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Biomedical subjects

Richard E Davis

Publications and source records attributed to Richard E Davis.

At least 19 recordsLinked to original sources
PubMed2006

Enzymatically stable 5' mRNA cap analogs: synthesis and binding studies with human DcpS decapping enzyme.

Four novel 5' mRNA cap analogs have been synthesized with one of the pyrophosphate bridge oxygen atoms of the triphosphate linkage replaced with a methylene group. The analogs were prepared via reaction of nucleoside phosphor/phosphon-1-imidazolidates with nucleoside phosphate/phosphonate in the presence of ZnCl2. Three of the new cap analogs are completely resistant to degradation by human DcpS, the enzyme responsible for hydrolysis of free cap resulting from 3' to 5' cellular mRNA decay. One of the new analogs has very high affinity for binding to human DcpS. Two of these analogs are Anti Reverse Cap Analogs which ensures that they are incorporated into mRNA chains exclusively in the correct orientation. These new cap analogs should be useful in a variety of biochemical studies, in the analysis of the cellular function of decapping enzymes, and as a basis for further development of modified cap analogs as potential anti-cancer and anti-parasite drugs.

Endoribonucleases↗
PubMed2005

The flatworm spliced leader 3'-terminal AUG as a translation initiator methionine.

Spliced leader (SL) RNA trans-splicing contributes the 5' termini to mRNAs in a variety of eukaryotes. In contrast with some transsplicing metazoan groups (e.g. nematodes), flatworm spliced leaders are variable in both sequence and length in different flatworm taxa. However, an absolutely conserved and unique feature of all flatworm spliced leaders is the presence of a 3'-terminal AUG. We previously suggested that the Schistosoma mansoni spliced leader AUG might contribute a required translation initiator methionine to recipient mRNAs. Here we identified and examined trans-spliced cDNAs from a large set of newly available schistosome cDNAs. 28% of the trans-spliced cDNAs have the SL AUG in-frame with the major open reading frame of the mRNA. We identified over 40 cDNAs (40% of the SL AUG in-frame clones) that require the SL AUG as an initiator methionine to synthesize phylogenetically conserved N-terminal residues characteristic of orthologous proteins. RNA transfection experiments using several schistosome stages demonstrated that the flatworm SL AUG can serve as a translation initiator methionine in vivo. We also present in vivo translation studies of the schistosome initiator methionine context and the effect of the spliced leader AUG added upstream and out-of-frame with the main open reading of recipient mRNAs. Overall, our data have provided evidence that another function of flatworm spliced leader trans-splicing is to provide some recipient mRNAs with an initiator methionine for translation initiation.

Amino Acid Sequence↗
PubMed2005

Caenorhabditis elegans decapping proteins: localization and functional analysis of Dcp1, Dcp2, and DcpS during embryogenesis.

Though posttranscriptional regulation is important for early embryogenesis, little is understood regarding control of mRNA decay during development. Previous work defined two major pathways by which normal transcripts are degraded in eukaryotes. However it is not known which pathways are key in mRNA decay during early patterning or whether developmental transcripts are turned over via specific pathways. Here we show that Caenorhabditis elegans Dcp2 is localized to distinct foci during embryogenesis, reminiscent of P-bodies, the sites of mRNA degradation in yeast and mammals. However the decapping enzyme of the 3' to 5' transcript decay system (DcpS) localizes throughout the cytoplasm, suggesting this degradation pathway is not highly organized. In addition we find that Dcp2 is localized to P-granules, showing that Dcp2 is stored and/or active in these structures. However RNAi of these decapping enzymes has no obvious effect on embryogenesis. In contrast we find that nuclear cap binding proteins (CBP-20 and 80), eIF4G, and PAB-1 are absolutely required for development. Together our data provides further evidence that pathways of general mRNA metabolism can be remarkably organized during development, with two different decapping enzymes localized in distinct cytoplasmic domains.

Animals↗
PubMed2005

Synthesis and biochemical properties of novel mRNA 5' cap analogs resistant to enzymatic hydrolysis.

