Strains of Escherichia coli have been produced which express very high levels of the tRNAleu1 isoacceptor. This was accomplished by transforming cells with plasmids containing the leuV operon which encodes three copies of the tRNALeu1 gene. Most transformants grew very slowly and exhibited a 15-fold increase in cellular concentrations of tRNALeu1 As a result, total cellular tRNA concentration was approximately doubled and 56% of the total was tRNALeu1. We examined a number of parameters which might be expected to be affected by imbalances in tRNA concentration: in vivo tRNA charging levels, misreading, ribosome step time, and tRNA modification. Surprisingly, no increase in intracellular ppGpp levels was detected even though only about 40% of total leucyl tRNA was found to be charged in vivo. Gross ribosomal misreading was not detected, and it was shown that ribosomal step times were reduced between two- and threefold. Analyses of leucyl tRNA isolated from these slow-growing strains showed that at least 90% of the detectable tRNALeu1 was hypomodified as judged by altered mobility on RPC-5 reverse-phase columns, and by specific modification assays using tRNA(m1G)-methyltransferase and pseudo-uridylate synthetase. Analysis of fast-growing revertants demonstrated that tRNA concentration per se may not explain growth inhibition because selected revertants which grew at wild-type growth rates displayed levels of tRNA comparable to that of control strains bearing the leuV operon. A synthetic tRNALeu1 operon under the control of the T7 promoter was prepared which, when induced, produced six- to sevenfold increases in tRNALeu1 levels. This level of tRNALeu1 titrated the modification system as judged by RPC-5 column chromatography. Overall, our results suggest that hypomodified tRNA may explain, in part, the observed effects on growth, and that the protein-synthesizing system can tolerate an enormous increase in the concentration of a single tRNA. 相似文献
SUP35is an omnipotent suppressor gene of Saccharomyces cerevisiae coding for a protein consisting of a C-terminal part similar to the elongation factor EF-1α and a unique N-terminal sequence of 253 amino acids. Twelve truncated versions of the SUP35 gene were generated by the deletion of fragments internal to the coding sequence. Functional studies of these deletion mutants showed that: (i) only the EF-1α-like C-terminal part of the Sup35 protein is essential for the cell viability; (ii) overexpression of either the N-terminal part of the Sup35 protein or the full-length Sup35 protein decreases translational fidelity, resulting in omnipotent suppression and reduced growth of [psi+] strains; (iii) expression of the C-terminal part of the Sup35 protein generates an antisuppressor phenotype; and (iv) both the N- or C-terminal segments of the Sup35 protein can bind to 80S ribosomes. Thus, the data obtained define two domains within the Sup35 protein which are responsible for different functions. 相似文献
To identify Rhodobacter capsulatus nif genes necessary for the alternative nitrogenase, strains carrying defined mutations in 32 genes and open reading frames of nif region A, B or C were constructed. The ability of these mutants to grow on nitrogen-free medium with molybdenum (Nif phenotype) or in a nifHDK deletion background on medium without molybdenum (Anf phenotype) was tested. Nine nif genes and nif-associated coding regions are absolutely essential for the alternative nitrogenase. These genes comprise nifV and nifB, the nif-specific ntr system (nifR1, R2, R4) and four open reading frames, which exhibit no homology to known genes. In addition, a significantly reduced activity of both the alternative nitrogenase and the molybdenum-dependent nitrogenase was found for fdxN mutants. By random Tn5 mutagenesis of a nifHDK deletion strain 42 Anf? mutants were isolated. Southern hybridization experiments demonstrated that 17 of these Tn5 mutants were localized in at least 13 different restriction fragments outside of known nif regions. Ten different Anf? Tn5 mutations are clustered on a 6 kb DNA fragment of the chromosome designated anf region A. DNA sequence analysis revealed that this region contained the structural genes of the alternative nitrogenase (anfHDGK). The identification of several Tn5 insertions mapping outside of anf region A indicated that at least 10 genes specific for the alternative nitrogenase are present in R. capsulatus.相似文献
Saxifraga section Saxifraga subsection Arachnoideae is a lineage of 12 species distributed mainly in the European Alps. It is unusual in terms of ecological diversification by containing both high elevation species from exposed alpine habitats and low elevation species from shady habitats such as overhanging rocks and cave entrances. Our aims are to explore which of these habitat types is ancestral, and to identify the possible drivers of this remarkable ecological diversification. Using a Hybseq DNA-sequencing approach and a complete species sample we reconstructed and dated the phylogeny of subsection Arachnoideae. Using Landolt indicator values, this phylogenetic tree was used for the reconstruction of the evolution of temperature, light and soil pH requirements in this lineage. Diversification of subsection Arachnoideae started in the late Pliocene and continued through the Pleistocene. Both diversification among and within clades was largely allopatric, and species from shady habitats with low light requirements are distributed in well-known refugia. We hypothesize that low light requirements evolved when species persisting in cold-stage refugia were forced into marginal habitats by more competitive warm-stage vegetation. While we do not claim that such competition resulted in speciation, it very likely resulted in adaptive evolution. 相似文献
The course of diseases such as hypertension, systolic heart failure and heart failure with a preserved ejection fraction is affected by interactions between the left ventricle (LV) and the vasculature. To study these interactions, a computationally efficient, biophysically based mathematical model for the circulatory system is presented. In a four-chamber model of the heart, the LV is represented by a previously described low-order, wall volume-preserving model that includes torsion and base-to-apex and circumferential wall shortening and lengthening, and the other chambers are represented using spherical geometries. Active and passive myocardial mechanics of all four chambers are included. The cardiac model is coupled with a wave propagation model for the aorta and a closed lumped-parameter circulation model. Parameters for the normal heart and aorta are determined by fitting to experimental data. Changes in the timing and magnitude of pulse wave reflections by the aorta are demonstrated with changes in compliance and taper of the aorta as seen in aging (decreased compliance, increased diameter and length), and resulting effects on LV pressure–volume loops and LV fiber stress and sarcomere shortening are predicted. Effects of aging of the aorta combined with reduced LV contractile force (failing heart) are examined. In the failing heart, changes in aortic properties with aging affect stroke volume and sarcomere shortening without appreciable augmentation of aortic pressure, and the reflected pressure wave contributes an increased proportion of aortic pressure.
