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The nitrate reductase (NR)-defective double mutant of Arabidopsis thaliana (nia1 nia2) has previously been shown to present a low endogenous content of NO in its leaves compared with the wild-type plants. In the present study, we analyzed the effect of NR mutation on floral induction and development of A. thaliana, as NO was recently described as one of the signals involved in the flowering process. The NO fluorescent probes diaminofluorescein-2 diacetate (DAF-2DA) and 1,2-diaminoanthraquinone (1,2-DAA) were used to localize NO production in situ by fluorescence microscopy in the floral structures of A. thaliana during floral development. Data were validated by incubating the intact tissues with DAF-2 and quantifying the DAF-2 triazole by fluorescence spectrometry. The results showed that NO is synthesized in specific cells and tissues in the floral structure and its production increases with floral development until anthesis. In the gynoecium, NO synthesis occurs only in differentiated stigmatic papillae of the floral bud, and, in the stamen, only anthers that are producing pollen grains synthesize NO. Sepals and petals do not show NO production. NR-deficient plants emitted less NO, although they showed the same pattern of NO emission in their floral organs. This mutant blossomed precociously when compared with wild-type plants, as measured by the increased caulinar/rosette leaf number and the decrease in the number of days to bolting and anthesis, and this phenotype seems to result from the markedly reduced NO levels in roots and leaves during vegetative growth. Overall, the results reveal a role for NR in the flowering process.

In the present study, we investigated the involvement of the mitochondrial permeability transition pore (PTP) in nitric oxide (NO)-induced plant cell death. NO donors such as sodium nitroprusside (SNP) and S-nitroso-N-acetylpenicillamine inhibited growth and caused death in suspension-cultured cells of Citrus sinensis. Cells treated with SNP showed chromatin condensation and fragmentation, characteristic of apoptosis. SNP caused loss of the mitochondrial membrane electrical potential, which was prevented by cyclosporin A (CsA), a specific inhibitor of PTP formation. CsA also prevented the nuclear apoptosis and subsequent Citrus cell death induced by NO. These findings indicate that mitochondrial PTP formation is involved in the signaling pathway by which NO induces apoptosis in cultured Citrus cells. ß 2002 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.

The genus Coffea L. has around 100 native species distributed in tropical and subtropical areas in Africa, and the most important economic species are C. arabica and C. canephora. C. arabica is exceptional in the genus since it is the only species so far analyzed which is self-compatible, and a natural polyploid with 2n=4x=44 chromosomes; it is considered to be a segmental allopolyploid because it presents a disomic inheritance and a regular meiotic behavior. All other species in the genus are selfincompatible diploids with 2n=2x=22. Cytogenetic studies in Coffea, undertaken since 1912, have followed various phases: initial studies were limited only to establishing chromosome counts. Subsequent studies characterized the karyotypes of various species using conventional cytological techniques. As the somatic metaphase chromosomes of coffee are very small (1 -3 μm) and morphologically symmetrical, these studies resulted in uniform karyotypes that show almost no differences among species. Since genetic improvement of coffee trees has progressed mainly by means of interspecific hybridizations involving wild species, analyses of microsporogenesis in species and hybrids were needed to establish their genetic affinity and relationships. The first successful attempts to differentiate coffee chromosomes longitudinally were made by mapping pachytene chromomeric patterns and by C and NOR banding techniques. From 1998 onwards, the use of banding techniques with the fluorochromes DAPI and CMA 3, and also the cytomolecular technique FISH using rDNA probes, has increased the longitudinal differentiation of coffee chromosomes. The use of the GISH technique with total genomic DNA has revealed the parental species that originated C. arabica species. Key words: Coffea, chromosomal characterization, karyotype, meiotic behavior, chromomere pattern, chromosome banding, molecular cytogenetics. Citogenética de café (Coffea L.):O gênero Coffea L., com cerca de 100 espécies nativas de regiões tropicais e subtropicais da África, tem as espécies Coffea arabica e C. canephora como comercialmente importantes. C. arabica é uma exceção no gênero por ser o único poliplóide natural com 2n=4x=44 cromossomos e autocompatível sendo considerado um alopoliplóide segmental devido à herança dissômica e comportamento meiótico regular. As demais espécies são diplóides autoincompatíveis com 2n=2x=22. Estudos citogenéticos em Coffea têm sido realizados, desde 1912, com diferentes finalidades. Os primeiros visaram apenas a contagem de cromossomos. Em seguida, estudos morfológicos em cromossomos somáticos com técnicas citológicas convencionais objetivaram a caracterização das diferentes espécies. Devido o fato dos cromossomos de café serem relativamente pequenos (1 -3 μm), e morfologicamente simétricos, estes estudos revelaram cariótipos homogêneos entre as diferentes espécies. O melhoramento genético do cafeeiro, realizado através de hibridações interespecífcas envolvendo espécies selvagens diplóides, demandou estudos da microsporogênese, em espécies e híbri...