A series of new dinucleotide cap analogs with methylene groups replacing oxygens within the pyrophosphate moieties have been synthesized. All the compounds were resistant to the human scavenger decapping hydrolase, DcpS. Binding constants of the modified caps to eIF4E are comparable to those obtained for m7GpppG. This suggests these methylene modifications in the pyrophosphate chain do not significantly affect cap-binding at least for eIF4E. These cap analogs are also good inhibitors of in vitro translation. mRNAs capped with novel analogs were translated similarly to the mRNA capped with the parent m7GpppG.

Animals↗
PubMed2005

Synthesis and properties of mRNA cap analogs containing phosphorothioate moiety in 5',5'-triphosphate chain.

Nucleosides and oligonucleotides with an oxygen replaced by sulfur atom are an interesting class of compounds because of their improved stability toward enzymatic cleavage by nucleases. We have synthesized several dinucleotide mRNA cap analogs containing a phosphorothioate moiety in the alpha, beta, or gamma position of 5',5'-triphosphate chain [m7Gp(s)ppG, m7Gpp(s)pG, and m7Gppp(s)G]. These are the first examples of the biologically important 5'mRNA cap analogs containing a phosphorothioate moiety, and these compounds may be useful in a variety of biochemical and biotechnological applications. Incorporation of a sulfur atom in the alpha or gamma position within the dinucleotide cap analog was achieved using PSCl3 in a nucleoside phosphorylation reaction followed by coupling the phosphorothioate of nucleoside with a second nucleotide. Synthesis of cap analogs with the phosphorothioate moiety in beta position was performed using an organic phosphorothioate salt in a coupling reaction with an activated nucleotide. The structures of newly synthesized compounds was confirmed using MS and 1H and 31P NMR spectroscopy. We present here the results of preliminary studies on their interaction with translation initiation factor eIF4E and enzymatic hydrolysis with human and nematode DcpS scavengers.

Animals↗
PubMed2005

Nasal valve surgery improves disease-specific quality of life.

OBJECTIVES/HYPOTHESIS: Disease-specific quality-of-life (QOL) assessment of patients with nasal valve compromise and symptomatic nasal obstruction has not been studied previously. The objectives of the study were to determine whether surgical treatment of the nasal valve improves disease-specific QOL and to identify clinical or demographic variables predictive of patients' baseline QOL or change in QOL. STUDY DESIGN: Prospective, multi-institutional outcomes study of 20 patients with nasal obstruction and a surgically treatable diagnosis of nasal valve compromise. METHODS: Disease-specific QOL assessment was performed using the Nasal Obstruction Symptom Evaluation (NOSE) scale preoperatively (n = 20) and at 3 (n = 14) and 6 months (n = 20) after surgery. Clinical and demographic data were collected, along with physician-reported assessments of degree of nasal obstruction. RESULTS: Mean NOSE scores significantly improved from baseline to 3 months after surgery (68.9 vs. 20.7 [P < .0001]), from baseline to 6 months after surgery (68.9 vs. 15.8 [P < .0001]), and from 3 to 6 months after surgery (20.7 vs. 15.8 [P = .0077]). Physician assessment of degree of nasal obstruction using a visual analogue scale was significantly correlated with baseline NOSE scores (P = .013) and change in NOSE scores at 6 months (P = .0015). No other clinical or demographic factors were found to be predictive. CONCLUSION: In patients with symptomatic nasal obstruction and nasal valve compromise, surgical repair of the nasal valve improves disease-specific QOL. Physician rating of degree of nasal obstruction was found to be significantly correlated with patient-reported QOL.

Adult↗
PubMed2005

Dcp2 Decaps m2,2,7GpppN-capped RNAs, and its activity is sequence and context dependent.

