SummaryThe AtMYB103 gene is a member of the R2R3 MYB gene family in Arabidopsis thaliana. Using the GUS reporter gene, AtMYB103 expression was found to be restricted to the tapetum of developing anthers. Employing RT-PCR and in situ hybridisation, we now show that AtMYB103 is also expressed in trichomes. GUS expression in trichomes was obtained by incorporating the coding and 3 H -untranslated regions of AtMYB103 into the promoter-GUS constructs. Sense and antisense technologies were used to downregulate AtMYB103 expression. In transgenic lines with reduced AtMYB103 transcript levels, pollen, tapetum and trichome development were altered. The majority of the pollen grains were distorted in shape and had reduced or no cytoplasmic content. Tapetal degeneration occurred early, and large opaque bodies appeared in the tapetal cytoplasm. In transgenic plants, trichomes on cauline and rosette leaves produced additional branches. These overbranched trichomes contained more nuclear DNA than the wild-type trichomes. The results indicate that AtMYB103 is required for tapetal development and microsporogenesis, and negatively regulates trichome endoreduplication linked to the trichome branching.

The Arabidopsis thaliana MYB5 gene is expressed in trichomes and seeds, including the seed coat. Constitutive expression of MYB5 resulted in the formation of more small trichomes and ectopic trichomes and a reduction in total leaf trichome numbers and branching. A myb5 mutant displayed minimal changes in trichome morphology, while a myb23 mutant produced increased numbers of small trichomes and two-branched trichomes. A myb5 myb23 double mutant developed more small rosette trichomes and two-branched trichomes than the single mutants. These results indicate that MYB5 and MYB23 regulate trichome extension and branching. The seed coat epidermal cells of myb5 and myb5 myb23 were irregular in shape, developed flattened columellae, and produced less mucilage than those of the wild type. Among the downregulated genes identified in the myb5 seeds using microarray analysis were ABE1 and ABE4 (a/b fold hydrolase/esterase genes), MYBL2, and GLABRA2. The same genes were also downregulated in transparent testa glabra1 (ttg1) seeds, suggesting that MYB5 collaborates with TTG1 in seed coat development. These genes were upregulated in leaves and roots by ectopically expressed MYB5. The MYBL2, ABE1, and ABE4 promoters were active in seeds, including seed coats, and the latter two also in trichomes. Models of the MYB5 regulatory networks involved in seed coat and trichome development are presented.

SummaryThe Arabidopsis AtMYB103 gene codes for an R2R3 MYB domain protein whose expression is restricted to the tapetum of developing anthers and to trichomes. Down-regulation of expression using anti-sense leads to abnormal tapetum and pollen development, although restorer constitute a reversible male sterility system which could be adapted for hybrid seed production. This is the first reversible male sterility system targeting a transcription factor essential for pollen development. Strategies for generating inducible male sterility and maintainable male sterility for the production of hybrid crops are discussed.

The nar operon, which encodes the three subunits of nitrate reductase in Escherichia coli, is (18,20). The anaerobic formation of nitrate reductase is under the control of thefnr gene product (2, 22), a positive pleiotropic regulator which also affects the formation of several other enzymes involved in anaerobic metabolism (10,19,24). A second positive regulator, the product of the narL gene, is required for the further induction of the anaerobic production of nitrate reductase by nitrate. A mutation in the narL locus permits the normal anaerobic induction of nitrate reductase but makes the cells unresponsive to nitrate (22). The narL mutations map in the same region of the chromosome (22), but are located outside the nar operon (E. S. Edwards and J. A. DeMoss, unpublished data).The nar operon, which encodes the three subunits of nitrate reductase, consists of three structural genes transcribed in the order narG (et subunit), narH (1 subunit) and narI (-y subunit) (1,4,5,23). The intact nar operon cloned into a multicopy plasmid overproduced the three subunits of nitrate reductase in the transformed strain under the control of the same factors which regulate the chromosomal operon (17).Recently, a portion of the 5' untranslated region of the nar operon was sequenced and the translation start site of narG was determined (11,14). Studies with deletions into the 5' end of the nar operon suggested that the 5' untranslated * Corresponding author. region includes a promoter and two regulatory regions (11). A regulatory domain immediately adjacent to the narG gene apparently includes a transcriptional start site and the sequence recognized by the fnr gene product, which induces the anaerobic expression of nar operon. A second regulatory domain extending further upstream determines the interaction with the narL product, which is activated by nitrate and leads to an increase of the anaerobic expression of the operon.In the studies reported here, we have sequenced the entire regulatory region, defined more precisely the 5' limit of the operon, and determined the transcriptional start site under the several conditions of induction. MATERIALS AND METHODSStrains and growth conditions. All bacterial strains used in this study are listed in

A novel MYB-Uke gene (AtMYB103) was isolated from a genomic library of Arabidopsis. Plants transgenic for chimeric AtMYB103 promoter/Gf/5' genes expressed the enzyme in early anthers. In situ hybridization of flower sections showed a high level of AtMYB103 mRNA in the tapetum and middle layer of developing anthers.