Plant lncRNAs Research Articles

Genome-wide identification and characterization of novel lncRNAs in Ginkgo biloba

We identified several novel lncRNAs that play important roles in Ginkgo biloba leaf development. A large number of long non-coding RNAs (lncRNAs) have emerged as important regulators of many biological processes in animals and plants. However, lncRNAs and their regulatory roles remain poorly characterized in woody plants, particularly in the gymnosperm G. biloba. We performed deep strand-specific RNA sequencing (ssRNA-seq), and obtained 27.44 GB raw data from all major developmental stages of G. biloba leaves. From these sequencing data, we identified 1323 novel lncRNAs. These lncRNAs were transcribed from 947 scaffolds of G. biloba, 57.6% of which came from intergenic regions. We also confirmed that Gb-lncRNAs were shorter and had fewer exons than protein-coding genes. Target prediction for 764 lncRNAs identified 1184 target genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation analysis revealed that these lncRNAs might be associated with photosynthesis, plant hormones, and secondary metabolism in leaves. Moreover, we constructed a network of regulatory interactions between lncRNAs and mRNAs; 89 lncRNAs were predicted as targets of 64 miRNAs and 33 lncRNAs functioned as miRNA precursors. Quantitative real-time PCR revealed that novel lncRNAs had stage- and tissue-specific expression patterns in G. biloba leaves. These results provide new insight into the function of lncRNAs in leaves and enhance our understanding of plant lncRNAs.

Reproductive Long Intergenic Noncoding RNAs Exhibit Male Gamete Specificity and Polycomb Repressive Complex 2-Mediated Repression.

Long noncoding RNAs (lncRNAs) have been characterized extensively in animals and are involved in several processes, including homeobox gene expression and X-chromosome inactivation. In comparison, there has been much less detailed characterization of plant lncRNAs, and the number of distinct lncRNAs encoded in plant genomes and their regulation by developmental and epigenetic mechanisms remain largely unknown. Here, we analyzed transcriptome data from Asian rice (Oryza sativa) and identified 6,309 long intergenic noncoding RNAs (lincRNAs), focusing on their expression in reproductive tissues and organs. Most O. sativa lincRNAs were expressed in a highly tissue-specific manner, with an unexpectedly high fraction specifically expressed in male gametes. Mutation of a component of the Polycomb Repressive Complex2 (PRC2) resulted in derepression of another large class of lincRNAs, whose expression is correlated with H3K27 trimethylation in developing panicles. Overlap with the sperm cell-specific lincRNAs suggests that epigenetic repression of lincRNAs in the panicles was partially relieved in the male germline. Expression of a subset of lincRNAs also showed modulation by drought in reproductive tissues. Comparison with other cereal genomes showed that the lincRNAs generally have low levels of conservation at both the sequence and structural levels. Use of a novelty detection support vector machine model enabled the detection of nucleotide sequence and structural homology in ∼10% and ∼4% of the lincRNAs in genomes of purple false brome (Brachypodium distachyon) and maize (Zea mays), respectively. This is the first study to report on a large number of lncRNAs that are targets of repression by PRC2 rather than mediating regulation via PRC2. That the vast majority of the lincRNAs reported here do not overlap with those of other rice studies indicates that these are a significant addition to the known lincRNAs in rice.

Long non-coding RNAs of switchgrass (Panicum virgatum L.) in multiple dehydration stresses

