Cryptochromes (CRYs) are photoreceptors that mediate light regulation of the circadian clock in plants and animals. Here we show that CRYs mediate blue-light regulation of N
-methyladenosine (m
A) ...modification of more than 10% of messenger RNAs in the Arabidopsis transcriptome, especially those regulated by the circadian clock. CRY2 interacts with three subunits of the METTL3/14-type N
-methyladenosine RNA methyltransferase (m
A writer): MTA, MTB and FIP37. Photo-excited CRY2 undergoes liquid-liquid phase separation (LLPS) to co-condense m
A writer proteins in vivo, without obviously altering the affinity between CRY2 and the writer proteins. mta and cry1cry2 mutants share common defects of a lengthened circadian period, reduced m
A RNA methylation and accelerated degradation of mRNA encoding the core component of the molecular oscillator circadian clock associated 1 (CCA1). These results argue for a photoregulatory mechanism by which light-induced phase separation of CRYs modulates m
A writer activity, mRNA methylation and abundance, and the circadian rhythms in plants.
Transposable elements (TEs) and repetitive sequences make up over 35% of the rice (Oryza sativa) genome. The host regulates the activity of different TEs by different epigenetic mechanisms, including ...DNA methylation, histone H3K9 methylation, and histone H3K4 demethylation. TEs can also affect the expression of host genes. For example, miniature inverted repeat TEs (MITEs), dispersed high copy-number DNA TEs, can influence the expression of nearby genes. In plants, 24-nt small interfering RNAs (siRNAs) are mainly derived from repeats and TEs. However, the extent to which TEs, particularly MITEs associated with 24-nt siRNAs, affect gene expression remains elusive. Here, we show that the rice Dicer-like 3 homolog OsDCL3a is primarily responsible for 24-nt siRNA processing. Impairing OsDCL3a expression by RNA interference caused phenotypes affecting important agricultural traits; these phenotypes include dwarfism, larger flag leaf angle, and fewer secondary branches. We used small RNA deep sequencing to identify 535,054 24-nt siRNA clusters. Of these clusters, ∼82% were OsDCL3a-dependent and showed significant enrichment of MITEs. Reduction of OsDCL3a function reduced the 24-nt siRNAs predominantly from MITEs and elevated expression of nearby genes. OsDCL3a directly targets genes involved in gibberellin and brassinosteroid homeostasis; OsDCL3a deficiency may affect these genes, thus causing the phenotypes of dwarfism and enlarged flag leaf angle. Our work identifies OsDCL3a-dependent 24-nt siRNAs derived from MITEs as broadly functioning regulators for fine-tuning gene expression, which may reflect a conserved epigenetic mechanism in higher plants with genomes rich in dispersed repeats or TEs.
SWI/SNF-type chromatin remodelers, such as BRAHMA (BRM), and H3K27 demethylases both have active roles in regulating gene expression at the chromatin level, but how they are recruited to specific ...genomic sites remains largely unknown. Here we show that RELATIVE OF EARLY FLOWERING 6 (REF6), a plant-unique H3K27 demethylase, targets genomic loci containing a CTCTGYTY motif via its zinc-finger (ZnF) domains and facilitates the recruitment of BRM. Genome-wide analyses showed that REF6 colocalizes with BRM at many genomic sites with the CTCTGYTY motif. Loss of REF6 results in decreased BRM occupancy at BRM-REF6 co-targets. Furthermore, REF6 directly binds to the CTCTGYTY motif in vitro, and deletion of the motif from a target gene renders it inaccessible to REF6 in vivo. Finally, we show that, when its ZnF domains are deleted, REF6 loses its genomic targeting ability. Thus, our work identifies a new genomic targeting mechanism for an H3K27 demethylase and demonstrates its key role in recruiting the BRM chromatin remodeler.
