51 research outputs found
Integrating the Genetic and Physical Maps of Arabidopsis thaliana: Identification of Mapped Alleles of Cloned Essential (EMB) Genes
The classical genetic map of Arabidopsis includes more than 130 genes with an embryo-defective (emb) mutant phenotype. Many of these essential genes remain to be cloned. Hundreds of additional EMB genes have been cloned and catalogued (www.seedgenes.org) but not mapped. To facilitate EMB gene identification and assess the current level of saturation, we updated the classical map, compared the physical and genetic locations of mapped loci, and performed allelism tests between mapped (but not cloned) and cloned (but not mapped) emb mutants with similar chromosome locations. Two hundred pairwise combinations of genes located on chromosomes 1 and 5 were tested and more than 1100 total crosses were screened. Sixteen of 51 mapped emb mutants examined were found to be disrupted in a known EMB gene. Alleles of a wide range of published EMB genes (YDA, GLA1, TIL1, AtASP38, AtDEK1, EMB506, DG1, OEP80) were discovered. Two EMS mutants isolated 30 years ago, T-DNA mutants with complex insertion sites, and a mutant with an atypical, embryo-specific phenotype were resolved. The frequency of allelism encountered was consistent with past estimates of 500 to 1000 EMB loci. New EMB genes identified among mapped T-DNA insertion mutants included CHC1, which is required for chromatin remodeling, and SHS1/AtBT1, which encodes a plastidial nucleotide transporter similar to the maize Brittle1 protein required for normal endosperm development. Two classical genetic markers (PY, ALB1) were identified based on similar map locations of known genes required for thiamine (THIC) and chlorophyll (PDE166) biosynthesis. The alignment of genetic and physical maps presented here should facilitate the continued analysis of essential genes in Arabidopsis and further characterization of a broad spectrum of mutant phenotypes in a model plant
The Tnt1 Retrotransposon Escapes Silencing in Tobacco, Its Natural Host
Retrotransposons' high capacity for mutagenesis is a threat that genomes need to control tightly. Transcriptional gene silencing is a general and highly effective control of retrotransposon expression. Yet, some retrotransposons manage to transpose and proliferate in plant genomes, suggesting that, as shown for plant viruses, retrotransposons can escape silencing. However no evidence of retrotransposon silencing escape has been reported. Here we analyze the silencing control of the tobacco Tnt1 retrotransposon and report that even though constructs driven by the Tnt1 promoter become silenced when stably integrated in tobacco, the endogenous Tnt1 elements remain active. Silencing of Tnt1-containing transgenes correlates with high DNA methylation and the inability to incorporate H2A.Z into their promoters, whereas the endogenous Tnt1 elements remain partially methylated at asymmetrical positions and incorporate H2A.Z upon induction. Our results show that the promoter of Tnt1 is a target of silencing in tobacco, but also that endogenous Tnt1 elements can escape this control and be expressed in their natural host
Arabidopsis HDA6 Regulates Locus-Directed Heterochromatin Silencing in Cooperation with MET1
Heterochromatin silencing is pivotal for genome stability in eukaryotes. In
Arabidopsis, a plant-specific mechanism called
RNA–directed DNA methylation (RdDM) is involved in heterochromatin
silencing. Histone deacetylase HDA6 has been identified as a component of such
machineries; however, its endogenous targets and the silencing mechanisms have
not been analyzed globally. In this study, we investigated the silencing
mechanism mediated by HDA6. Genome-wide transcript profiling revealed that the
loci silenced by HDA6 carried sequences corresponding to the RDR2-dependent
24-nt siRNAs, however their transcript levels were mostly unaffected in the
rdr2 mutant. Strikingly, we observed significant overlap of
genes silenced by HDA6 to those by the CG DNA methyltransferase MET1.
Furthermore, regardless of dependence on RdDM pathway, HDA6 deficiency resulted
in loss of heterochromatic epigenetic marks and aberrant enrichment for
euchromatic marks at HDA6 direct targets, along with ectopic expression of these
loci. Acetylation levels increased significantly in the hda6
mutant at all of the lysine residues in the H3 and H4 N-tails, except H4K16.
Interestingly, we observed two different CG methylation statuses in the
hda6 mutant. CG methylation was sustained in the
hda6 mutant at some HDA6 target loci that were surrounded
by flanking DNA–methylated regions. In contrast, complete loss of CG
methylation occurred in the hda6 mutant at the HDA6 target loci
that were isolated from flanking DNA methylation. Regardless of CG methylation
status, CHG and CHH methylation were lost and transcriptional derepression
occurred in the hda6 mutant. Furthermore, we show that HDA6
binds only to its target loci, not the flanking methylated DNA, indicating the
profound target specificity of HDA6. We propose that HDA6 regulates
locus-directed heterochromatin silencing in cooperation with MET1, possibly
recruiting MET1 to specific loci, thus forming the foundation of silent
chromatin structure for subsequent non-CG methylation
Nomad DNA — A model for movement and duplication of DNA sequences in plant genomes
Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/43425/1/11103_2004_Article_BF00019160.pd
Real-Time Lineage Analysis to Study Cell Division Orientation in the Arabidopsis Shoot Meristem
Cells in the Arabidopsis shoot apical meristem are small and divide frequently throughout the life-time of the organism making them good candidates for studying the mechanisms of cell division in plants. But tracking these cell divisions requires multiple images to be taken of the same specimen over time which means the specimen must stay alive throughout the process. This chapter provides details on how to prepare plants for live imaging, keep them alive and growing through multiple time points, and how to process the data to extract cell boundary coordinates from three-dimensional images
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