BackgroundLong non-coding RNAs (lncRNAs) play important roles in plant growth and stress responses. Studies of lncRNAs in non-model plants are quite limited, especially those investigating multiple dehydration stresses. In this study, we identified novel lncRNAs and analyzed their functions in dehydration stress memory in switchgrass, an excellent biofuel feedstock and soil-conserving plant in the Gramineae family.ResultsWe analyzed genome-wide transcriptional profiles of leaves of 5-week-old switchgrass plantlets grown via tissue culture after primary and secondary dehydration stresses (D1 and D2) and identified 16,551 novel lncRNAs, including 4554 annotated lncRNAs (targeting 3574 genes), and 11,997 unknown lncRNAs. Gene ontology and pathway enrichment analysis of annotated genes showed that the differentially expressed lncRNAs were related to abscisic acid (ABA) and ethylene (ETH) biosynthesis and signal transduction, and to starch and sucrose metabolism. The upregulated lncRNAs and genes were related to ABA synthesis and its signal transduction, and to trehalose synthesis. Meanwhile, lncRNAs and genes related to ETH biosynthesis and signal transduction were suppressed. LncRNAs and genes involved in ABA metabolism were verified using quantitative real-time PCR, and the endogenous ABA content was determined via high performance liquid chromatography mass spectrometry (HPLC-MS). These results showed that ABA accumulated significantly during dehydration stress, especially in D2. Furthermore, we identified 307 dehydration stress memory lncRNAs, and the ratios of different memory types in switchgrass were similar to those in Arabidopsis and maize.ConclusionsThe molecular responses of switchgrass lncRNAs to multiple dehydration stresses were researched systematically, revealing novel information about their transcriptional regulatory behavior. This study provides new insights into the response mechanism to dehydration stress in plants. The lncRNAs and pathways identified in this study provide valuable information for genetic modification of switchgrass and other crops.

Prediction of plant lncRNA by ensemble machine learning classifiers

BackgroundIn plants, long non-protein coding RNAs are believed to have essential roles in development and stress responses. However, relative to advances on discerning biological roles for long non-protein coding RNAs in animal systems, this RNA class in plants is largely understudied. With comparatively few validated plant long non-coding RNAs, research on this potentially critical class of RNA is hindered by a lack of appropriate prediction tools and databases. Supervised learning models trained on data sets of mostly non-validated, non-coding transcripts have been previously used to identify this enigmatic RNA class with applications largely focused on animal systems. Our approach uses a training set comprised only of empirically validated long non-protein coding RNAs from plant, animal, and viral sources to predict and rank candidate long non-protein coding gene products for future functional validation.ResultsIndividual stochastic gradient boosting and random forest classifiers trained on only empirically validated long non-protein coding RNAs were constructed. In order to use the strengths of multiple classifiers, we combined multiple models into a single stacking meta-learner. This ensemble approach benefits from the diversity of several learners to effectively identify putative plant long non-coding RNAs from transcript sequence features. When the predicted genes identified by the ensemble classifier were compared to those listed in GreeNC, an established plant long non-coding RNA database, overlap for predicted genes from Arabidopsis thaliana, Oryza sativa and Eutrema salsugineum ranged from 51 to 83% with the highest agreement in Eutrema salsugineum. Most of the highest ranking predictions from Arabidopsis thaliana were annotated as potential natural antisense genes, pseudogenes, transposable elements, or simply computationally predicted hypothetical protein. Due to the nature of this tool, the model can be updated as new long non-protein coding transcripts are identified and functionally verified.ConclusionsThis ensemble classifier is an accurate tool that can be used to rank long non-protein coding RNA predictions for use in conjunction with gene expression studies. Selection of plant transcripts with a high potential for regulatory roles as long non-protein coding RNAs will advance research in the elucidation of long non-protein coding RNA function.

Classification and experimental identification of plant long non-coding RNAs

Our understanding of gene regulation is constantly evolving. It is now clear that the majority of cellular transcripts are non-coding RNAs. The spectrum of non-coding RNAs is diverse and includes short (<200 nt) and long non-coding RNAs (lncRNAs) (>200 nt). LncRNAs regulate gene expression through diverse mechanisms. In this review, we describe the emerging roles of lncRNA mediated plant gene regulation. We discuss the current classification of lncRNAs and their role in genome organization and gene regulation. We also highlight a subset of lncRNAs that are epigenetic regulators of plant gene expression. Lastly, we provide an overview of emerging techniques and databases that are employed for the identification and characterization of plant lncRNAs.

CRISPR/Cas9-mediated mutagenesis of lncRNA1459 alters tomato fruit ripening.

