SummaryWe have produced 22 090 primary transgenic rice plants that carry a T-DNA insertion, which has resulted in 18 358 fertile lines. Genomic DNA gel-blot and PCR analyses have shown that approximately 65% of the population contains more than one copy of the inserted T-DNA. Hygromycin resistance tests revealed that transgenic plants contain an average of 1.4 loci of T-DNA inserts. Therefore, it can be estimated that approximately 25 700 taggings have been generated. The binary vector used in the insertion contained the promoterless b-glucuronidase (GUS) reporter gene with an intron and multiple splicing donors and acceptors immediately next to the right border. Therefore, this gene trap vector is able to detect a gene fusion between GUS and an endogenous gene, which is tagged by T-DNA. Histochemical GUS assays were carried out in the leaves and roots from 5353 lines, mature¯owers from 7026 lines, and developing seeds from 1948 lines. The data revealed that 1.6±2.1% of tested organs were GUS-positive in the tested organs, and that their GUS expression patterns were organ-or tissue-speci®c or ubiquitous in all parts of the plant. The large population of T-DNA-tagged lines will be useful for identifying insertional mutants in various genes and for discovering new genes in rice.

Loss of green color in leaves results from chlorophyll (Chl) degradation in chloroplasts, but little is known about how Chl catabolism is regulated throughout leaf development. Using the staygreen (sgr) mutant in rice (Oryza sativa), which maintains greenness during leaf senescence, we identified Sgr, a senescence-associated gene encoding a novel chloroplast protein. Transgenic rice overexpressing Sgr produces yellowish-brown leaves, and Arabidopsis thaliana pheophorbide a oxygenase-impaired mutants exhibiting a stay-green phenotype during dark-induced senescence have reduced expression of Sgr homologs, indicating that Sgr regulates Chl degradation at the transcriptional level. We show that the leaf staygreenness of the sgr mutant is associated with a failure in the destabilization of the light-harvesting chlorophyll binding protein (LHCP) complexes of the thylakoid membranes, which is a prerequisite event for the degradation of Chls and LHCPs during senescence. Transient overexpression of Sgr in Nicotiana benthamiana and an in vivo pull-down assay show that Sgr interacts with LHCPII, indicating that the Sgr-LHCPII complexes are formed in the thylakoid membranes. Thus, we propose that in senescing leaves, Sgr regulates Chl degradation by inducing LHCPII disassembly through direct interaction, leading to the degradation of Chls and Chl-free LHCPII by catabolic enzymes and proteases, respectively.