Hydrolysis of the mRNA cap plays a pivotal role in initiating and completing mRNA turnover. In nematodes, mRNA metabolism and cap-interacting proteins must deal with two populations of mRNAs, spliced leader trans-spliced mRNAs with a trimethylguanosine cap and non-trans-spliced mRNAs with a monomethylguanosine cap. We describe here the characterization of nematode Dcp1 and Dcp2 proteins. Dcp1 was inactive in vitro on both free cap and capped RNA and did not significantly enhance Dcp2 activity. Nematode Dcp2 is an RNA-decapping protein that does not bind cap and is not inhibited by cap analogs but is effectively inhibited by competing RNA irrespective of RNA sequence and cap. Nematode Dcp2 activity is influenced by both 5' end sequence and its context. The trans-spliced leader sequence on mRNAs reduces Dcp2 activity approximately 10-fold, suggesting that 5'-to-3' turnover of trans-spliced RNAs may be regulated. Nematode Dcp2 decaps both m(7)GpppG- and m(2,2,7)GpppG-capped RNAs. Surprisingly, both budding yeast and human Dcp2 are also active on m(2,2,7)GpppG-capped RNAs. Overall, the data suggest that Dcp2 activity can be influenced by both sequence and context and that Dcp2 may contribute to gene regulation in multiple RNA pathways, including monomethyl- and trimethylguanosine-capped RNAs.

Animals↗
PubMed2004

Coherent differential absorption lidar measurements of CO2.

A differential absorption lidar has been built to measure CO2 concentration in the atmosphere. The transmitter is a pulsed single-frequency Ho:Tm:YLF laser at a 2.05-microm wavelength. A coherent heterodyne receiver was used to achieve sensitive detection, with the additional capability for wind profiling by a Doppler technique. Signal processing includes an algorithm for power measurement of a heterodyne signal. Results show a precision of the CO2 concentration measurement of 1%-2% 1sigma standard deviation over column lengths ranging from 1.2 to 2.8 km by an average of 1000 pulse pairs. A preliminary assessment of instrument sensitivity was made with an 8-h-long measurement set, along with correlative measurements with an in situ sensor, to determine that a CO2 trend could be detected.

Journal Article↗
PubMed2004

Contribution of trans-splicing, 5' -leader length, cap-poly(A) synergism, and initiation factors to nematode translation in an Ascaris suum embryo cell-free system.

Trans-splicing introduces a common 5' 22-nucleotide sequence with an N-2,2,7-trimethylguanosine cap (m (2,2,7)(3)GpppG or TMG-cap) to more than 70% of transcripts in the nematodes Caenorhabditis elegans and Ascaris suum. Using an Ascaris embryo cell-free translation system, we found that the TMG-cap and spliced leader sequence synergistically collaborate to promote efficient translation, whereas addition of either a TMG-cap or spliced leader sequence alone decreased reporter activity. We cloned an A. suum embryo eIF4E homolog and demonstrate that this recombinant protein can bind m(7)G- and TMG-capped mRNAs in cross-linking assays and that binding is enhanced by eIF4G. Both the cap structure and the spliced leader (SL) sequence affect levels of A. suum eIF4E cross-linking to mRNA. Furthermore, the differential binding of eIF4E to a TMG-cap and to trans-spliced and non-trans-spliced RNAs is commensurate with the translational activity of reporter RNAs observed in the cell-free extract. Together, these binding data and translation assays with competitor cap analogs suggest that A. suum eIF4E-3 activity may be sufficient to mediate translation of both trans-spliced and non-trans-spliced mRNAs. Bioinformatic analyses demonstrate the SL sequence tends to trans-splice close to the start codon in a diversity of nematodes. This evolutionary conservation is functionally reflected in the optimal SL to AUG distance for reporter mRNA translation in the cell-free system. Therefore, trans-splicing of the SL1 leader sequence may serve at least two functions in nematodes, generation of an optimal 5'-untranslated region length and a specific sequence context (SL1) for optimal translation of trimethylguanosine capped transcripts.

5' Untranslated Regions↗
PubMed2004

Technical factors in the creation of a "floppy" Nissen fundoplication.

The original description of "floppy" Nissen fundoplication was that of an open procedure involving a large esophageal bougie, complete fundic mobilization, ligation of the short gastric vessels, and placement of a finger or a dilator under the completed fundoplication to assure laxity. A maneuver equivalent to the latter step during laparoscopic Nissen fundoplication has not been described. The consensus in the literature appears to be that the element of a fundoplication that defines it as floppy is complete mobilization of the fundus. We report an intraoperative maneuver that assures the fundoplication has adequate laxity. We also discuss several other maneuvers to assure proper formation of the fundoplication and to minimize side effects such as long-term dysphagia.

