Allopolyploid hybridization serves as a major pathway for plant evolution, but in its early stages it is associated with phenotypic and genomic instabilities that are poorly understood. We have investigated allopolyploidization between Arabidopsis thaliana (2 n ‫؍‬ 2 x ‫؍‬ 10; n , gametic chromosome number; x , haploid chromosome number) and Cardaminopsis arenosa (2 n ‫؍‬ 4 x ‫؍‬ 32). The variable phenotype of the allotetraploids could not be explained by cytological abnormalities. However, we found suppression of 20 of the 700 genes examined by amplified fragment length polymorphism of cDNA. Independent reverse transcription-polymerase chain reaction analyses of 10 of these 20 genes confirmed silencing in three of them, suggesting that ‫ف‬ 0.4% of the genes in the allotetraploids are silenced. These three silenced genes were characterized. One, called K7, is repeated and similar to transposons. Another is RAP2.1 , a member of the large APETALA2 (AP2) gene family, and has a repeated element upstream of its 5 Ј end. The last, L6, is an unknown gene close to ALCOHOL DEHYDROGENASE on chromosome 1. CNG DNA methylation of K7 was less in the allotetraploids than in the parents, and the element varied in copy number. That K7 could be reactivated suggests epigenetic regulation. L6 was methylated in the C. arenosa genome. The present evidence that gene silencing accompanies allopolyploidization opens new avenues to this area of research.

Centromeric H3-like histones, which replace histone H3 in the centromeric chromatin of animals and fungi, have not been reported in plants. We identified a histone H3 variant from Arabidopsis thaliana that encodes a centromere-identifying protein designated HTR12. By immunological detection, HTR12 localized at centromeres in both mitotic and meiotic cells. HTR12 signal revealed tissue-and stage-specific differences in centromere morphology, including a distended bead-like structure in interphase root tip cells. The anti-HTR12 antibody also detected spherical organelles in meiotic cells. Although the antibody does not label centromeres in the closely related species Arabidopsis arenosa , HTR12 signal was found on all centromeres in allopolyploids of these two species. Comparison of the HTR12 genes of A. thaliana and A. arenosa revealed striking adaptive evolution in the N-terminal tail of the protein, similar to the pattern seen in its counterpart in Drosophila . This finding suggests that the same evolutionary forces shape centromeric chromatin in both animals and plants. INTRODUCTIONCentromeres are the specialized chromosomal sites necessary for poleward movement during mitosis and meiosis in eukaryotes. Commonly, a centromere is evident as a prominent constriction within the heterochromatin of each metaphase chromosome. The attachment to and movement of chromosomes along the spindle is mediated by the proteinaceous kinetochores, which form at the centromeres during cell division.Despite this highly conserved function, centromeric DNA sequences are not conserved between organisms. For example, human centromeres consist of large blocks (200 kb to several megabases) of tandemly repeated 171-bp ␣ -satellite (Willard, 1998), but the sequences can differ from those of apes on homologous chromosomes (Haaf and Willard, 1997). Similarly, Drosophila melanogaster centromeric regions contain blocks of 5-to 12-bp satellite repeats that do not appear to be shared by homologous centromeres of sibling species (Lohe and Brutlag, 1987).Plant centromeric regions resemble their mammalian counterparts in that both have large arrays of tandem repeats, frequently of approximately nucleosomal size. In centromere regions of maize (Alfenito and Birchler, 1993;Ananiev et al., 1998), pearl millet (Kamm et al., 1994), rice (Dong et al., 1998), sugarcane (Nagaki et al., 1998), sorghum (Zwick et al., 2000), the Australian daisy Brachycome dichromosomatica (Leach et al., 1995), rape (Xia et al., 1993), and the wild beet Beta procumbens (Gindullis et al., 2001), tandem repeats have been found that differ in sequence but that all have lengths in the range of ‫ف‬ 140 to 180 bp. Particular repeat arrays in cereals have been estimated to be hundreds of kilobases long (Dong et al., 1998;Kaszás and Birchler, 1998). Cereal tandem repeat arrays are interrupted by TY3/gypsy -like retrotransposons and other retrotransposons (Ananiev et al., 1998;Kumekawa et al., 2001;Nonomura and Kurata, 2001). The satellite arrays and interspersed retrotransposons o...

SummaryPolyploids are common and arise frequently by genome duplication (autopolyploids) or interspecific hybridization (allopolyploids). Neoallopolyploids display sterility, lethality, phenotypic instability, gene silencing and epigenetic changes. Little is known about the molecular basis of these phenomena, and how much genomic remodeling happens upon allopolyploidization. Extensive genomic remodeling has been documented in wheat, but little remodeling occurs in cotton. Newly synthesized Arabidopsis allopolyploids, which display phenotypic instability and low fertility, displayed several, possibly related mechanisms that can remodel genomes. We detected transcriptional activity of several transposons although their transposition was limited. One represents a new family of conditionally active En-Spm-like transposons of Arabidopsis thaliana, which underwent remodeling of CG methylation upon allopolyploidization. A random amplified fragment length polymorphism survey suggested remodeling at few, specific loci. Meiotic analyses revealed the appearance of chromosomal fragments in a substantial fraction of anther meiocytes. In several individuals produced by hybrids between the synthetic and a natural allopolyploid pollen viability inversely correlated with meiotic instability. Activity of selected DNA transposons and the possibly related chromosomal breaks could cause changes by inducing translocations and rearrangements.

Allopolyploid hybridization serves as a major pathway for plant evolution, but in its early stages it is associated with phenotypic and genomic instabilities that are poorly understood. We have investigated allopolyploidization between Arabidopsis thaliana (2n = 2x = 10; n, gametic chromosome number; x, haploid chromosome number) and Cardaminopsis arenosa (2n = 4x = 32). The variable phenotype of the allotetraploids could not be explained by cytological abnormalities. However, we found suppression of 20 of the 700 genes examined by amplified fragment length polymorphism of cDNA. Independent reverse transcription-polymerase chain reaction analyses of 10 of these 20 genes confirmed silencing in three of them, suggesting that approximately 0.4% of the genes in the allotetraploids are silenced. These three silenced genes were characterized. One, called K7, is repeated and similar to transposons. Another is RAP2.1, a member of the large APETALA2 (AP2) gene family, and has a repeated element upstream of its 5' end. The last, L6, is an unknown gene close to ALCOHOL DEHYDROGENASE on chromosome 1. CNG DNA methylation of K7 was less in the allotetraploids than in the parents, and the element varied in copy number. That K7 could be reactivated suggests epigenetic regulation. L6 was methylated in the C. arenosa genome. The present evidence that gene silencing accompanies allopolyploidization opens new avenues to this area of research.