A possible epigenetic mechanism for the persistence of memory.
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Animals↗
Biomedical subjects
K Si
Publications and source records attributed to K Si.
The Saccharomyces cerevisiae TIF6 gene encoding translation initiation factor 6 is required for 60S ribosomal subunit biogenesis.
Eukaryotic translation initiation factor 6 (eIF6), a monomeric protein of about 26 kDa, can bind to the 60S ribosomal subunit and prevent its association with the 40S ribosomal subunit. In Saccharomyces cerevisiae, eIF6 is encoded by a single-copy essential gene. To understand the function of eIF6 in yeast cells, we constructed a conditional mutant haploid yeast strain in which a functional but a rapidly degradable form of eIF6 fusion protein was synthesized from a repressible GAL10 promoter. Depletion of eIF6 from yeast cells resulted in a selective reduction in the level of 60S ribosomal subunits, causing a stoichiometric imbalance in 60S-to-40S subunit ratio and inhibition of the rate of in vivo protein synthesis. Further analysis indicated that eIF6 is not required for the stability of 60S ribosomal subunits. Rather, eIF6-depleted cells showed defective pre-rRNA processing, resulting in accumulation of 35S pre-rRNA precursor, formation of a 23S aberrant pre-rRNA, decreased 20S pre-rRNA levels, and accumulation of 27SB pre-rRNA. The defect in the processing of 27S pre-rRNA resulted in the reduced formation of mature 25S and 5.8S rRNAs relative to 18S rRNA, which may account for the selective deficit of 60S ribosomal subunits in these cells. Cell fractionation as well as indirect immunofluorescence studies showed that c-Myc or hemagglutinin epitope-tagged eIF6 was distributed throughout the cytoplasm and the nuclei of yeast cells.
Blotting, Western↗
Stretch of mammalian nerve in vitro: effect on compound action potentials.
Stretch of nerve has been reported to decrease the amplitude of the compound action potential (CAP) with a complete block appearing in approximately 30 minutes. But for the most part, those experiments were carried out in vivo, and it is generally accepted that the failure of responses was due to a closure of vessels supplying the nerve with a resulting ischemia and anoxia. These studies were undertaken to determine if stretch of nerve has effects that are independent of interference with its vascular supply. In the studies, lengths of rat sciatic and dog peroneal nerves were removed and placed in a chamber supplied with oxygen in which their CAPs were continuously elicited and recorded. This in vitro preparation obviated interference with the nerve's metabolism on stretching. We have previously shown that the form change termed 'beading,' appearing within 10 seconds and reversing as quickly on relaxation, can be elicited with tensions of only several grams. We wished to determine if stretch adequate to produce beading could alter CAPs with the same rapidity. Tensions below 2 g had little effect. On applying tensions of 10-100 g, levels well above those needed to bead the fibers, both increases and decreases of CAP amplitude were seen. The changes occurred within 10 seconds of stretch application, the time at which beading arises with stretch. Although the decreases of CAP amplitudes could be accounted for by beading, the degree of CAP change did not correspond to the amount of tension applied. We hypothesize that the constrictions in the beaded fibers increase axial resistivity and diminish local currents so as to block conduction. The lack of an increasing degree of decreased CAP amplitude with increases in tension is ascribed to the inhibition of elongation offered by the collagen fibrils present in nerve. Collagenase applied to nerves allowed a further increase in length, producing a 'hyperbeading,' showing much longer lengths of beading constrictions on stretch. This would further increase axial resistance and is taken to account for the greater decreases of CAP amplitudes seen following collagenase treatment. To account for those cases where increases of CAP amplitude were seen on stretch, we hypothesize that stretch can also cause an increase in the excitability of the nodes. The outcome of stretch in any given nerve would be the resultant of two opposing actions; beading of the internodes causes a decrease of local currents leading to block of CAPs, while an increased excitability of the nodes acts to augment the responses.
Action Potentials↗
Initial characterization of the effects of Aloe vera at a crayfish neuromuscular junction.