Fundoplication↗
PubMed2004

Nematode m7GpppG and m3(2,2,7)GpppG decapping: activities in Ascaris embryos and characterization of C. elegans scavenger DcpS.

A spliced leader contributes the mature 5'ends of many mRNAs in trans-splicing organisms. Trans-spliced metazoan mRNAs acquire an m3(2,2,7)GpppN cap from the added spliced leader exon. The presence of these caps, along with the typical m7GpppN cap on non-trans-spliced mRNAs, requires that cellular mRNA cap-binding proteins and mRNA metabolism deal with different cap structures. We have developed and used an in vitro system to examine mRNA degradation and decapping activities in nematode embryo extracts. The predominant pathway of mRNA decay is a 3' to 5' pathway with exoribonuclease degradation of the RNA followed by hydrolysis of resulting mRNA cap by a scavenger (DcpS-like) decapping activity. Direct decapping of mRNA by a Dcp1/Dcp2-like activity does occur, but is approximately 15-fold less active than the 3' to 5' pathway. The DcpS-like activity in nematode embryo extracts hydrolyzes both m7GpppG and m3(2,2,7)GpppG dinucleoside triphosphates. The Dcp1/Dcp2-like activity in extracts also hydrolyzes these two cap structures at the 5' ends of RNAs. Interestingly, recombinant nematode DcpS differs from its human ortholog in its substrate length requirement and in its capacity to hydrolyze m3(2,2,7)GpppG.

Animals↗
PubMed2003

Is analysis of lower esophageal sphincter vector volumes of value in diagnosing gastroesophageal reflux disease?

AIM: With successful surgical treatment of gastroesophageal reflux disease (GERD), there is interest in understanding the anti-reflux barrier and its mechanisms of failure. To date, the potential use of vector volumes to predict the DeMeester score has not been adequately explored. METHODS: 627 patients in the referral database received esophageal manometry and ambulatory 24-hour pH monitoring. Study data included LES resting pressure (LESP), overall LES length (OL) and abdominal length (AL), total vector volume (TVV) and intrabdominal vector volume (IVV). RESULTS: In cases where LESP, TVV or IVV were all below normal, there was an 81.4 % probability of a positive DeMeester score. In cases where all three were normal, there was an 86.9 % probability that the DeMeester score would be negative. Receiver-operating characteristics (ROC) for LESP, TVV and IVV were nearly identical and indicated no useful cut-off values. Logistic regression demonstrated that LESP and IVV had the strongest association with a positive DeMeester score; however, the regression formula was only 76.1 % accurate. CONCLUSION: While the indices based on TVV, IVV and LESP are more sensitive and specific, respectively, than any single measurement, the measurement of vector volumes does not add significantly to the diagnosis of GERD.

Adolescent↗
PubMed2003

Postmyotomy dysphagia after laparoscopic surgery for achalasia.

AIM: To determine predictive factors for postoperative dysphagia after laparoscopic myotomy for achalasia. METHODS: Logistic regression was used to investigate the possible association between the response (postoperative dysphagia, with two levels: none/mild and moderate/severe) and several plausible predictive factors. RESULTS: Eight patients experienced severe or moderate postoperative dysphagia. The logistic regression revealed that only the severity of preoperative dysphagia (with four levels: mild, moderate, severe, and liquid) was a marginally significant (P=0.0575) predictive factor for postoperative dysphagia. CONCLUSION: The severity of postoperative dysphagia is strongly associated with preoperative dysphagia. Preoperative symptomatology can significantly impact patient outcome.

Deglutition Disorders↗
PubMed2002

Assessment of the Wiener-Retinex process.

We assess the performance of the Retinex transformation, the Wiener restoration, and the combined Wiener-Retinex process in the context of the end-to-end model of color visual-communication channels for near-surface acquisition. We extend the formulations of panchromatic imaging for the nonlinear Retinex transformation and the linear Wiener and Wiener-Retinex restoration filters to color imaging. We further assess the accuracy by which colors are restored in the cases of uniform and spatially varying irradiance levels, and the shadow depth for which color consistency is still possible with reasonable accuracy. Finally, we assess the sensitivity of these processes to perturbations in the visual channel, including the image-gathering design and the spatial support of the Retinex transformation.

Journal Article↗