111 research outputs found
The FACT Histone Chaperone: Tuning Gene Transcription in the Chromatin Context to Modulate Plant Growth and Development
FACT is a heterodimeric histone chaperone consisting of the SSRP1 and SPT16 proteins and is conserved among eukaryotes. It interacts with the histones H2A-H2B and H3-H4 as well as with DNA. Based on in vitro and in vivo studies mainly in yeast and mammalian cells, FACT can mediate nucleosome disassembly and reassembly and thus facilitates in the chromatin context DNA-dependent processes including transcription, replication and repair. In plants, primarily the role of FACT related to RNA polymerase II transcription has been examined. FACT was found to associate with elongating Arabidopsis RNA polymerase II (RNAPII) as part of the transcript elongation complex and it was identified as repressor of aberrant intragenic transcriptional initiation. Arabidopsis mutants depleted in FACT subunits exhibit various defects in vegetative and reproductive development. Strikingly, FACT modulates important developmental transitions by promoting expression of key repressors of these processes. Thus, FACT facilitates expression of DOG1 and FLC adjusting the switch from seed dormancy to germination and from vegetative to reproductive development, respectively. In the central cell of the female gametophyte, FACT can facilitate DNA demethylation especially within heterochromatin, and thereby contributes to gene imprinting during Arabidopsis reproduction. This review discusses results particularly from the plant perspective about the contribution of FACT to processes that involve reorganisation of nucleosomes with a main focus on RNAPII transcription and its implications for diverse areas of plant biology
Plant Proteins Containing High Mobility Group Box DNA-Binding Domains Modulate Different Nuclear Processes
Transcript elongation by RNA polymerase II in plants: factors, regulation and impact on gene expression
Transcriptional elongation by RNA polymerase II (RNAPII) through chromatin is a dynamic and highly regulated step of eukaryotic gene expression. A combination of transcript elongation factors (TEFs) including modulators of RNAPII activity and histone chaperones facilitate efficient transcription on nucleosomal templates. Biochemical and genetic analyses, primarily performed in Arabidopsis, provided insight into the contribution of TEFs to establish gene expression patterns during plant growth and development. In addition to summarising the role of TEFs in plant gene expression, we emphasise in our review recent advances in the field. Thus, mechanisms are presented how aberrant intragenic transcript initiation is suppressed by repressing transcriptional start sites within coding sequences. We also discuss how transcriptional interference of ongoing transcription with neighbouring genes is prevented. Moreover, it appears that plants make no use of promoter-proximal RNAPII pausing in the way mammals do, but there are nucleosome-defined mechanism(s) that determine the efficiency of mRNA synthesis by RNAPII. Accordingly, a still growing number of processes related to plant growth, development and responses to changing environmental conditions prove to be regulated at the level of transcriptional elongation
Critical Role of Transcript Cleavage in Arabidopsis RNA Polymerase II Transcriptional Elongation
Transcript elongation factors associate with elongating RNA polymerase II (RNAPII) to control the efficiency of mRNA synthesis and consequently modulate plant growth and development. Encountering obstacles during transcription such as nucleosomes or particular DNA sequences may cause backtracking and transcriptional arrest of RNAPII. The elongation factor TFIIS stimulates the intrinsic transcript cleavage activity of the polymerase, which is required for efficient rescue of backtracked/arrested RNAPII. A TFIIS mutant variant (TFIISmut) lacks the stimulatory activity to promote RNA cleavage, but instead efficiently inhibits unstimulated transcript cleavage by RNAPII. We could not recover viable Arabidopsis (Arabidopsis thaliana) tfIIs plants constitutively expressing TFIISmut. Induced, transient expression of TFIISmut in tfIIs plants provoked severe growth defects, transcriptomic changes and massive, transcription-related redistribution of elongating RNAPII within transcribed regions toward the transcriptional start site. The predominant site of RNAPII accumulation overlapped with the 11 nucleosome, suggesting that upon inhibition of RNA cleavage activity, RNAPII arrest prevalently occurs at this position. In the presence of TFIISmut, the amount of RNAPII was reduced, which could be reverted by inhibiting the proteasome, indicating proteasomal degradation of arrested RNAPII. Our findings suggest that polymerase backtracking/arrest frequently occurs in plant cells, and RNAPII-reactivation is essential for correct transcriptional output and proper growth/development
