Mechanisms of metal resistance in plants: aluminum and heavy metals

Plants have evolved sophisticated mechanisms to deal with toxic levels of metals in the soil. In this paper, an overview of recent progress with regards to understanding fundamental molecular and physiological mechanisms underlying plant resistance to both aluminum (Al) and heavy metals is presented. The discussion of plant Al resistance will focus on recent advances in our understanding of a mechanism based on Al exclusion from the root apex, which is facilitated by Al-activated exudation of organic acid anions. The consideration of heavy metal resistance will focus on research into a metal hyperaccumulating plant species, the Zn/Cd hyperaccumulator, Thlaspi caerulescens, as an example for plant heavy metal research. Based on the specific cases considered in this paper, it appears that quite different strategies are used for Al and heavy metal resistance. For Al, our current understanding of a resistance mechanism based on excluding soil-borne Al from the root apex is presented. For heavy metals, a totally different strategy based on extreme tolerance and metal hyperaccumulation is described for a hyperaccumulator plant species that has evolved on naturally metalliferous soils. The reason these two strategies are the focus of this paper is that, currently, they are the best understood mechanisms of metal resistance in terrestrial plants. However, it is likely that other mechanisms of Al and/or heavy metal resistance are also operating in certain plant species, and there may be common features shared for dealing with Al and heavy resistance. Future research may uncover a number of novel metal resistance mechanisms in plants. Certainly the complex genetics of Al resistance in some crop plant species, such as rice and maize, suggests that a number of presently unidentified mechanisms are part of an overall strategy of metal resistance in crop plants.     

Mechanochemical coupling in spin-labeled, active, isometric muscle

Observed effects of inorganic phosphate (P(i)) on active isometric muscle may provide the answer to one of the fundamental questions in muscle biophysics: how are the free energies of the chemical species in the myosin-catalyzed ATP hydrolysis (ATPase) reaction coupled to muscle force?. Pflugers Arch. 414:73-81) showed that active, isometric muscle force varies logarithmically with [P(i)]. Here, by simultaneously measuring electron paramagnetic resonance and the force of spin-labeled muscle fibers, we show that, in active, isometric muscle, the fraction of myosin heads in any given biochemical state is independent of both [P(i)] and force. These direct observations of mechanochemical coupling in muscle are immediately described by a muscle equation of state containing muscle force as a state variable. These results challenge the conventional assumption mechanochemical coupling is localized to individual myosin heads in muscle.     

The joint evolutionary histories of Wolbachia and mitochondria in Hypolimnas bolina

Background  

The interaction between the Blue Moon butterfly, Hypolimnas bolina, and Wolbachia has attracted interest because of the high prevalence of male-killing achieved within the species, the ecological consequences of this high prevalence, the intensity of selection on the host to suppress the infection, and the presence of multiple Wolbachia infections inducing different phenotypes. We examined diversity in the co-inherited marker, mtDNA, and the partitioning of this between individuals of different infection status, as a means to investigate the population biology and evolutionary history of the Wolbachia infections.     

Regulators of cell division in plant tissues

The cytokinins in certain fractions prepared from extracts of immature sweet-corn (Zea mays L.) kernels using polystyrene ion-exchange resins have been further investigated. Cytokinins active in the radish cotyledon bioassay were purified from these fractions and identified as 9--D-glucopyranosylzeatin, 9--D-glucopyranosyldihydrozeatin, O--D-glucopyranosylzeatin. and O--D-glucopyranosyl-9--D-ribofuranosylzeatin. In addition, compounds which resemble zeatin and its glycosides in chromatographic behaviour and in ultraviolet absorption characteristics were purified from extracts of the same material by high-performance liquid chromatography. In addition to zeatin and zeatin riboside, the following compounds were identified unambiguously: O--D-glucopyranosyl-9--D-ribofuranosyldihydrozeatin, O--D-glucopyranosyldihydrozeatin, and hihydrozeatin riboside. A further compound was tentatively identified as O--D-glucopyranosylzeatin, and at least two unidentified compounds appeared to be new derivatives of zeatin. In identifying the above compounds, chemical-ionization mass spectrometry proved to be an invaluable complementary technique, yielding spectra showing intense protonated-molecular-ion peaks and also prominent structure-related fragmentation that was either not evident or very minor in the electron-impact spectra. An assessment of the relative importance of the various possible mechanisms for cytokinin modification and inactivation in mature sweet-corn kernels was made by supplying [³H]zeatin and [³H]zeatin riboside to such kernels after excision. The principal metabolites of zeatin were adenine nucleotides, adenosine and adenine, while little of the metabolite radioactivity was attributable to known O-glucosides. Adenine nucleotides and adenine were the principal metabolites of zeatin riboside, while lesser metabolites were identified as adenosine, dihydrozeatin, and the O-glucosides of dihydrozeatin and dihydrozeatin riboside. Side-chain cleavage, rather than side-chain modification, appears to be the dominant form of cytokinin metabolism in mature sweet-corn kernels.Abbreviations CI-MS chemical-ionization mass spectrum - EIMS electron-impact mass spectrum - GC-MS combined gas chromatography-mass spectrometry - HPLC high performance liquid chromatography - M⁺ molecular ion - MH⁺ protonated molecular ion - TLC thin-layer chromatography - TMS trimethylsilyl - UV ultraviolet XXVII=Letham et al. (1979)     

