 year, in this group was 54 %. In the remaining four patients, partial cuff detachment was observed at follow-up endoscopy. The percentage EWL was lower in this group. Of the six patients that reached a year with a fully attached device, five were followed at an average of 14-months post-explant (26 months from the time of device implant). These five maintained an average percentage EWL of 30 % at the 14-month post-explant follow-up. Co-morbidites measured included diabetes mellitus, hypertension, hyperlipidemia, and use of antihyperglycemics. Each of the measured comorbidities showed improvement during the 12-month trial. DISCUSSION: The endoluminal, GJBS can be safely placed and retrieved. The short-term data show it is well tolerated with a good safety profile. It achieves excellent weight loss results with over 70 % of all comorbidities resolved or significantly improved.

Aquaporin-2 levels in vitro and in vivo are regulated by VACM-1, a cul 5 gene.

BACKGROUND: In the renal collecting duct, vasopressin regulates water permeability by a process that involves stimulation of adenylyl cyclase activity, cAMP production and subsequent translocation of water channel aquaporin-2 (AQP2) into the apical plasma membrane. We have previously shown that in cos 1 cells in vitro, both adenylyl cyclase activity and cAMP production can be regulated by VACM-1, a cul 5 gene that forms complexes involved in protein ubiquitination and subsequent degradation. METHODS: To extend these observations further, the effects of changes in hydration state on the expression of VACM-1 at the mRNA and the protein level were examined in rats deprived of water (WD) for 24 hrs. RESULTS: In the kidney of WD rats Western blot analyses of kidney tissue showed that the decrease in VACM-1 protein concentration was correlated with the increase in the AQP2 protein level. The immunostaining data suggested that VACM-1/cul5 may be decreased in renal collecting duct but increases in the vasculature of the inner medullary region in response to WD. To determine the possible consequences of the WD dependent decrease in VACM-1/cul5, we next examined the effects of VACM-1 expression on AQP2 protein in vitro. Immunocytochemistry and Western blot analyses data indicate that VACM-1/cul5 expression in MDCK line stably expressing AQP2 gene and in cos 1 cells co-transfected with the AQP2 and VACM-1/cul5 cDNAs decreased AQP2 protein concentration when compared to the vector transfected control groups. CONCLUSION: In summary, our data demonstrate that VACM-1 is involved in the regulation of AQP2 protein concentration and may play a role in regulating water balance.

Cerebral fat embolism without intracardiac shunt: A novel presentation.

Fat embolism syndrome (FES) is defined as an uncommon life-threatening disease process consisting of pulmonary, central nervous system (CNS), and cutaneous manifestations. The pathophysiology of this secondary injury is poorly understood. In the setting of the multiply injured patient, the diagnosis of FES is difficult to ascertain. A case report of a posttraumatic death caused by acute dissemination of diffuse fat emboli to the brain and lungs in the absence of a right-to-left heart defect after femur fracture is presented. The transesophageal echo cardiogram with bubble study failed to demonstrate an intracardiac defect or AV malformation in the lung further supporting a biochemical process. The acute decompensation of the patient within 2 h of the injury would favor mechanical emboli. Supportive care continues to be the mainstay of treatment for FES. Cerebral fat embolism should be considered in traumatically injured patients with unexplained decline in their neurologic examination. Cerebral fat embolism may occur without an intracardiac shunt.

Human experience with an endoluminal, endoscopic, gastrojejunal bypass sleeve.

BACKGROUND: This report describes the authors' experience with a unique endoluminal, endoscopically delivered and retrieved gastroduodenojejunal bypass sleeve, including short-term weight loss and changes in comorbidities. METHODS: A prospective, single-center trial was designed. The patients were morbidly obese individuals who met the National Institutes of Health criteria for bariatric surgery. The device used was a unique gastroduodenojejunal bypass sleeve secured at the esophagogastric junction with endoscopic and laparoscopic techniques and designed to create an endoluminal gastroduodenojejunal bypass. At completion of the trial, the device was explanted with endoscopic retrieval. The primary end points were safety and incidence of adverse events. The secondary outcomes included the percentage of excess weight loss and changes in comorbidities, specifically glucose control, use of antihyperglycemic medications, and changes in hemoglobin A1c levels. RESULTS: From July 2008 to February 2010, 24 patients were enrolled in the trial. The gastroduodenojejunal bypass sleeve was implanted, left in situ, and then retrieved. The 7 men and 17 women in the study had a mean preoperative body mass index of 42 kg/m(2). The device was successfully delivered in 22 of the 24 patients (92%) and retrieved endoscopically from all 22 patients in whom it was implanted (100%). Two patients were excluded from the study preprocedurally. The one patient was excluded preoperatively due to noncompliance with the preoperative liquid diet. For the other excluded patient, the device was not attempted endoscopically due to significant inflammation at the gastroesophageal junction at the time of laparoscopic evaluation. Of the 22 patients who had the device implanted, 17 maintained it (77%) and completed the full 12-week trial. These patients had 39.7% excess weight loss at completion of the study. The primary reason for early explantation of the device was early postoperative dysphagia. The seven patients with preoperative diabetes mellitus all had normal blood glucose levels throughout the trial, and none required antihyperglycemic medications. All four patients with elevated hemoglobin A1c levels preoperatively showed improvement by the end of the trial. CONCLUSIONS: This trial demonstrated that the endoluminal gastroduodenojejunal sleeve can achieve excellent weight loss at 12 weeks. No patient safety issues were encountered. Adverse effects were minimal and resolved at endoscopic device removal. Effective glycemic control was demonstrated through use of the device during the trial. Long-term results are needed.

Which presidents are most memorable?

BACKGROUND/RATIONALE: Episodic memory loss is a hall-mark of Alzheimer's disease (AD), with recall of recent events becoming progressively difficult. A commonly used tool, the recollection of US presidents, was assessed in evaluating episodic versus semantic memory loss among AD patients compared with spouse controls. METHODS: A total of 36 patients (12 men, 24 women) with possible or probable AD were asked to "give the names of 5 US presidents" and concurrently administered the Mini-Mental State Examination (MMSE). Twenty-three spouses (12 men, 11 women) were controls. The year 1980 demarcated "remote" versus "recent" presidents. RESULTS: Patients were older, had lower MMSE scores (P < .001), and recalled fewer presidents than controls (P < .005), after controlling for age. Among patients, men were more educated than women (P < .05) and recalled more presidents (P < .001). No gender differences were observed in controls. CONCLUSIONS: Patients with AD preferentially recalled remote presidents, supporting retention of semantic memory in this group. There were no gender differences between groups.

Bedraggled, a putative transporter, influences the tissue polarity complex during the R3/R4 fate decision in the Drosophila eye.

The tissue polarity pathway is required for the establishment of epithelial polarity in a variety of vertebrate and invertebrate organs. Core tissue polarity proteins act in a dynamically regulated complex to direct the polarization of the Drosophila eye. We report the identification and characterization of bedraggled (bdg), a novel gene that regulates one output of the tissue polarity pathway--the establishment of the R3/R4 photoreceptor fates. bdg encodes a novel, putative transporter protein and interacts genetically with all of the core polarity genes to influence the specification of the R3 and R4 cell fates. Finally, bdg is required for both viability and the initial stages of imaginal disc development.