Different elongation factors distinctly modulate RNA polymerase II transcription in Arabidopsis
Various transcript elongation factors (TEFs) including modulators of RNA polymerase II (RNAPII) activity and histone chaperones tune the efficiency of transcription in the chromatin context. TEFs are involved in establishing gene expression patterns during growth and development in Arabidopsis, while little is known about the genomic distribution of the TEFs and the way they facilitate transcription. We have mapped the genome-wide occupancy of the elongation factors SPT4–SPT5, PAF1C and FACT, relative to that of elongating RNAPII phosphorylated at residues S2/S5 within the carboxyterminal domain. The distribution of SPT4–SPT5 along transcribed regions closely resembles that of RNAPII-S2P, while the occupancy of FACT and PAF1C is rather related to that of RNAPII-S5P. Under transcriptionally challenging heat stress conditions, mutant plants lacking the corresponding TEFs are differentially impaired in transcript synthesis. Strikingly, in plants deficient in PAF1C, defects in transcription across intron/exon borders are observed that are cumulative along transcribed regions. Upstream of transcriptional start sites, the presence of FACT correlates with nucleosomal occupancy. Under stress conditions FACT is particularly required for transcriptional upregulation and to promote RNAPII transcription through +1 nucleosomes. Thus, Arabidopsis TEFs are differently distributed along transcribed regions, and are distinctly required during transcript elongation especially upon transcriptional reprogramming
Structural insights into the mechanism of negative regulation of single-box high mobility group proteins by the acidic tail domain.
The Drosophila and plant (maize) functional counterparts of the abundant vertebrate chromosomal protein HMGB1 (HMG-D and ZmHMGB1, respectively) differ from HMGB1 in having a single HMG box, as well as basic and acidic flanking regions that vary greatly in length and charge. We show that despite these variations, HMG-D and ZmHMGB1 exist in dynamic assemblies in which the basic HMG boxes and linkers associate with their intrinsically disordered, predominantly acidic, tails in a manner analogous to that observed previously for HMGB1. The DNA-binding surfaces of the boxes and linkers are occluded in "auto-inhibited" forms of the protein, which are in equilibrium with transient, more open structures that are "binding-competent." This strongly suggests that the mechanism of auto-inhibition may be a general one. HMG-D and ZmHMGB1 differ from HMGB1 in having phosphorylation sites in their tail and linker regions. In both cases, in vitro phosphorylation of serine residues within the acidic tail stabilizes the assembled form, suggesting another level of regulation for interaction with DNA, chromatin, and other proteins that is not possible for the uniformly acidic (hence unphosphorylatable) tail of HMGB1.This work was supported by the Biotechnology and Biological Sciences Research Council through the award of Grant BB/D002257/1 (to J. O. T.) and a grant from the Deutsche Forschungsgemeinschaft (DFG) (to K. D. G.).This is the final published version. It first appeared at http://www.jbc.org/content/289/43/29817.long
Distinct role of subunits of the Arabidopsis RNA polymerase II elongation factor PAF1C in transcriptional reprogramming
Transcript elongation by RNA polymerase II (RNAPII) is dynamic and highly regulated, thereby contributing to the implementation of gene expression programs during plant development or in response to environmental cues. The heterohexameric polymerase-associated factor 1 complex (PAF1C) stabilizes the RNAPII elongation complex promoting efficient transcript synthesis. In addition, PAF1C links transcriptional elongation with various post-translational histone modifications at transcribed loci. We have exposed