With the development of high-throughput sequencing, many long non-coding RNAs (lncRNAs) have been found to play important roles in diverse biological processes. However, the biological functions of most plant lncRNAs are still unknown. We have previously discovered a tomato ripening-related lncRNA, lncRNA1459. Here, we cloned the full-length lncRNA1459, giving two transcript isoforms. In addition, lncRNA1459 exhibited a specific location in the nucleus. Furthermore, in order to fully identify the function of lncRNA1459 in tomato ripening, loss-of-function mutants of lncRNA1459 were developed using clustered regularly interspaced short palindromic repeats (CRISPR)/-associated protein 9 (Cas9)-induced genome editing technology. Compared with wild-type fruits, the tomato ripening process was significantly repressed in lncRNA1459 mutants. Ethylene production and lycopene accumulation were largely repressed in lncRNA1459 mutants. Additionally, genes related to ethylene and carotenoid biosynthesis were distinctly downregulated in lncRNA1459 mutants compared with wild-type fruits. Moreover, expression of numerous ripening-related genes was changed significantly when lncRNA1459 was knocked out. Expression of potential tomato ripening-related lncRNAs was also specifically changed after knocking out lncRNA1459. Taken together, these results provide insight into the role of lncRNA1459 in tomato fruit ripening.

Genome-Wide Identification and Characterization of Long Non-Coding RNA in Wheat Roots in Response to Ca2+ Channel Blocker.

It remains unclear whether plant lncRNAs are responsive to Ca2+-channel blocking. When using the Ca2+-channel blocker, LaCl3, to treat germinated wheat seeds for 24 h, we found that both root length and mitosis were inhibited in the LaCl3-treated groups. The effect of the Ca2+-channel blocker was verified in three ways: a [Ca2+]cyt decrease detected using Fluo-3/AM staining, a decrease in the Ca content measured using inductively coupled plasma mass spectrometry, and an inhibition of Ca2+ influx detected using Non-invasive Micro-test Technology. Genome-wide high throughput RNA-seq and bioinformatical methods were used to identify lncRNAs, and found 177 differentially expressed lncRNAs that might be in responsive to Ca2+-channel blocking. Among these, 108 were up-regulated and 69 were down-regulated. The validity of identified lncRNAs data from RNA-seq was verified using qPCR. GO and KEGG analysis indicated that a number of lncRNAs might be involved in diverse biological processes upon Ca2+-channel blocking. Further GO analysis showed that 23 lncRNAs might play roles as transcription factor (TF); Moreover, eight lncRNAs might participate in cell cycle regulation, and their relative expressions were detected using qPCR. This study also provides diverse data on wheat lncRNAs that can deepen our understanding of the function and regulatory mechanism of Ca2+-channel blocking in plants.

Stress-responsive regulation of long non-coding RNA polyadenylation in Oryza sativa.

Recently, long non-coding RNAs (lncRNAs) have been demonstrated to be involved in many biological processes of plants; however, a systematic study on transcriptional and, in particular, post-transcriptional regulation of stress-responsive lncRNAs in Oryza sativa (rice) is lacking. We sequenced three types of RNA libraries (poly(A)+, poly(A)- and nuclear RNAs) under four abiotic stresses (cold, heat, drought and salt). Based on an integrative bioinformatics approach and ~200 high-throughput data sets, ~170 of which have been published, we revealed over 7000 lncRNAs, nearly half of which were identified for the first time. Notably, we found that the majority of the ~500 poly(A) lncRNAs that were differentially expressed under stress were significantly downregulated, but approximately 25% were found to have upregulated non-poly(A) forms. Moreover, hundreds of lncRNAs with downregulated polyadenylation (DPA) tend to be highly conserved, show significant nuclear retention and are co-expressed with protein-coding genes that function under stress. Remarkably, these DPA lncRNAs are significantly enriched in quantitative trait loci (QTLs) for stress tolerance or development, suggesting their potential important roles in rice growth under various stresses. In particular, we observed substantially accumulated DPA lncRNAs in plants exposed to drought and salt, which is consistent with the severe reduction of RNA 3'-end processing factors under these conditions. Taken together, the results of this study reveal that polyadenylation and subcellular localization of many rice lncRNAs are likely to be regulated at the post-transcriptional level. Our findings strongly suggest that many upregulated/downregulated lncRNAs previously identified by traditional RNA-seq analyses need to be carefully reviewed to assess the influence of post-transcriptional modification.

Novel phosphate deficiency-responsive long non-coding RNAs in the legume model plant Medicago truncatula.