Development and standardization of multiplexed antibody microarrays for use in quantitative proteomics

BACKGROUND: Quantitative proteomics is an emerging field that encompasses multiplexed measurement of many known proteins in groups of experimental samples in order to identify differences between groups. Antibody arrays are a novel technology that is increasingly being used for quantitative proteomics studies due to highly multiplexed content, scalability, matrix flexibility and economy of sample consumption. Key applications of antibody arrays in quantitative proteomics studies are identification of novel diagnostic assays, biomarker discovery in trials of new drugs, and validation of qualitative proteomics discoveries. These applications require performance benchmarking, standardization and specification. RESULTS: Six dual-antibody, sandwich immunoassay arrays that measure 170 serum or plasma proteins were developed and experimental procedures refined in more than thirty quantitative proteomics studies. This report provides detailed information and specification for manufacture, qualification, assay automation, performance, assay validation and data processing for antibody arrays in large scale quantitative proteomics studies. CONCLUSION: The present report describes development of first generation standards for antibody arrays in quantitative proteomics. Specifically, it describes the requirements of a comprehensive validation program to identify and minimize antibody cross reaction under highly multiplexed conditions; provides the rationale for the application of standardized statistical approaches to manage the data output of highly replicated assays; defines design requirements for controls to normalize sample replicate measurements; emphasizes the importance of stringent quality control testing of reagents and antibody microarrays; recommends the use of real-time monitors to evaluate sensitivity, dynamic range and platform precision; and presents survey procedures to reveal the significance of biomarker findings.     

Retardation of soybean leaf senescence and associated effects on seed composition

Soybean leaf senescence, leaf abscission, and pod yellowing were markedly delayed by sprays of 10^–4 M 6-benzylamino-9-(tetrahydropyran-2-yl)purine plus 5×10^–5 M -naphthalene acetic acid. The pods on the sprayed plants turned yellow 5–7 days later than those on the control plants, and the treated leaves remained dark green even when the pods had already desiccated. The antisenescence spray did not change pod numbers, seed numbers, seed size, or the yield. By retarding senescence, seed nitrogen content was increased in both TCA-soluble and TCA-insoluble fractions. Seed total protein, buffer-extractable total protein, and globulin were increased by 26, 28, and 33 mg/g of seed flour, respectively, and albumin was decreased by 6 mg/g. The overall increase in seed protein caused by spray treatment is confined to the globulin portion.     

Regulators of cell division in plant tissues

6-[³H]Benzylaminopurine was supplied through the transpiration stream to de-rooted Phaseolus vulgaris L. seedlings. The principal metabolite formed was identified as -(6-benzylaminopurin-9-yl)alanine by comparison with the synthetic compound.Abbreviations BAP 6-benzylaminopurine - TLC thin-layer chromatography XXVI=Letham et al. (1978)     

The complex of O-glucosylzeatin derivatives formed in Populus species

When zeatin was supplied to excised leaves of Populus alba, the principal metabolites formed were adenosine, O-β-d-glucopyranosyl-cis-zeatin (derived from cis-zeatin in the commercial zeatin used), O-β-d-glucopyranosylzeatin, and two new metabolites, namely, O-β-d-glucopyranosyldihydrozeatin and O-β-d-glycopyranosyl-9-β-d-ribofuranosyldihydrozeatin, the structures of which were confirmed by unambiguous synthesis. Chromatographic studies indicated that adenosine 5′-phosphate, zeatin 7-glucopyranoside, zeatin 9-glucopyranoside, dihydrozeatin and zeatin 9-riboside were minor metabolites. The principal metabolites of zeatin 9-riboside in P. nigra leaves were the new metabolites O-β-d-glucopyranosyl-9-β-d-ribofuranosylzeatin (synthesized chemically) and O-β-d-glucopyranosl-9-β-d-ribofuranosyldihydrozeatin.     

Glucosylation of cytokinin analogues

Glucosylation of adenine and 6-methylaminopurine was not detected in derooted 10-day-old radish seedlings. However, 4-(purin-6-ylamino)butanoic amide and 6-(3,4-dimethoxybenzylamino)purine (N⁶-substituted adenines with negligible cytokinin activity), like the highly active cytokinin 6-benzylaminopurine, were converted to 7-glucopyranosides. The N²-substituted guanine, 2-benzylaminopurin-6-one, and 6-benzylamino-2-(2-hydroxy-ethylamino)purine were also metabolized to glucosides which were probably 7-glucopyranosides. Hence glucosylation of purines is not restricted to N⁶-substituted adenines with strong cytokinin activity. Although only ca 1.6% of 6-benzylamino-9-(4-chlorobutyl)purine taken up by the derooted seedlings could be accounted for as 7- and 9-glucosides, a considerable proportion was metabolized to these glucosides in cotyledons excised from 2-day-old radish seedlings. The high cytokinin activity of this 9-substituted compound may be a consequence of cleavage of the 4-chlorobutyl groud at N-9.