Two diploid (2n=22 chromosomes) and self-incompatible coffee species, Coffea canephora Pierre ex Froehner and C. dewevrei De Wild, et Th. Dur., were subjected to 2% acetic orcein and C-banding techniques with the purpose of mitotic chromosome characterizations. Karyological analysis were done in root cell chromosomes. These two diploid species of coffee could be distinguished mainly by some chromosome absolute (u,m) and relative (%) lenght mean values. In both species chromosome 5, 10 and 11 were metacentrics and the others were submetacentries; chromosomes 1 and 3 had secondary constriction beside centromere. The data suggested chromosome 3 had the NOR. Haploid karyotype formula was 1M + 2m + 7sm + lsm s for C. canephora, and 3m + 7sm + lsm s for C. dewevrei. Total karyotype length mean values were 16.97 ± 0.95u,m for C. canephora and 15.60± l.ll|imfor C. dewev rei and the difference was not significant. C-banded karyotypes were very similar in both species with prevailing centromeric/pericentromeric C-bands. The total C-band heterochromatin values expressed in percentage were 15.76 + 0.32% for C. canephora and 16.11 ± 0.26% for C. dewevrei. These values were not significantly different to separate both species. Similarly, no distinction could be made on karyotype asymmetry index mean values between the two species.

SUMMARY -A detailed morphological analysis of Coffea arabica pachytene chromosomes is presented for the first time. The pachytene chromomeric patterns revealed that there occurs some structural similarities in about 54% of the 22 bivalents. This fact is supported by two secondary associations observed, one of them involving one nucleolar bivalent. Based on these results, and other evidences from literature, it is suggested that C. arabica is a segmental alIopolyploid with genetically controlled chromosome pairing.

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Despite the recent advances in plant population genetic studies, the lack of information regarding pedigree, ploidy level, or mode of inheritance for many polyploids can compromise the analysis of the molecular data produced. The aim of this study was to examine both microsatellite and cytogenetic characteristics of the pioneer tree Croton floribundus Spreng. (Euphorbiaceae) to test for the occurrence of polyploidy in the species and to evaluate its implications for the appropriate use of SSR markers. Seven microsatellite markers were developed and screened for 62 individuals from a semi-deciduous tropical forest in Brazil. Chromosome number, meiotic behavior, and pollen viability were evaluated from male flower buds. All SSR loci were highly polymorphic. The number of bivalents observed in meiosis n = 56 (2n = 8× = 112) and the maximum number of alleles per individual (Ni = 8) demonstrated the occurrence of polyploidy in C. floribundus. The normal meiotic pairing and the high pollen viability suggested that C. floribundus is a regular and stable polyploid, most likely an allopolyploid. The combined SSR and cytogenetic data provided new evidence on the origin and evolution of the species as well as assured the accurate use of SSR loci for population genetic studies of the polyploid pioneer species.