Emerging evidence indicates that long non-coding RNAs (lncRNAs) play important roles in the regulation of many biological processes. Inhibition of plant growth due to deficiency in soil inorganic phosphate (Pi) occurs widely across natural and agricultural ecosystems; however, we know little about the function of plant lncRNAs in response to Pi deficiency. To address this issue, we first identified 10 785 lncRNAs in the legume model species Medicago truncatula by sequencing eight strand-specific libraries. Out of these lncRNAs, 358 and 224 were responsive to Pi deficiency in the leaves and roots, respectively. We further predicted and classified the putative targets of those lncRNAs and the results revealed that they may be involved in the processes of signal transduction, energy synthesis, detoxification, and Pi transport. Finally, we functionally characterized three Phosphate Deficiency-Induced LncRNAs (PDILs) using their corresponding Tnt1 mutants. The results showed that PDIL1 suppressed degradation of MtPHO2, which encodes a ubiquitin-conjugating E2 enzyme regulated by miR399, while PDIL2 and PDIL3 directly regulated Pi transport at the transcriptional level. These findings demonstrate that PDILs can regulate Pi-deficiency signaling and Pi transport, highlighting the involvement of lncRNAs in the regulation of responses of plants to Pi deficiency.

Genome-wide identification of miRNAs and lncRNAs in Cajanus cajan

BackgroundNon-coding RNAs (ncRNAs) are important players in the post transcriptional regulation of gene expression (PTGR). On one hand, microRNAs (miRNAs) are an abundant class of small ncRNAs (~22nt long) that negatively regulate gene expression at the levels of messenger RNAs stability and translation inhibition, on the other hand, long ncRNAs (lncRNAs) are a large and diverse class of transcribed non-protein coding RNA molecules (> 200nt) that play both up-regulatory as well as down-regulatory roles at the transcriptional level. Cajanus cajan, a leguminosae pulse crop grown in tropical and subtropical areas of the world, is a source of high value protein to vegetarians or very poor populations globally. Hence, genome-wide identification of miRNAs and lncRNAs in C. cajan is extremely important to understand their role in PTGR with a possible implication to generate improve variety of crops.ResultsWe have identified 616 mature miRNAs in C. cajan belonging to 118 families, of which 578 are novel and not reported in MirBase21. A total of 1373 target sequences were identified for 180 miRNAs. Of these, 298 targets were characterized at the protein level. Besides, we have also predicted 3919 lncRNAs. Additionally, we have identified 87 of the predicted lncRNAs to be targeted by 66 miRNAs.ConclusionsmiRNA and lncRNAs in plants are known to control a variety of traits including yield, quality and stress tolerance. Owing to its agricultural importance and medicinal value, the identified miRNA, lncRNA and their targets in C. cajan may be useful for genome editing to improve better quality crop. A thorough understanding of ncRNA-based cellular regulatory networks will aid in the improvement of C. cajan agricultural traits.

PLncPRO for prediction of long non-coding RNAs (lncRNAs) in plants and its application for discovery of abiotic stress-responsive lncRNAs in rice and chickpea.

Long non-coding RNAs (lncRNAs) make up a significant portion of non-coding RNAs and are involved in a variety of biological processes. Accurate identification/annotation of lncRNAs is the primary step for gaining deeper insights into their functions. In this study, we report a novel tool, PLncPRO, for prediction of lncRNAs in plants using transcriptome data. PLncPRO is based on machine learning and uses random forest algorithm to classify coding and long non-coding transcripts. PLncPRO has better prediction accuracy as compared to other existing tools and is particularly well-suited for plants. We developed consensus models for dicots and monocots to facilitate prediction of lncRNAs in non-model/orphan plants. The performance of PLncPRO was quite better with vertebrate transcriptome data as well. Using PLncPRO, we discovered 3714 and 3457 high-confidence lncRNAs in rice and chickpea, respectively, under drought or salinity stress conditions. We investigated different characteristics and differential expression under drought/salinity stress conditions, and validated lncRNAs via RT-qPCR. Overall, we developed a new tool for the prediction of lncRNAs in plants and showed its utility via identification of lncRNAs in rice and chickpea.