Nanopore sequencing from Oxford Nanopore Technologies (ONT) and Pacific BioSciences (PacBio) single-molecule real-time (SMRT) long-read isoform sequencing (Iso-Seq) are revolutionizing the way ...transcriptomes are analyzed. These methods offer many advantages over most widely used high-throughput short-read RNA sequencing (RNA-Seq) approaches and allow a comprehensive analysis of transcriptomes in identifying full-length splice isoforms and several other post-transcriptional events. In addition, direct RNA-Seq provides valuable information about RNA modifications, which are lost during the PCR amplification step in other methods. Here, we present a comprehensive summary of important applications of these technologies in plants, including identification of complex alternative splicing (AS), full-length splice variants, fusion transcripts, and alternative polyadenylation (APA) events. Furthermore, we discuss the impact of the newly developed nanopore direct RNA-Seq in advancing epitranscriptome research in plants. Additionally, we summarize computational tools for identifying and quantifying full-length isoforms and other co/post-transcriptional events and discussed some of the limitations with these methods. Sequencing of transcriptomes using these new single-molecule long-read methods will unravel many aspects of transcriptome complexity in unprecedented ways as compared to previous short-read sequencing approaches. Analysis of plant transcriptomes with these new powerful methods that require minimum sample processing is likely to become the norm and is expected to uncover novel co/post-transcriptional gene regulatory mechanisms that control biological outcomes during plant development and in response to various stresses.
Auxin is essential for plant growth and development. Although substantial progress has been made in understanding auxin pathways in model plants such as Arabidopsis and rice, little is known in moso ...bamboo which is famous for its fast growth resulting from the rapid cell elongation and division.
Here we showed that exogenous auxin has strong effects on crown and primary roots. Genes involved in auxin action, including 13 YUCCA (YUC) genes involved in auxin synthesis, 14 PIN-FORMED/PIN-like (PIN/PILS) and 7 AUXIN1/LIKE-AUX1 (AUX1/LAX) members involved in auxin transport, 10 auxin receptors (AFB) involved in auxin perception, 43 auxin/indole-3-aceticacid (AUX/IAA) genes, and 41 auxin response factors (ARF) involved in auxin signaling were identified through genome-wide analysis. Phylogenetic analysis of these genes from Arabidopsis, Oryza sativa and bamboo revealed that auxin biosynthesis, transport, and signaling pathways are conserved in these species. A comprehensive study of auxin-responsive genes using RNA sequencing technology was performed, and the results also supported that moso bamboo shared a conserved regulatory mechanism for the expression of auxin pathway genes; meanwhile it harbors its own specific properties.
In summary, we generated an overview of the auxin pathway in bamboo, which provides information for uncovering the precise roles of auxin pathway in this important species in the future.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Alternative splicing (AS) is a major post‐transcriptional mechanism to enhance the diversity of proteome in response to environmental signals. Among the numerous external signals perceived by plants, ...light is the most crucial one. Plants utilize complex photoreceptor signaling networks to sense different light conditions and adjust their growth and development accordingly. Although light‐mediated gene expression has been widely investigated, little is known regarding the mechanism of light affecting AS to modulate mRNA at the post‐transcriptional level. In this minireview, we summarize current progresses on how light affects AS, and how sensory photoreceptors and retrograde signaling pathways may coordinately regulate AS of pre‐mRNAs. In addition, we also discuss the possibility that AS of the mRNAs encoding photoreceptors may be involved in feedback control of AS. We hypothesize that light regulation of the expression and activity of splicing factors would be a major mechanism of light‐mediated AS. The combination of genetic study and high‐throughput analyses of AS and splicing complexes in response to light is likely to further advance our understanding of the molecular mechanisms underlying light control of AS and plant development.
In plant, light affects alternative splicing mainly via three primary types of sensory photoreceptor pathways or retrograde signals from chloroplast, while photoreceptors themselves are also under the regulation of alternative splicing. The light regulation of splicing factors would be a major mechanism of light‐mediated alternative splicing that represents functions on dominant negative regulation, subcellular localization, or taking part in nonsense‐mediated mRNA decay pathway.
The process of RNA splicing influences many physiological processes, including plant immunity. However, how plant parasites manipulate host RNA splicing process remains unknown. Here we demonstrate ...that PsAvr3c, an avirulence effector from oomycete plant pathogen Phytophthora sojae, physically binds to and stabilizes soybean serine/lysine/arginine-rich proteins GmSKRPs. The SKRPs are novel proteins that associate with a complex that contains plant spliceosome components, and are negative regulators of plant immunity. Analysis by RNA-seq data indicates that alternative splicing of pre-mRNAs from 401 soybean genes, including defense-related genes, is altered in GmSKRP1 and PsAvr3c overexpressing lines compared to control plants. Representative splicing events mediated by GmSKRP1 and PsAvr3c are tested by infection assays or by transient expression in soybean plants. Our results show that plant pathogen effectors can reprogram host pre-mRNA splicing to promote disease, and we propose that pathogens evolved such strategies to defeat host immune systems.