This study examines the effects of Aloe vera on neurotransmission processes in a well-established invertebrate neuromuscular junction preparation. We studied concentration-response relationships of an Aloe vera extract on excitatory junctional potentials (EJPs) at the opener muscle of the dactyl in the first and second walking limbs of crayfish (Procambarus clarkii and simulans). We observed concentration-dependent depolarizations of the muscle fibre membrane resting potential, depression of EJP amplitudes and an increase in latency to onset of the EJP following electrical stimulation of the isolated excitatory axon in the meropodite. These effects occurred with Aloe concentrations within the 1%-10% (wt-vol) range. Effects of lower concentrations, ranging to a minimum of 0.01% were equivocal. The effects of Aloe were at least partially, and in a majority of cases totally, reversible. EJPs reduced by Aloe could be restored by increasing the nerve stimulation amplitude. This, along with the latency increase, suggests a depression of action potential generation and conduction. The results provide a preliminary characterization of the effects of Aloe vera on the neurotransmission process and suggest that these effects may at least partially account for Aloe's analgesic and antiinflammatory effects. This study shows that the crayfish NMJ preparation should be useful for further elucidating the location(s) and mechanism(s) of action of Aloe on the nervous system.
Aloe↗
The Saccharomyces cerevisiae homologue of mammalian translation initiation factor 6 does not function as a translation initiation factor.
Eukaryotic translation initiation factor 6 (eIF6) binds to the 60S ribosomal subunit and prevents its association with the 40S ribosomal subunit. The Saccharomyces cerevisiae gene that encodes the 245-amino-acid eIF6 (calculated Mr 25,550), designated TIF6, has been cloned and expressed in Escherichia coli. The purified recombinant protein prevents association between 40S and 60S ribosomal subunits to form 80S ribosomes. TIF6 is a single-copy gene that maps on chromosome XVI and is essential for cell growth. eIF6 expressed in yeast cells associates with free 60S ribosomal subunits but not with 80S monosomes or polysomal ribosomes, indicating that it is not a ribosomal protein. Depletion of eIF6 from yeast cells resulted in a decrease in the rate of protein synthesis, accumulation of half-mer polyribosomes, reduced levels of 60S ribosomal subunits resulting in the stoichiometric imbalance in the 40S/60S subunit ratio, and ultimately cessation of cell growth. Furthermore, lysates of yeast cells depleted of eIF6 remained active in translation of mRNAs in vitro. These results indicate that eIF6 does not act as a true translation initiation factor. Rather, the protein may be involved in the biogenesis and/or stability of 60S ribosomal subunits.
Base Sequence↗
[Rapid microdose 14C-urea breath test for detection of Helicobacter pylori infection].
The purpose of this study was to develop and evaluate a rapid microdose 14C-urea breath test (14C-UBT) with a simplified protocol for detecting the infection of hilicobacter pylori (HP). 157 fasting patients who underwent endoscopy with histological examination and rapid urease test (RUT) were given a drink of 37 kBq of 14C-urea. Samples of breath carbon dioxide (1 mmol) were collected at baseline and 10, 20 and 30 min after administration by trapping in hyamine solution. 14C activity was measured by liquid scintillation counting. Results were expressed as cpm. Histolal examination and RUT were used as gold standard for the detection of HP infection. The cutoff value was selected as 200 cpm at 10 min. The results showed that the sensitivity, specificity, positive predictive value, negative predictive value and accuracy were 94.79%, 90.16%, 93.81%, 91.60% and 92.99% respectively. In this study, a 10 min, single sample, 37 kBq 14C-urea breath test for detection HP was developed. The test has good diagnostic accuracy with minimal radiation exposure and low cost. Thus, the authors considered the test to be reliable, safe, convenient and cost-effective to clinical use.
Adolescent↗
Molecular cloning and functional expression of a human cDNA encoding translation initiation factor 6.