Identification and characterization of novel lncRNAs in Arabidopsis thaliana

Long noncoding RNAs (lncRNAs) are important regulators of various biological processes, but few studies have identified lncRNAs in plants; genome-wide discovery of novel lncRNAs is thus required. We used deep strand-specific sequencing (ssRNA-seq) to obtain approximately 62 million reads from all developmental stages of Arabidopsis thaliana and identified 156 novel lncRNAs that we classified according to their localization. These novel identified lncRNAs showed low expression levels and sequence conservation. Bioinformatic analysis predicted potential target genes or cis-regulated genes of 91 antisense and 32 intergenic lncRNAs. Functional annotation of these potential targets and sequence motif analysis indicated that the lncRNAs participate in various biological processes underlying Arabidopsis growth and development. Seventeen of the lncRNAs were predicted targets of 22 miRNAs, and a network of interactions between ncRNAs and mRNAs was constructed. In addition, nine lncRNAs functioned as miRNA precursors. Finally, qRT-PCR revealed that novel lncRNAs have stage- and tissue-specific expression patterns in A. thaliana. Our study provides insight into the potential functions and regulatory interactions of novel Arabidopsis lncRNAs, and enhances our understanding of plant lncRNAs, which will facilitate functional research.

Present Scenario of Long Non-Coding RNAs in Plants.

Small non-coding RNAs have been extensively studied in plants over the last decade. In contrast, genome-wide identification of plant long non-coding RNAs (lncRNAs) has recently gained momentum. LncRNAs are now being recognized as important players in gene regulation, and their potent regulatory roles are being studied comprehensively in eukaryotes. LncRNAs were first reported in humans in 1992. Since then, research in animals, particularly in humans, has rapidly progressed, and a vast amount of data has been generated, collected, and organized using computational approaches. Additionally, numerous studies have been conducted to understand the roles of these long RNA species in several diseases. However, the status of lncRNA investigation in plants lags behind that in animals (especially humans). Efforts are being made in this direction using computational tools and high-throughput sequencing technologies, such as the lncRNA microarray technique, RNA-sequencing (RNA-seq), RNA capture sequencing, (RNA CaptureSeq), etc. Given the current scenario, significant amounts of data have been produced regarding plant lncRNAs, and this amount is likely to increase in the subsequent years. In this review we have documented brief information about lncRNAs and their status of research in plants, along with the plant-specific resources/databases for information retrieval on lncRNAs.

Long Noncoding RNAs in Plants.

The eukaryotic genomes are pervasively transcribed. In addition to protein-coding RNAs, thousands of long noncoding RNAs (lncRNAs) modulate key molecular and biological processes. Most lncRNAs are found in the nucleus and associate with chromatin, but lncRNAs can function in both nuclear and cytoplasmic compartments. Emerging work has found that many lncRNAs regulate gene expression and can affect genome stability and nuclear domain organization both in plant and in the animal kingdom. Here, we describe the major plant lncRNAs and how they act, with a focus on research in Arabidopsis thaliana and our emerging understanding of lncRNA functions in serving as molecular sponges and decoys, functioning in regulation of transcription and silencing, particularly in RNA-directed DNA methylation, and in epigenetic regulation of flowering time.

