Increased leakiness of the tetracycline-inducible Triple-Op promoter in dividing cells renders it unsuitable for high inducible levels of a dominant negative CDC2aAt gene

A tetracycline-inducible promoter system was used to generate transgenic tobacco plants that confer inducible expression of the wild type or a dominant negative allele of the gene coding for the cyclin-dependent kinase (CDK) of Arabidopsis thaliana CDC2aAt. Although the total extractable CDK activity was doubled, the induced expression of the wild-type CDC2aAt did not correlate with any change of the cell cycle kinetics. An increase of CDK activity upon CDC2aAt expression was only seen in dividing cell populations, demonstrating that CDC2aAt expression itself is not sufficient to induce CDK activation. Induced expression of the dominant negative CDC2aAt.N146 correlated with a reduction of CDK activity to 66% of the level found in non-induced cells. This decrease was not sufficient to block cell division. The isolation of plants showing only low inducible levels of CDC2aAt.N146 suggests that a counterselection against strong inducible lines had occurred. Accordingly, Triple-Op promoter activity was found in dividing cells in the absence of tetracycline.     

A common F-box gene regulates the leucine homeostasis of Medicago truncatula and Arabidopsis thaliana

The F-box domain is a conserved structural protein motif that most frequently interacts with the SKP1 protein, the core of the SCFs (SKP1-CULLIN-F-box protein ligase) E3 ubiquitin protein ligases. As part of the SCF complexes, the various F-box proteins recruit substrates for degradation through ubiquitination. In this study, we functionally characterized an F-box gene (MtF-box) identified earlier in a population of Tnt1 retrotransposon-tagged mutants of Medicago truncatula and its Arabidopsis thaliana homolog (AtF-box) using gain- and loss-of-function plants. We highlighted the importance of MtF-box in leaf development of M. truncatula. Protein–protein interaction analyses revealed the 2-isopropylmalate synthase (IPMS) protein as a common interactor partner of MtF-box and AtF-box, being a key enzyme in the biosynthesis pathway of the branched-chain amino acid leucine. For further detailed analysis, we focused on AtF-box and its role during the cell division cycle. Based on this work, we suggest a mechanism for the role of the studied F-box gene in regulation of leucine homeostasis, which is important for growth.

     

Herbicide Safener-Inducible Gene Expression in Arabidopsis thaliana

The potential use of a new chemical-inducibie gene expressionsystem in Arabidopsis thaliana has been examined. The systemis based on the maize In2-2 promoter which is activated by benzenesulfonamideherbicide safe-ners. Plants transformed with the ß-glucuronidase(gus) reporter gene under the control of the In2-2 promoterwere grown in the presence of different safeners and the inducedGUS activity pattern was studied histochemically. In the absenceof safeners, the In2-2 promoter was not active. Applicationof different safeners induced distinct gus expression patterns,including expression in the root, hy-dathodes, and the shootapical meristem. Plants maintained continuously on inducingconcentrations of the safeners were retarded in growth. Thegrowth inhibition effects of the Sa5 safener could be overcomein a sul-fonylurea-resistant background. In2-2 promoter activitycould also be induced by the sulfonylurea herbicide chlor-sulfuron.In the sulfonylurea-resistant background, which derives fromherbicide-resistant acetolactate synthase activity, inductionof the In2-2 promoter by chlorsulfuron was lower. Furthermore,branched-chain amino acids, known to inhibit acetolactate synthaseactivity, also induced In2-2 promoter activity. Our data suggesta strong correlation between In2-2 expression and inhibitionof the acetolactate synthase activity. (Received November 12, 1996; Accepted February 21, 1997)