Background: It has become increasingly evident that dietary Se plays a significant role in reducing the incidence of lung, colorectal and prostate cancer in humans. Different forms of Se vary in their chemopreventative efficacy, with Se-methylselenocysteine being one of the most potent. Interestingly, the Se accumulating plant Astragalus bisulcatus (Two-grooved poison vetch) contains up to 0.6% of its shoot dry weight as Se-methylselenocysteine. The ability of this Se accumulator to biosynthesize Se-methylselenocysteine provides a critical metabolic shunt that prevents selenocysteine and selenomethionine from entering the protein biosynthetic machinery. Such a metabolic shunt has been proposed to be vital for Se tolerance in A. bisulcatus. Utilization of this mechanism in other plants may provide a possible avenue for the genetic engineering of Se tolerance in plants ideally suited for the phytoremediation of Se contaminated land. Here, we describe the overexpression of a selenocysteine methyltransferase from A. bisulcatus to engineer Se-methylselenocysteine metabolism in the Se non-accumulator Arabidopsis thaliana (Thale cress).

A hybrid of sweet corn, Zea mays L. ('1720'; Rogers Brothers Seed Co.), was found to be comprised of glycinebetaine-positive and glycinebetaine-deficient individuals in a 1:1 mixture. This phenomenon was traced to segregation for a single, nuclear, dominant gene determining leaf glycinebetaine content within the female inbred parent of this hybrid. Selection for homozygous recessive (glycinebetaine-deficient) and homozygous dominant (glycinebetaine-positive) genotypes of the female inbred parent enabled production of two isogenic versions of hybrid '1720' differing with respect to a single copy of the dominant allele, by mating these female parent selections with the common homozygous recessive (glycinebetaine-deficient) male parent. These two isogenic hybrids are shown to differ by a factor of 300-to 400-fold in glycinebetaine titer of young expanding leaves of salinized plants, but exhibit no striking differences in the levels of free amino acids or the level of N-methyinicotinic acid (nicotinic acid betaine; trigonelline). The only significant difference between the two hybrids in terms of amino acid composition was found to be in the level of alanine under nonsalinized conditions. The betaine-deficient hybrid exhibited a 14% lower alanine level than the betaine-positive hybrid. Betaine deficiency was not associated with altered stress-induced accumulation of amino acids such as proline, serine, and asparagine plus aspartate, attesting to the high specificity of the genetic difference between these isogenic hybrids with respect to betaine accumulation. This germplasm offers unique opportunities to test whether a single dominant allele determining stress-induced betaine accumulation capacity influences stress resistance in maize.Osmotic adjustment is widely considered to be an adaptive response to water deficits and salinity stress (6,7,14,20,21). Bet2 (betaine) accumulation in response to salinity stress is proposed to play an important role in osmotic adjustment in halophytic members of the Chenopodiaceae and Gramineae (7,(18)(19)(20)22), and may function as a compatible osmotic solute of the cytoplasm and/or chloroplast (17,22).

Screening of a range of public maize (Zea mays L.) inbred lines for glycinebetaine (betaine) content over two growing seasons (1987 and 1988), using fast atom bombardment mass spectrometry, has identified 19 public inbred lines which all exhibit low betaine levels (<100 nanomoles per gram fresh weight). These include common inbreds such as A188, A619, B37, H95, N6, and Oh43. Several inbreds exhibit high betaine levels (3000 to 10000 nanomoles per gram fresh weight); in these strongly betainepositive inbreds, betaine levels tended to be, on average, 1.38-fold greater in the 1988 growing season presumably in part due to field water deficits experienced during the drought of 1988. Where several different sources of the same inbred line were available (including cytoplasmic male sterile and restored lines of A632, B37, B73, Oh43, and WF9), betaine levels were found to be similar when the inbreds were tested in the same environment. Because W22-R/r-X1 was found to be strongly betaine-positive, it should be possible to map the putative recessive gene(s) determining betaine deficiency to specific chromosome(s) from monosomics resulting from crosses between W22-R/r-X1 and betaine-deficient lines.

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A serious limitation to the use of N(O,S)-heptafluorobutyryl isobutyl amino acid derivatives in the analysis of 15N-labeling kinetics of amino acids in plant tissues, is that the amides glutamine and asparagine undergo acid hydrolysis to glutamate and aspartate, respectively, during derivatization. This led us to consider an altemative procedure (G Fortier et a/. [1986] (N-HFBI3) derivatives of amino acids are convenient for routine quantification of amino acid pools in higher plant tissues by GC, and determination of the 'sN abundance of the majority of their aamino groups by GC-MS (2,(11)(12)(13)(14). However, a serious limitation ofthis analytical methodology in '5N-tracer studies is that during derivatization, glutamine and asparagine undergo deamidation to glutamate and aspartate, respectively (12,14). Thus, independent quantification of glutamate, glutamine, asparagine, and aspartate requires prior separation of the neutral + basic and acidic amino acid fractions using ion exchange chromatography, and, more importantly, two key N groups (the amide moieties of glutamine and asparagine) are lost as NH4' during derivatization of the neutral + basic amino acid fraction (2,(11)(12)(13)(14). The latter problem has prompted us to seek methods for sensitively estimating the amide-['5N] abundance of glutamine and asparagine, using N(O,S)-Heptafluorobutyryl isobutylLemna minor as a model plant system.A specific solution to the problem of estimating the amide-['5N] abundance of asparagine was found by noting that a small fraction (0.5-2%) of the asparagine in neutral + basic amino acid fractions routinely undergoes dehydration and nitrile formation at the amide position during derivatization, yielding the N-HFBI derivative of j3-cyanoalanine; a derivative which contains both the amide and amino nitrogen groups of asparagine. However, this procedure requires relatively large quantities of asparagine (>500 nmol/derivatization) in order to reliably detect the dehydration product at sufficient levels to obtain meaningful electron ionization mass spectra and hence isotope abundance, and moreover, a comparable dehydration product of glutamine is not obtained. A fraction of the glutamine undergoes deamidation and cyclization to yield the N-HFBI derivative of pyroglutamate (2-pyrrolidone-5-carboxylate), but the latter contains only the amino nitrogen moiety ofglutamine (12). We therefore sought alternative derivatization schemes for the amides which would produce derivatives containing both the amide and amino nitrogen moieties of glutamine. Woo et al. (19) have successfully employed trimethylsilyl-3