25 aromatic carboxylic acids which are analogs of benzoic acid were tested in the rat diaphragm preparation for effects on chloride conductance (G(Cl)). Of the 25, 19 were shown to reduce membrane G(Cl) with little effect on other membrane parameters, although their apparent K(i) varied widely. This inhibition was reversible if exposure times were not prolonged. The most effective analog studied was anthracene-9-COOH (9-AC; K(i) = 1.1 x 10(-5) M). Active analogs produced concentration-dependent inhibition of a type consistent with interaction at a single site or group of sites having similar binding affinities, although a correlation could also be shown between lipophilicity and K(i). Structure-activity analysis indicated that hydrophobic ring substitution usually increased inhibitory activity while para polar substitutions reduced effectiveness.
These compounds do not appear to inhibit G(Cl) by altering membrane surface charge and the inhibition produced is not voltage dependent. Qualitative characteristics of the I-V relationship for Cl(-) current are not altered. Conductance to all anions is not uniformly altered by these acids as would be expected from steric occlusion of a common channel. Concentrations of 9-AC reducing G(Cl) by more than 90 percent resulted in slight augmentation of G(I). The complete conductance sequence obtained at high levels of 9-AC was the reverse of that obtained under control conditions. Permeability sequences underwent progressive changes with increasing 9-AC concentration and ultimately inverted at high levels of the analog. Aromatic carboxylic acids appear to inhibit G(Cl) by binding to a specific intramembrane site and altering the selectivity sequence of the membrane anion channel.
Cell death was studied in barley (cv Himalaya) aleurone cells treated with abscisic acid and gibberellin. Aleurone protoplasts incubated in abscisic acid remained viable in culture for at least 3 weeks, but exposure to gibberellin initiated a series of events that resulted in death. Between 4 and 8 days after incubation in gibberellin, >70% of all protoplasts died. Death, which occurred after cells became highly vacuolated, was manifest by an abrupt loss of plasma membrane integrity followed by rapid shrinkage of the cell corpse. Hydrolysis of DNA began before death and occurred as protoplasts ceased production of alpha-amylase. DNA degradation did not result in the accumulation of discrete low molecular weight fragments. DNA degradation and cell death were prevented by LY83583, an inhibitor of gibberellin signaling in barley aleurone. We conclude that cell death in aleurone cells is hormonally regulated and is the final step of a developmental program that promotes successful seedling establishment. 相似文献
Light microscopy was used to study the structure and function of vacuoles in living protoplasts of barley (Hordeum vulgare cv Himalaya) aleurone. Light microscopy showed that aleurone protoplasts contain two distinct types of vacuole: the protein storage vacuole and a lysosome-like organelle, which we have called the secondary vacuole. Fluorescence microscopy using pH-sensitive fluorescent probes and a fluorogenic substrate for cysteine proteases showed that both protein storage vacuoles and secondary vacuoles are acidic, lytic organelles. Ratio imaging showed that the pH of secondary vacuoles was lower in aleurone protoplasts incubated in gibberellic acid than in those incubated in abscisic acid. Uptake of fluorescent probes into intact, isolated protein storage vacuoles and secondary vacuoles required ATP and occurred via at least two types of vanadate-sensitive, ATP-dependent tonoplast transporters. One transporter catalyzed the accumulation of glutathione-conjugated probes, and another transported probes not conjugated to glutathione. 相似文献
alpha 1-Proteinase inhibitors (alpha 1-PIs) are members of the serpin
superfamily of proteinase inhibitors, and are important in the maintenance
of homeostasis in a wide variety of animal taxa. Previous studies have
shown that in mice (genus Mus), evolution of alpha 1-PIs is characterized
by gene amplification, region-specific concerted evolution, and rapid
accumulation of amino acid substitutions. The latter occurs primarily in
the reactive center, which is the region of the alpha 1-PI molecule that
determines the inhibitor's specificity for target proteinases. The P1
residue within the reactive center, which is methionine in so-called
orthodox alpha 1-PIs and an amino acid other than methionine in unorthodox
alpha 1-PIs, is a primary determinant of inhibitor specificity. In the
present study, we find that the expression of mRNAs encoding unorthodox
alpha 1-PIs is polymorphic within Mus species, i.e., among individuals or
inbred strains. This is in striking contrast to mRNAs that encode orthodox
alpha 1-PIs, whose concentrations are relatively invariant. The
intraspecies variations in mRNA expression represent polymorphisms in the
structure of the alpha 1- PI gene family. The results, taken together with
previously described aspects of alpha 1-PI evolution, indicate that the
dissimilar levels of polymorphism exhibited by orthodox and unorthodox
alpha 1-PIs, which likely have distinct physiological functions, may
reflect different levels of selective constraint. The significance of this
finding to the evolution of gene families is discussed.
