Mutations in the predicted DNA polymerase subunit POLD3 result in more rapid flowering of Brachypodium distachyon.

The timing of reproduction is a critical developmental decision in the life cycle of many plant species. Fine mapping of a rapid-flowering mutant was done using whole-genome sequence data from bulked DNA from a segregating F2 mapping populations. The causative mutation maps to a gene orthologous with the third subunit of DNA polymerase δ (POLD3), a previously uncharacterized gene in plants.

Establishment of a vernalization requirement in requires .

A requirement for vernalization, the process by which prolonged cold exposure provides competence to flower, is an important adaptation to temperate climates that ensures flowering does not occur before the onset of winter. In temperate grasses, vernalization results in the up-regulation of () to establish competence to flower; however, little is known about the mechanism underlying repression of in the fall season, which is necessary to establish a vernalization requirement. Here, we report that a plant-specific gene containing a bromo-adjacent homology and transcriptional elongation factor S-II domain, which we named (), represses before vernalization in That is upstream of is supported by the observations that is precociously elevated in an mutant, resulting in rapid flowering without cold exposure, and the rapid-flowering phenotype is dependent on The precocious expression in is associated with reduced levels of the repressive chromatin modification H3K27me3 at , which is similar to the reduced H3K27me3 in vernalized plants.

Subunit compositions of Arabidopsis RNA polymerases I and III reveal Pol I- and Pol III-specific forms of the AC40 subunit and alternative forms of the C53 subunit.

Using affinity purification and mass spectrometry, we identified the subunits of Arabidopsis thaliana multisubunit RNA polymerases I and III (abbreviated as Pol I and Pol III), the first analysis of their physical compositions in plants. In all eukaryotes examined to date, AC40 and AC19 subunits are common to Pol I (a.k.

PHYTOCHROME C is an essential light receptor for photoperiodic flowering in the temperate grass, Brachypodium distachyon.

We show that in the temperate grass, Brachypodium distachyon, PHYTOCHROME C (PHYC), is necessary for photoperiodic flowering. In loss-of-function phyC mutants, flowering is extremely delayed in inductive photoperiods. PHYC was identified as the causative locus by utilizing a mapping by sequencing pipeline (Cloudmap) optimized for identification of induced mutations in Brachypodium.

Memory of the vernalized state in plants including the model grass Brachypodium distachyon.

Plant species that have a vernalization requirement exhibit variation in the ability to "remember" winter - i.e., variation in the stability of the vernalized state.

Interaction of photoperiod and vernalization determines flowering time of Brachypodium distachyon.

Timing of flowering is key to the reproductive success of many plants. In temperate climates, flowering is often coordinated with seasonal environmental cues such as temperature and photoperiod. Vernalization is an example of temperature influencing the timing of flowering and is defined as the process by which a prolonged exposure to the cold of winter results in competence to flower during the following spring.

Intersection of small RNA pathways in Arabidopsis thaliana sub-nuclear domains.

In Arabidopsis thaliana, functionally diverse small RNA (smRNA) pathways bring about decreased RNA accumulation of target genes via several different mechanisms. Cytological experiments have suggested that the processing of microRNAs (miRNAs) and heterochromatic small interfering RNAs (hc-siRNAs) occurs within a specific nuclear domain that can present Cajal Body (CB) characteristics. It is unclear whether single or multiple smRNA-related domains are found within the same CB and how specialization of the smRNA pathways is determined within this specific sub-compartment.

In vitro transcription activities of Pol IV, Pol V, and RDR2 reveal coupling of Pol IV and RDR2 for dsRNA synthesis in plant RNA silencing.

In Arabidopsis, RNA-dependent DNA methylation and transcriptional silencing involves three nuclear RNA polymerases that are biochemically undefined: the presumptive DNA-dependent RNA polymerases Pol IV and Pol V and the putative RNA-dependent RNA polymerase RDR2. Here we demonstrate their RNA polymerase activities in vitro. Unlike Pol II, Pols IV and V require an RNA primer, are insensitive to α-amanitin, and differ in their ability to displace the nontemplate DNA strand during transcription.

Functional consequences of subunit diversity in RNA polymerases II and V.

Multisubunit RNA polymerases IV and V (Pol IV and Pol V) evolved as specialized forms of Pol II that mediate RNA-directed DNA methylation (RdDM) and transcriptional silencing of transposons, viruses, and endogenous repeats in plants. Among the subunits common to Arabidopsis thaliana Pols II, IV, and V are 93% identical alternative ninth subunits, NRP(B/D/E)9a and NRP(B/D/E)9b. The 9a and 9b subunit variants are incompletely redundant with respect to Pol II; whereas double mutants are embryo lethal, single mutants are viable, yet phenotypically distinct.

RNA polymerase V transcription guides ARGONAUTE4 to chromatin.

Retrotransposons and repetitive DNA elements in eukaryotes are silenced by small RNA-directed heterochromatin formation. In Arabidopsis, this process involves 24-nt siRNAs that bind to ARGONAUTE4 (AGO4) and facilitate the targeting of complementary loci via unknown mechanisms. Nuclear RNA polymerase V (Pol V) is an RNA silencing enzyme recently shown to generate noncoding transcripts at loci silenced by 24-nt siRNAs.

Subunit compositions of the RNA-silencing enzymes Pol IV and Pol V reveal their origins as specialized forms of RNA polymerase II.

In addition to RNA polymerases I, II, and III, the essential RNA polymerases present in all eukaryotes, plants have two additional nuclear RNA polymerases, abbreviated as Pol IV and Pol V, that play nonredundant roles in siRNA-directed DNA methylation and gene silencing. We show that Arabidopsis Pol IV and Pol V are composed of subunits that are paralogous or identical to the 12 subunits of Pol II. Four subunits of Pol IV are distinct from their Pol II paralogs, six subunits of Pol V are distinct from their Pol II paralogs, and four subunits differ between Pol IV and Pol V.

Mitochondrial DNA transfer to the nucleus generates extensive insertion site variation in maize.

Mitochondrial DNA (mtDNA) insertions into nuclear chromosomes have been documented in a number of eukaryotes. We used fluorescence in situ hybridization (FISH) to examine the variation of mtDNA insertions in maize. Twenty overlapping cosmids, representing the 570-kb maize mitochondrial genome, were individually labeled and hybridized to root tip metaphase chromosomes from the B73 inbred line.

A test for ectopic exchange catalyzed by Cre recombinase in maize.

A maize line expressing Cre recombinase as well as the recipient line without the transgene were assayed for evidence of ectopic recombination within the maize genome. Such a test is valuable for understanding the action of Cre as well as for its use to recombine two target lox sites present in the chromosomes. Pollen examination and seed set tests of material expressing Cre provided no evidence of ectopic recombination, which would be manifested in the production of translocations or inversions and result in pollen abortion and reduced seed set.

Nonadditive gene expression in diploid and triploid hybrids of maize.

The molecular basis of hybrid vigor (heterosis) has remained unknown despite the importance of this phenomenon in evolution and in practical breeding programs. To formulate a molecular basis of heterosis, an understanding of gene expression in inbred and hybrid states is needed. In this study, we examined the amount of various transcripts in hybrid and inbred individuals (B73 and Mo17) to determine whether the quantities of specific messenger RNAs were additive or nonadditive in the hybrids.