Rice contains several MADS box genes. It has been demonstrated previously that one of these genes, OsMADS1 (for Oryza sativa MADS box gene1 ), is expressed preferentially in flowers and causes early flowering when ectopically expressed in tobacco plants. In this study, we demonstrated that ectopic expression of OsMADS1 in rice also results in early flowering. To further investigate the role of OsMADS1 during rice flower development, we generated transgenic rice plants expressing altered OsMADS1 genes that contain missense mutations in the MADS domain. There was no visible alteration in the transgenic plants during the vegetative stage. However, transgenic panicles typically exhibited phenotypic alterations, including spikelets consisting of elongated leafy paleae and lemmas that exhibit a feature of open hull, two pairs of leafy palea-like and lemma-like lodicules, a decrease in stamen number, and an increase in the number of carpels. In addition, some spikelets generated an additional floret from the same rachilla. These characteristics are very similar to those of leafy hull sterile1 ( lhs1 ). The map position of OsMADS1 is closely linked to that of lhs1 on chromosome 3. Examination of lhs1 revealed that it contains two missense mutations in the OsMADS1 MADS domain. A genetic complementation experiment showed that the 11.9-kb genomic DNA fragment containing the wild-type OsMADS1 gene rescued the mutant phenotypes. In addition, ectopic expression of the OsMADS1 gene isolated from the lhs1 line resulted in lhs1 -conferred phenotypes. These lines of evidence demonstrate that OsMADS1 is the lhs1 gene. INTRODUCTIONIn response to floral induction, the inflorescence meristem becomes committed to flowering. LEAFY ( LFY ) and APE-TALA1 ( AP1 ) in Arabidopsis and FLORICAULA ( FLO ) and SQUAMOSA ( SQUA ) in Antirrhinum are responsible for promoting the specification of floral meristem identity (reviewed in Ma, 1994). The genes required for specifying the fate of floral organ primordia include AP1 , AP2 , AGAMOUS ( AG ), PISTILATA ( PI ), and AP3 in Arabidopsis and SQUA , PLENA ( PLE ), GLOBOSA ( GLO ), and DEFICIENS ( DEF ) in Antirrhinum (reviewed in Weigel and Meyerowitz, 1994). Excluding AP2 , these floral homeotic genes encode MADS box proteins that are highly conserved transcription factors in plants, animals, yeast, and fungi and that are regulated by the floral meristem identity gene LFY (Parcy et al., 1998;Wagner et al., 1999).Several other MADS box genes have more subtle functions associated with floral meristem and floral organ identity. Expression of AG-LIKE2 ( AGL2 ), AGL4 , and AGL9 of Arabidopsis begins after the onset of expression of floral meristem identity genes but before the activation of floral organ identity genes (Flanagan and Ma, 1994;Savidge et al., 1995;Mandel and Yanofsky, 1998). DEFH72 and DEFH200 of Antirrhinum appear to function in mediating interactions between the meristem and organ identity genes through direct interaction with PLE (Davies et al., 1996). FLORAL BINDING PROTEIN2 ( FBP2 ) o...

Positive phototaxis systems have been well studied in bacteria; however, the photoreceptor(s) and their downstream signaling components that are responsible for negative phototaxis are poorly understood. Negative phototaxis sensory systems are important for cyanobacteria, oxygenic photosynthetic organisms that must contend with reactive oxygen species generated by an abundance of pigment photosensitizers. The unicellular cyanobacterium Synechocystis sp. PCC6803 exhibits type IV pilus-dependent negative phototaxis in response to unidirectional UV-A illumination. Using a reverse genetic approach, together with biochemical, molecular genetic, and RNA expression profiling analyses, we show that the cyanobacteriochrome locus ( slr1212/uirS ) of Synechocystis and two adjacent response regulator loci ( slr1213/uirR and the PatA-type regulator slr1214/lsiR ) encode a UV-A–activated signaling system that is required for negative phototaxis. We propose that UirS, which is membrane-associated via its ETR1 domain, functions as a UV-A photosensor directing expression of lsiR via release of bound UirR, which targets the lsiR promoter. Constitutive expression of LsiR induces negative phototaxis under conditions that normally promote positive phototaxis. Also induced by other stresses, LsiR thus integrates light inputs from multiple photosensors to determine the direction of movement.

To understand the transcriptional regulatory mechanism of host genes during the activation of defense responses in rice, we isolated WRKY transcription factors whose expressions were altered upon attack of the fungal pathogen Magnaporthe grisea, the causal agent of the devastating rice blast disease. A systematic expression analysis of OsWRKYs (Oryza sativa L. WRKYs) revealed that among 45 tested genes the expression of 15 genes was increased remarkably in an incompatible interaction between rice and M. grisea. Twelve of the M. grisea-inducible OsWRKY genes were also differentially regulated in rice plants infected with the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo). In experiments with defense signaling molecules, the expression of two genes, OsWRKY45 and OsWRKY62, was increased in salicylic acid (SA)-treated leaves and the expression of three genes, OsWRKY10, OsWRKY82, and OsWRKY85 was increased by jasmonic acid (JA) treatment. OsWRKY30 and OsWRKY83 responded to both SA- and JA treatments. The expression profiles suggest that a large number of WRKY DNA-binding proteins are involved in the transcriptional activation of defense-related genes in response to rice pathogens.