Sulphur starvation induces the expression of microRNA-395 and one of its target genes but in different cell types.

Plants play an important role in the global sulphur cycle because they assimilate sulphur from the environment and build it into methionine and cysteine. Several genes of the sulphur assimilation pathway are regulated by microRNA-395 (miR395) that is itself induced by a low-sulphur (-S) environment. Here, we show that the six Arabidopsis miR395 loci are induced differently. We find that MIR395 loci are expressed in the vascular system of roots and leaves and root tips. Induction of miR395 by a -S environment in both roots and leaves suggests that translocation of miR395 from leaves to roots through the phloem is not necessary for plants growing on -S soil/medium. We also demonstrate that induction of miR395 is controlled by SLIM1, a key transcription factor in the sulphur assimilation pathway. Unexpectedly, the mRNA level of a miR395 target gene, SULTR2;1, strongly increases during miR395 induction in roots. We show that the spatial expression pattern of MIR395 transcripts in the vascular system does not appear to overlap with the expression pattern previously reported for SULTR2;1 mRNA. These results illustrate that negative temporal correlation between the expression level of a miRNA and its target gene in a complex tissue cannot be a requirement for target gene validation.

Characterization and expression analysis of a serine acetyltransferase gene family involved in a key step of the sulfur assimilation pathway in Arabidopsis.

Ser acetyltransferase (SATase; EC 2.3.1.30) catalyzes the formation of O-acetyl-Ser from L-Ser and acetyl-CoA, leading to synthesis of Cys. According to its position at the decisive junction of the pathways of sulfur assimilation and amino acid metabolism, SATases are subject to regulatory mechanisms to control the flux of Cys synthesis. In Arabidopsis (Arabidopsis thaliana) there are five genes encoding SATase-like proteins. Two isoforms, Serat3;1 and Serat3;2, were characterized with respect to their enzymatic properties, feedback inhibition by L-Cys, and subcellular localization. Functional identity of Serat3;1 and Serat3;2 was established by complementation of a SATase-deficient mutant of Escherichia coli. Cytosolic localization of Serat3;1 and Serat3;2 was confirmed by using fusion construct with the green fluorescent protein. Recombinant Serat3;1 was not inhibited by L-Cys, while Serat3;2 was a strongly feedback-inhibited isoform. Quantification of expression patterns indicated that Serat2;1 is the dominant form expressed in most tissues examined, followed by Serat1;1 and Serat2;2. Although Serat3;1 and Serat3;2 were expressed weakly in most tissues, Serat3;2 expression was significantly induced under sulfur deficiency and cadmium stress as well as during generative developmental stages, implying that Serat3;1 and Serat3;2 have specific roles when plants are subjected to distinct conditions. Transgenic Arabidopsis plants expressing the green fluorescent protein under the control of the five promoters indicated that, in all Serat genes, the expression was predominantly localized in the vascular system, notably in the phloem. These results demonstrate that Arabidopsis employs a complex array of compartment-specific SATase isoforms with distinct enzymatic properties and expression patterns to ensure the provision of Cys in response to developmental and environmental changes.

Heavy metal tolerance of transgenic tobacco plants over-expressing cysteine synthase.

Cysteine synthase [O-acetyl-L-serine(thiol)lyase] catalyzes the final step for L-cysteine biosynthesis in plants. The tolerance of transgenic tobacco plants over-expressing cysteine synthase cDNA in cytosol (3F), chloroplasts (4F) and in both organelles (F1) was investigated towards heavy metals such as Cd, Se, Ni, Pb and Cu. The transgenic plants were significantly more tolerant than wild-type plants in agar medium containing Cd, Se and Ni. The F1 transgenic plants had a higher resistance than other transgenic lines towards these metals and could enhance accumulation of Cd in shoot. These results suggest that the transgenic plants over-expressing cysteine synthase both in cytosol and chloroplasts can be applicable to phyto-remediation of Cd from contaminated soils.