Arabidopsis mutants deficient in the PAF1C subunits ELF7 or CDC73 to elevated NaCl concentrations to provoke a transcriptional response. The growth of elf7 plants was reduced relative to that of wildtype under these challenging
conditions, whereas cdc73 plants exhibited rather enhanced tolerance. Profiling of the transcriptional changes upon NaCl exposure revealed that cdc73 responded similar to wildtype. Relative to wildtype and cdc73, the transcriptional response of elf7 plants was severely reduced in accord with their greater susceptibility to NaCl. The data also imply that CDC73 is more
relevant for the transcription of longer genes. Despite the fact that both ELF7 and CDC73 are part of PAF1C the strikingly different transcriptional response of the mutants upon NaCl exposure suggests that the subunits have (partially) specific functions
Histone 2B monoubiquitination complex integrates transcript elongation with RNA processing at circadian clock and flowering regulators
Altres ajuts: CERCA Programme/Generalitat de CatalunyaHISTONE MONOUBIQUITINATION1 (HUB1) and its paralog HUB2 act in a conserved heterotetrameric complex in the chromatin-mediated transcriptional modulation of developmental programs, such as flowering time, dormancy, and the circadian clock. The KHD1 and SPEN3 proteins were identified as interactors of the HUB1 and HUB2 proteins with in vitro RNA-binding activity. Mutants in SPEN3 and KHD1 had reduced rosette and leaf areas. Strikingly, in spen3 mutants, the flowering time was slightly, but significantly, delayed, as opposed to the early flowering time in the hub1-4 mutant. The mutant phenotypes in biomass and flowering time suggested a deregulation of their respective regulatory genes CIRCADIAN CLOCK-ASSOCIATED1 (CCA1) and FLOWERING LOCUS C (FLC) that are known targets of the HUB1-mediated histone H2B monoubiquitination (H2Bub). Indeed, in the spen3-1 and hub1-4 mutants, the circadian clock period was shortened as observed by luciferase reporter assays, the levels of the CCA1α and CCA1β splice forms were altered, and the CCA1 expression and H2Bub levels were reduced. In the spen3-1 mutant, the delay in flowering time was correlated with an enhanced FLC expression, possibly due to an increased distal versus proximal ratio of its antisense COOLAIR transcript. Together with transcriptomic and double-mutant analyses, our data revealed that the HUB1 interaction with SPEN3 links H2Bub during transcript elongation with pre-mRNA processing at CCA1. Furthermore, the presence of an intact HUB1 at the FLC is required for SPEN3 function in the formation of the FLC-derived antisense COOLAIR transcripts
The Arabidopsis condensin CAP‐D subunits arrange interphase chromatin
Condensins are best known for their role in shaping chromosomes. Other functions such as organizing interphase chromatin and transcriptional control have been reported in yeasts and animals, but little is known about their function in plants. To elucidate the specific composition of condensin complexes and the expression of CAP-D2 (condensin I) and CAP-D3 (condensin II), we performed biochemical analyses in Arabidopsis. The role of CAP-D3 in interphase chromatin organization and function was evaluated using cytogenetic and transcriptome analysis in cap-d3 T-DNA insertion mutants. CAP-D2 and CAP-D3 are highly expressed in mitotically active tissues. In silico and pull-down experiments indicate that both CAP-D proteins interact with the other condensin I and II subunits. In cap-d3 mutants, an association of heterochromatic sequences occurs, but the nuclear size and the general histone and DNA methylation patterns remain unchanged. Also, CAP-D3 influences the expression of genes affecting the response to water, chemicals, and stress. The expression and composition of the condensin complexes in Arabidopsis are similar to those in other higher eukaryotes. We propose a model for the CAP-D3 function during interphase in which CAP-D3 localizes in euchromatin loops to stiffen them and consequently separates centromeric regions and 45S rDNA repeats
Chromatin-associated HMGA and HMGB proteins: versatile co-regulators of DNA-dependent processes
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