식물의 긴비암호화 RNA들의 생물학적 기능

차세대 염기서열 분석기술의 발달로 대량의 전사수준의 단위체들이 발견되었는데, 이것들 중 대부분의 전사체들은 비암호화 RNA들이다. 이들 중 긴 비암호화 RNA (lncRNA)들은 200개 이상의 뉴클레오티드를 가지며 단백질로 번역이 되지 않는 기능적 RNA 분자들이다. 식물의 lncRNA들은 RNA Pol II, Pol III, Pol IV, Pol V에 의해 전사체가 만들어지고, 전사 후 이들 lncRNA들은 추가적인 splicing과 polyadenylation 과정이 일어난다. 식물의 lncRNA들은 그 발현수준이 매우 낮고, 조직 특이적으로 발현 되지만, 이들 lncRNA들은 외부자극에 의해 강한 발현이 유도된다. 환경스트레스를 포함한 각기 다른 외부자극에 의해 많은 식물 lncRNA들의 발현이 유도되었기 때문에 이들 lncRNA들은 식물의 다양한 생물학적 기능과 식물의 생장발달 과정에 있어 새로운 조절 인자로 고려되고 있다. 특히 후성유전학적인 유전자 억제, 크로마틴 변형, 타겟모방, 광형태형성, 단백질 재배치, 환경스트레스 반응, 병원균 감염등에 관련하여 기능을 한다. 또한 어떤 lncRNA들은 short RNA들의 전구체로 역할도 한다. 최근 식물에서 많은 lncRNA들이 분리 동정되었지만, 이들 lncRNA들의 생리학적인 기능에 대한 현재의 이해는 여전히 제한적이고, 이들의 세부적인 조절 메커니즘 연구는 지속적으로 이루어져야 할 것이다. 이 총설에서는 식물 lncRNA들의 생합성 및 조절 메커니즘과 현재까지 밝혀진 분자수준에서의 중요한 기능들을 요약 정리하였다. With the development of next generation sequencing (NGS), large numbers of transcriptional molecules have been discovered. Most transcripts are non -coding RNAs (ncRNAs). Among them, long non-coding RNAs (lncRNAs) with more than 200 nucleotides represent functional RNA molecule that will not be translated into protein. In plants, lncRNAs are transcribed by RNA polymerase II (Pol II) or Pol III, Pol VI and Pol V. After transcription of these lncRNAs, more RNA processing mechanisms such as splicing and polyadenylation occurs. The expression of plant lncRNAs is very low and is tissue specific. However, these lncRNAs are strongly induced by specific external stimuli. Because different external stimuli including environmental stresses induce a large number of plant lncRNAs, these lncRNAs have been gradually considered as new regulatory factors of various biological and development processes such as epigenetic repression, chromatin modification, target mimicry, photomorphogenesis, protein relocalization, environmental stress response, pathogen infection in plants. Moreover, some lncRNAs act as precursor of short RNAs. Although a large number of lncRNAs have been predicted and identified in plants, our current understanding of the biological function of these lncRNAs is still limited and their detailed regulatory mechanisms should be elucidated continuously. Here, we reviewed the biogenesis and regulation mechanisms of lncRNAs and summarized the molecular functions unraveled in plants.

Genome-wide identification of potato long intergenic noncoding RNAs responsive to Pectobacterium carotovorum subspecies brasiliense infection.

BackgroundLong noncoding RNAs (lncRNAs) represent a class of RNA molecules that are implicated in regulation of gene expression in both mammals and plants. While much progress has been made in determining the biological functions of lncRNAs in mammals, the functional roles of lncRNAs in plants are still poorly understood. Specifically, the roles of long intergenic nocoding RNAs (lincRNAs) in plant defence responses are yet to be fully explored.ResultsIn this study, we used strand-specific RNA sequencing to identify 1113 lincRNAs in potato (Solanum tuberosum) from stem tissues. The lincRNAs are expressed from all 12 potato chromosomes and generally smaller in size compared to protein-coding genes. Like in other plants, most potato lincRNAs possess single exons. A time-course RNA-seq analysis between a tolerant and a susceptible potato cultivar showed that 559 lincRNAs are responsive to Pectobacterium carotovorum subsp. brasiliense challenge compared to mock-inoculated controls. Moreover, coexpression analysis revealed that 17 of these lincRNAs are highly associated with 12 potato defence-related genes.ConclusionsTogether, these results suggest that lincRNAs have potential functional roles in potato defence responses. Furthermore, this work provides the first library of potato lincRNAs and a set of novel lincRNAs implicated in potato defences against P. carotovorum subsp. brasiliense, a member of the soft rot Enterobacteriaceae phytopathogens.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2967-9) contains supplementary material, which is available to authorized users.

Population genomic analysis of gibberellin-responsive long non-coding RNAs in Populus.

