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The multigene family encoding the small subunit of ribulose 1,5-bisphosphate carboxylase/oxygenase [EC 4.1.1.39] in Phaseolus vulgaris L. contains,at least three members. cDNA clones representing 3 members of the family (rbcS genes) have been characterised in terms of structure and expression. These rbcS genes show very high nucleotide sequence homologies in the coding regions for both the transit peptide and the mature polypeptide. The nucleotide sequences for the 5' - and 3' -untranslated regions of these genes which were present in the cDNA clones showed far more divergence. One of the rbcS cDNA clones contained two regions of intronic sequences, the first lying between the codons for the 2nd and 3rd amino acids of the mature polypeptide and the second lying between the codons for the 47th and 48th amino acids of the mature polypeptide.

Many environmental stresses result in increased generation of active oxygen species (AOS) in plant cells, leading to the induction of protective mechanisms. In this study, signalling components linking AOS perception to downstream responses were examined, with particular emphasis on H2O2 signalling. All AOS investigated had an early [Ca2+]cyt peak in common, but differed in other aspects of their Ca2+ signatures, indicating that the plant is able to discriminate between different types of AOS. An early event in AOS signal transduction may involve changes in the cellular redox balance as reduction of glutathione levels prior to stress application increased the height of the first [Ca2+]cyt peak. Inhibiting or enhancing the height of the H2O2-triggered Ca2+ signature lead to inhibition or enhancement of GST1 and APX1 induction, respectively, demonstrating that the Ca2+ signature is required for induction of genes encoding antioxidant enzymes. OX1, encoding a putative ser/thr kinase, was shown to be involved in signal transduction in response to H2O2-generating stresses. Transcript levels of OX1 were increased upon treatment with H2O2 and a range of abiotic and biotic stresses as well as ABA, all of which have been shown to result in H2O2 accumulation. Inhibition of stress-induced [Ca2+]cyt elevations inhibited OX1 induction, placing the OX1 kinase downstream of Ca2+ in the signalling chain. OX1 is required for full activation of AtMPKS and AtMPK6 in response to ozone fumigation, indicating that OX1 functions upstream of these MAP kinases. An ox1 null-mutant displayed enhanced susceptibility to infection with a virulent Peronospora parasitica isolate as well as reduced induction of several defence genes. In addition, the ox1 mutant exhibited shorter root hairs and an early flowering phenotype. AOS treatment induced several genes encoding AtERF transcription factors, but did not have an effect on other members of this family. Induction occurred in an ethylene-independent but Ca2+-dependent manner.

Increases in temperature damage plant cells, and plants react to heat stress by inducing of a number of protective mechanisms. In this study, it has been shown that heat damages cells both directly while the plants are being heated, and indirectly through heat induced oxidative stress during recovery from heating. Different stress response pathways are induced in each case. Evidence implicates salicylic acid, abscisic acid, ACC (a precursor of ethylene), calcium ions and active oxygen species in pathways resulting in thermotolerance (i.e. increased survival at high temperature). Addition of these potential second messengers results in increased survival and decreased oxidative damage after heating (as measured using the TBARS assay), while silencing the pathways through use of mutants, inhibitors or transgenes results in decreased thermotolerance. In vivo calcium measurements show cytosolic calcium transients only at initiation of cooling after heat stress. Calcium chloride also induces thermotolerance when added after heating, as does ACC. These substances can induce early increases in ascorbate peroxidase activity after heating, and induce expression of antioxidant genes. Thus they may play a role in heat induced oxidative stress response pathways in recovery. Two different pathways induced during heating appear to result in the expression of genes for heat shock proteins. The predominant pathway induced at 30oC involves ABA, while that at 35-40oC involves an oxidative burst generated through the NADPH oxidase, atrbohB. Addition of SA can induce HSP (heat shock protein) expression, but there is little evidence that this occurs endogenously in plant cells, although nahG plants unable to signal via SA are thermosensitive. Results in this study indicate that there are several signalling pathways associated with heat shock. The pathway induced during recovery from heating involves calcium ions and ethylene, and results in increased antioxidant capacity. During heating two pathways induce expression of HSPs: one involving abscisic acid and possibly salicylic acid and one involving an oxidative burst. At least one further pathway is believed to exist, which involves protein kinases and phosphatases as heat shock causes up-regulation of expression of certain genes for these signalling components.