Aspartic proteases (APs) are a class of aspartic peptidases belonging to nine proteolytic enzyme families whose members are widely distributed in biological organisms. APs play essential functions ...during plant development and environmental adaptation. However, there are few reports about APs in fast-growing moso bamboo.
In this study, we identified a total of 129 AP proteins (PhAPs) encoded by the moso bamboo genome. Phylogenetic and gene structure analyses showed that these 129 PhAPs could be divided into three categories (categories A, B and C). The PhAP gene family in moso bamboo may have undergone gene expansion, especially the members of categories A and B, although homologs of some members in category C have been lost. The chromosomal location of PhAPs suggested that segmental and tandem duplication events were critical for PhAP gene expansion. Promoter analysis revealed that PhAPs in moso bamboo may be involved in plant development and responses to environmental stress. Furthermore, PhAPs showed tissue-specific expression patterns and may play important roles in rapid growth, including programmed cell death, cell division and elongation, by integrating environmental signals such as light and gibberellin signals.
Comprehensive analysis of the AP gene family in moso bamboo suggests that PhAPs have experienced gene expansion that is distinct from that in rice and may play an important role in moso bamboo organ development and rapid growth. Our results provide a direction and lay a foundation for further analysis of plant AP genes to clarify their function during rapid growth.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Accurate and comprehensive annotation of transcript sequences is essential for transcript quantification and differential gene and transcript expression analysis. Single-molecule long-read sequencing ...technologies provide improved integrity of transcript structures including alternative splicing, and transcription start and polyadenylation sites. However, accuracy is significantly affected by sequencing errors, mRNA degradation, or incomplete cDNA synthesis.
We present a new and comprehensive Arabidopsis thaliana Reference Transcript Dataset 3 (AtRTD3). AtRTD3 contains over 169,000 transcripts-twice that of the best current Arabidopsis transcriptome and including over 1500 novel genes. Seventy-eight percent of transcripts are from Iso-seq with accurately defined splice junctions and transcription start and end sites. We develop novel methods to determine splice junctions and transcription start and end sites accurately. Mismatch profiles around splice junctions provide a powerful feature to distinguish correct splice junctions and remove false splice junctions. Stratified approaches identify high-confidence transcription start and end sites and remove fragmentary transcripts due to degradation. AtRTD3 is a major improvement over existing transcriptomes as demonstrated by analysis of an Arabidopsis cold response RNA-seq time-series. AtRTD3 provides higher resolution of transcript expression profiling and identifies cold-induced differential transcription start and polyadenylation site usage.
AtRTD3 is the most comprehensive Arabidopsis transcriptome currently. It improves the precision of differential gene and transcript expression, differential alternative splicing, and transcription start/end site usage analysis from RNA-seq data. The novel methods for identifying accurate splice junctions and transcription start/end sites are widely applicable and will improve single-molecule sequencing analysis from any species.
Nuclear-localized RNA binding proteins are involved in various aspects of RNA metabolism, which in turn modulates gene expression. However, the functions of nuclear-localized RNA binding proteins in ...plants are poorly understood. Here, we report the functions of two proteins containing RNA recognition motifs, RZ-1B and RZ-1C, in Arabidopsis thaliana. RZ-1B and RZ-1C were localized to nuclear speckles and interacted with a spectrum of serine/arginine-rich (SR) proteins through their C termini. RZ-1C preferentially bound to purine-rich RNA sequences in vitro through its N-terminal RNA recognition motif. Disrupting the RNA binding activity of RZ-1C with SR proteins through overexpression of the C terminus of RZ-1C conferred defective phenotypes similar to those observed in rz-1b rz-1c double mutants, including delayed seed germination, reduced stature, and serrated leaves. Loss of function of RZ-1B and RZ-1C was accompanied by defective splicing of many genes and global perturbation of gene expression. In addition, we found that RZ-1C directly targeted FLOWERING LOCUS C (FLC), promoting efficient splicing of FLC introns and likely also repressing FLC transcription. Our findings highlight the critical role of RZ-1B/1C in regulating RNA splicing, gene expression, and many key aspects of plant development via interaction with proteins including SR proteins.
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