Long non-coding RNAs (lncRNAs) participate in a wide range of biological processes, but lncRNAs in plants remain largely unknown; in particular, we lack a systematic identification of plant lncRNAs involved in hormone responses. Moreover, allelic variation in lncRNAs remains poorly characterized at a large scale. Here, we conducted high-throughput RNA-sequencing of leaves from control and gibberellin (GA)-treated Populus tomentosa and identified 7655 reliably expressed lncRNAs. Among the 7655 lncRNAs, the levels of 410 lncRNAs changed in response to GA. Seven GA-responsive lncRNAs were predicted to be putative targets of 18 miRNAs, and one GA-responsive lncRNA (TCONS_00264314) was predicted to be a target mimic of ptc-miR6459b. Computational analysis predicted 939 potential cis-regulated target genes and 965 potential trans-regulated target genes for GA-responsive lncRNAs. Functional annotation of these potential target genes showed that they participate in many different biological processes, including auxin signal transduction and synthesis of cellulose and pectin, indicating that GA-responsive lncRNAs may influence growth and wood properties. Finally, single nucleotide polymorphism (SNP)-based association analysis showed that 112 SNPs from 52 GA-responsive lncRNAs and 1014 SNPs from 296 potential target genes were significantly associated with growth and wood properties. Epistasis analysis also provided evidence for interactions between lncRNAs and their potential target genes. Our study provides a comprehensive view of P. tomentosa lncRNAs and offers insights into the potential functions and regulatory interactions of GA-responsive lncRNAs, thus forming the foundation for future functional analysis of GA-responsive lncRNAs in P. tomentosa.

Molecular functions of long noncoding transcripts in plants

Long noncoding RNAs (lncRNAs) are transcribed from the genomes of various eukaryotes. LncRNAs are potent regulators of various developmental processes and respond to external stimuli. Recent studies have shown that lncRNAs function as potent cis- and trans-regulators of gene expression and as modular scaffolds of chromatin-modifying complexes. Several plant lncRNAs transcribed in response to various external signals and stresses have been identified. However, only a few of these lncRNAs have been characterized, and information regarding their regulatory mechanisms is limited. Here, we describe our current understanding of the biological functions of plant lncRNAs.

Targeting Non-Coding RNAs in Plants with the CRISPR-Cas Technology is a Challenge yet Worth Accepting.

Non-coding RNAs (ncRNAs) have emerged as versatile master regulator of biological functions in recent years. MicroRNAs (miRNAs) are small endogenous ncRNAs of 18–24 nucleotides in length that originates from long self-complementary precursors. Besides their direct involvement in developmental processes, plant miRNAs play key roles in gene regulatory networks and varied biological processes. Alternatively, long ncRNAs (lncRNAs) are a large and diverse class of transcribed ncRNAs whose length exceed that of 200 nucleotides. Plant lncRNAs are transcribed by different RNA polymerases, showing diverse structural features. Plant lncRNAs also are important regulators of gene expression in diverse biological processes. There has been a breakthrough in the technology of genome editing, the CRISPR-Cas9 (clustered regulatory interspaced short palindromic repeats/CRISPR-associated protein 9) technology, in the last decade. CRISPR loci are transcribed into ncRNA and eventually form a functional complex with Cas9 and further guide the complex to cleave complementary invading DNA. The CRISPR-Cas technology has been successfully applied in model plants such as Arabidopsis and tobacco and important crops like wheat, maize, and rice. However, all these studies are focused on protein coding genes. Information about targeting non-coding genes is scarce. Hitherto, the CRISPR-Cas technology has been exclusively used in vertebrate systems to engineer miRNA/lncRNAs, but it is still relatively unexplored in plants. While briefing miRNAs, lncRNAs and applications of the CRISPR-Cas technology in human and animals, this review essentially elaborates several strategies to overcome the challenges of applying the CRISPR-Cas technology in editing ncRNAs in plants and the future perspective of this field.

GREENC: a Wiki-based database of plant lncRNAs.

Long non-coding RNAs (lncRNAs) are functional non-translated molecules greater than 200 nt. Their roles are diverse and they are usually involved in transcriptional regulation. LncRNAs still remain largely uninvestigated in plants with few exceptions. Experimentally validated plant lncRNAs have been shown to regulate important agronomic traits such as phosphate starvation response, flowering time and interaction with symbiotic organisms, making them of great interest in plant biology and in breeding. There is still a lack of lncRNAs in most sequenced plant species, and in those where they have been annotated, different methods have been used, so making the lncRNAs less useful in comparisons within and between species. We developed a pipeline to annotate lncRNAs and applied it to 37 plant species and six algae, resulting in the annotation of more than 120 000 lncRNAs. To facilitate the study of lncRNAs for the plant research community, the information gathered is organised in the Green Non-Coding Database (GreeNC, http://greenc.sciencedesigners.com/).