相似文献
Internal eliminated sequences (IESs) often interrupt ciliate genes in the
silent germline nucleus but are exactly excised and eliminated from the
developing somatic nucleus from which genes are then expressed. Some long
IESs are transposons, supporting the hypothesis that short IESs are ancient
transposon relics. In light of that hypothesis and to explore the
evolutionary history of a collection of IESs, we have compared various
alleles of a particular locus (the 81 locus) of the ciliated protozoa
Oxytricha trifallax and O. fallax. Three short IESs that interrupt two
genes of the locus are found in alleles from both species, and thus must be
relatively ancient, consistent with the hypothesis that short IESs are
transposon relics. In contrast, TBE1 transposon interruptions of the locus
are allele-specific and probably the results of recent transpositions.
These IESs (and the TBE1s) are precisely excised from the DNA of the
developing somatic macronucleus. Each IES interrupts a highly conserved
sequence. A few nucleotides at the ends of each IES are also conserved,
suggesting that they interact critically with IES excision machinery.
However, most IES nucleotide positions have evolved at high rates, showing
little or no selective constraint for function. Nonetheless, the length of
each IES has been maintained (+/- 3 bp). While one IES is approximately 33
bp long, three other IESs have very similar sizes, approximately 70 bp
long. Two IESs are surrounded by direct repeats of the sequence TTCTT. No
other sequence similarities were found between any of the four IESs.
However, the ends of one IES do match the inverted terminal repeat
consensus sequence of the "TA" IESs of Paramecium. Three O. trifallax
alleles appear to have been recipients in recent conversion events that
could have been provoked by double-strand breaks associated with IES ends
subsequent to IES transposition. Our findings support the hypothesis that
short IESs evolved from ancient transposons that have lost most of their
sequences, except those necessary for precise excision during macronuclear
development.
相似文献
Antigen/antibody complexes can efficiently target antigen presenting cells to allow stimulation of the cellular immune response. Due to the difficulty of manufacture and their inherent instability complexes have proved inefficient cancer vaccines. However, anti-idiotypic antibodies mimicking antigens have been shown to stimulate both antibody and T cell responses. The latter are due to T cell mimotopes expressed within the complementarity-determining regions (CDRs) of antibodies that are efficiently presented to dendritic cells in vivo. Based on this observation we have designed a DNA vaccine platform called ImmunoBody™, where cytotoxic T lymphocyte (CTL) and helper T cell epitopes replace CDR regions within the framework of a human IgG1 antibody. The ImmunoBody™ expression system has a number of design features which allow for rapid production of a wide range of vaccines. The CDR regions of the heavy and light chain have been engineered to contain unique restriction endonuclease sites, which can be easily opened, and oligonucleotides encoding the T cell epitopes inserted. The variable and constant regions of the ImmunoBody™ are also flanked by restriction sites, which permit easy exchange of other IgG subtypes. Here we show a range of T cell epitopes can be inserted into the ImmunoBody™ vector and upon immunization these T cell epitopes are efficiently processed and presented to stimulate high frequency helper and CTL responses capable of anti-tumor activity.Key words: DNA vaccines, cancer vaccines, melanoma, CTL, helper T cells相似文献
There is growing concern about mitigation-driven translocations that move animals from anthropogenic threats at donor sites because of their failure rate and lack of application of scientific principles and best practice. We reviewed all known lizard translocations in New Zealand between 1988 and 2013 and identified 85 translocations of 30 lizard taxa to 46 release sites. Most translocations (62%) were motivated by conservation goals for the species or the release site, and one-third were mitigation-driven translocations, typically motivated by habitat loss due to development. Mitigation-driven translocations began in 2003, and since that time have equalled the number of conservation-motivated translocations. Conservation-motivated translocations usually released lizards on islands without mammalian predators, whereas mitigation-driven translocations usually relocated lizards to mainland sites with introduced predators. Long-term monitoring has been sparse and often rudimentary. Eight lizard translocations have recorded population growth, including one mitigation-driven translocation that was into a fenced reserve. Research on commonly used management techniques to mitigate human-related impacts is recommended to establish whether these techniques benefit lizards in the long term. 相似文献