The inaugural Diversity and Inclusion in Science Session was held during the 2021 Society for Melanoma Research (SMR) congress. The goal of the session was to discuss diversity, equity, and inclusion in the melanoma research community and strategies to promote the advancement of underrepresented melanoma researchers. An international survey was conducted to assess the diversity, equity, and inclusion (DEI) climate among researchers and clinicians within the Society for Melanoma Research (SMR). The findings suggest there are feelings and experiences of inequity, bias, and harassment within the melanoma community that correlate with one's gender, ethnic/racial group, and/or geographic location. Notably, significant reports of inequity in opportunity, discrimination, and sexual harassment demonstrate there is much work remaining to ensure all scientists in our community experience an academic workplace culture built on mutual respect, fair access, inclusion, and equitable opportunity. 相似文献
Biochar and manure can be used for sustainable land management. However, little is known about how soil amendments might affect surface and belowground microbial processes and subsequent wood decomposition. In a split-split-split plot design, we amended soil with two rates of manure (whole plot; 0 and 9 Mg ha−1) and biochar (split plot; 0 and 10 Mg ha−1). Wood stakes of three species (hybrid poplar, triploid Populus tomentosa Carr.; aspen, Populus tremuloides Michx.; and pine, Pinus taeda L.) were placed in two positions (horizontally on the soil surface, and inserted vertically in the mineral soil), which served as a substrate for fungal growth. In 3 years, the decomposition rate (density loss), moisture content, and fungal community (via high-throughput sequencing methods) of stakes were evaluated. Results indicated that biochar and/or manure increased the wood stake decomposition rates, moisture content, and operational taxonomic unit abundance. However, the richness and diversity of fungi were dependent on wood stake position (surface > mineral), species (pine > the two Populus), and sample dates. This study highlights that soil amendment with biochar and/or manure can alter the fungal community, which in turn can enhance an important soil process (i.e., decomposition). 相似文献
Treatment-resistant depression (TRD) is common and associated with multiple serious public health implications. A consensus definition of TRD with demonstrated predictive utility in terms of clinical decision-making and health outcomes does not currently exist. Instead, a plethora of definitions have been proposed, which vary significantly in their conceptual framework. The absence of a consensus definition hampers precise estimates of the prevalence of TRD, and also belies efforts to identify risk factors, prevention opportunities, and effective interventions. In addition, it results in heterogeneity in clinical practice decision-making, adversely affecting quality of care. The US Food and Drug Administration (FDA) and the European Medicines Agency (EMA) have adopted the most used definition of TRD (i.e., inadequate response to a minimum of two antidepressants despite adequacy of the treatment trial and adherence to treatment). It is currently estimated that at least 30% of persons with depression meet this definition. A significant percentage of persons with TRD are actually pseudo-resistant (e.g., due to inadequacy of treatment trials or non-adherence to treatment). Although multiple sociodemographic, clinical, treatment and contextual factors are known to negatively moderate response in persons with depression, very few factors are regarded as predictive of non-response across multiple modalities of treatment. Intravenous ketamine and intranasal esketamine (co-administered with an antidepressant) are established as efficacious in the management of TRD. Some second-generation antipsychotics (e.g., aripiprazole, brexpiprazole, cariprazine, quetiapine XR) are proven effective as adjunctive treatments to antidepressants in partial responders, but only the olanzapine-fluoxetine combination has been studied in FDA-defined TRD. Repetitive transcranial magnetic stimulation (TMS) is established as effective and FDA-approved for individuals with TRD, with accelerated theta-burst TMS also recently showing efficacy. Electroconvulsive therapy is regarded as an effective acute and maintenance intervention in TRD, with preliminary evidence suggesting non-inferiority to acute intravenous ketamine. Evidence for extending antidepressant trial, medication switching and combining antidepressants is mixed. Manual-based psychotherapies are not established as efficacious on their own in TRD, but offer significant symptomatic relief when added to conventional antidepressants. Digital therapeutics are under study and represent a potential future clinical vista in this population. 相似文献