Eukaryotic translation initiation factor 6 (eIF6) binds to the 60S ribosomal subunit and prevents its association with the 40S ribosomal subunit. In this paper, we devised a procedure for purifying eIF6 from rabbit reticulocyte lysates and immunochemically characterized the protein by using antibodies isolated from egg yolks of laying hens immunized with rabbit eIF6. By using these monospecific antibodies, a 1.096-kb human cDNA that encodes an eIF6 of 245 amino acids (calculated Mr 26,558) has been cloned and expressed in Escherichia coli. The purified recombinant human protein exhibits biochemical properties that are similar to eIF6 isolated from mammalian cell extracts. Database searches identified amino acid sequences from Saccharomyces cerevisiae, Drosophila, and the nematode Caenorhabditis elegans with significant identity to the deduced amino acid sequence of human eIF6, suggesting the presence of homologues of human eIF6 in these organisms.
Amino Acid Sequence↗
Function of eukaryotic translation initiation factor 1A (eIF1A) (formerly called eIF-4C) in initiation of protein synthesis.
We have used an efficient in vitro translation initiation system to show that the mammalian 17-kDa eukaryotic initiation factor, eIF1A (formerly designated eIF-4C), is essential for transfer of the initiator Met-tRNAf (as Met-tRNAf.eIF2.GTP ternary complex) to 40 S ribosomal subunits in the absence of mRNA to form the 40 S preinitiation complex (40 S.Met-tRNAf.eIF2.GTP). Furthermore, eIF1A acted catalytically in this reaction to mediate highly efficient transfer of the Met-tRNAf.eIF2.GTP ternary complex to 40 S ribosomal subunits. The 40 S complex formed was free of eIF1A indicating that its role in 40 S preinitiation complex formation is not to stabilize the binding of Met-tRNAf to 40 S ribosomes. Additionally, the eIF1A-mediated 40 S initiation complex formed in the presence of AUG codon efficiently joined 60 S ribosomal subunits in an eIF5-dependent reaction to form a functional 80 S initiation complex. In contrast to other reports, we found that eIF1A plays no role either in the subunit joining reaction or in the generation of ribosomal subunits from 80 S ribosomes. Our results indicate that the major function of eIF1A is to mediate the transfer of Met-tRNAf to 40 S ribosomal subunits to form the 40 S preinitiation complex.
Animals↗
Characterization of multiple mRNAs that encode mammalian translation initiation factor 5 (eIF-5).
Eukaryotic translation initiation factor 5 (eIF-5) interacts with the 40 S initiation complex (40S.mRNA.MettRNAf.eIF-2.GTP) to promote the hydrolysis of bound GTP with the concomitant joining of the 60 S ribosomal subunit to the 40 S initiation complex to form a functional 80 S initiation complex. In this paper, the multiple mRNAs that encode mammalian eIF-5 have been characterized. In rat tissues, three major eIF-5 mRNAs of 3.5, 2.8, and 2.2 kilobases in length are detected. All major eIF-5 mRNAs are initiated from a single transcription initiation site, contain identical 5'-untranslated and coding regions, but differ from one another only in the length of their 3'-untranslated regions. The different lengths of the 3'-untranslated region of eIF-5 mRNAs are generated by the use of alternative polyadenylation signals. Additionally, we demonstrate tissue-specific variations in eIF-5 mRNA expression as well as preference for polyadenylation sites. These results should lead to increased understanding of the regulation of eIF-5 gene expression.
Alternative Splicing↗
Characterization of Vibrio cholerae EIT or typing phage D10.
The Vibrio cholerae EITor typing phage D10 was characterized. The adsorption kinetics of the phage on V. cholerae MAK757 strain were biphasic in nature. Intracellular growth was characterized by an eclipse period, latent period and burst size which were 20 min, 25 min and 80 particles per cell respectively. The phage yield was dependent on the concentration and time of addition of DNA synthesis inhibitors such as nalidixic acid and novobiocin, and RNA synthesis inhibitors such as rifampicin. The 32+/-0.2 kb linear double-stranded DNA molecule has unique termini. A restriction map of the phage DNA was constructed with the enzymes BamHI, HindIII and PstI.
Adsorption↗