PRL1, an RNA-Binding Protein, Positively Regulates the Accumulation of miRNAs and siRNAs in Arabidopsis

The evolutionary conserved WD-40 protein PRL1 plays important roles in immunity and development. Here we show that PRL1 is required for the accumulation of microRNAs (miRNAs) and small interfering RNAs (siRNAs). PRL1 positively influences the processing of miRNA primary transcripts (pri-miRNAs) and double-stranded RNAs (dsRNAs). Furthermore, PRL1 interacts with the pri-miRNA processor, DCL1, and the dsRNA processors (DCL3 and DCL4). These results suggest that PRL1 may function as a general factor to promote the production of miRNAs and siRNAs. We also show that PRL1 is an RNA-binding protein and associates with pri-miRNAs in vivo. In addition, prl1 reduces pri-miRNA levels without affecting pri-miRNA transcription. These results suggest that PRL1 may stabilize pri-miRNAs and function as a co-factor to enhance DCL1 activity. We further reveal the genetic interaction of PRL1 with CDC5, which interacts with PRL1 and regulates transcription and processing of pri-miRNAs. Both miRNA and pri-miRNA levels are lower in cdc5 prl1 than those in either cdc5 or prl1. However, the processing efficiency of pri-miRNAs in cdc5 prl1 is similar to that in cdc5 and slightly lower than that in prl1. Based on these results, we propose that CDC5 and PRL1 cooperatively regulate pri-miRNA levels, which results in their synergistic effects on miRNA accumulation, while they function together as a complex to enhance DCL1 activity

microRNA biogenesis, degradation and activity in plants

microRNAs (miRNAs) are important regulators of gene expression. After excised from primary miRNA transcript by dicer-like1 (DCL1, an RNAse III enzyme), miRNAs bind and guide their effector protein named argonaute 1 (AGO1) to silence the expression of target RNAs containing their complementary sequences in plants. miRNA levels and activities are tightly controlled to ensure their functions in various biological processes such as development, metabolism and responses to abiotic and biotic stresses. Studies have identified many factors that involve in miRNA accumulation and activities. Characterization of these factors in turn greatly improves our understanding of the processes related to miRNAs. Here, we review recent progress of mechanisms underlying miRNA expression and functions in plants

microRNA biogenesis, degradation and activity in plants

microRNAs (miRNAs) are important regulators of gene expression. After excised from primary miRNA transcript by dicer-like1 (DCL1, an RNAse III enzyme), miRNAs bind and guide their effector protein named argonaute 1 (AGO1) to silence the expression of target RNAs containing their complementary sequences in plants. miRNA levels and activities are tightly controlled to ensure their functions in various biological processes such as development, metabolism and responses to abiotic and biotic stresses. Studies have identified many factors that involve in miRNA accumulation and activities. Characterization of these factors in turn greatly improves our understanding of the processes related to miRNAs. Here, we review recent progress of mechanisms underlying miRNA expression and functions in plants

CDC5, a DNA binding protein, positively regulates posttranscriptional processing and/or transcription of primary microRNA transcripts

CDC5 is a MYB-related protein that exists in plants, animals, and fungi. In Arabidopsis, CDC5 regulates both growth and immunity through unknown mechanisms. Here, we show that CDC5 from Arabidopsis positively regulates the accumulation of microRNAs (miRNAs), which control many biological processes including development and adaptations to environments in plants. CDC5 interacts with both the promoters of genes encoding miRNAs (MIR) and the DNA-dependent RNA polymerase II. As a consequence, lack of CDC5 reduces the occupancy of polymerase II at MIR promoters, as well as MIR promoter activities. In addition, CDC5 is associated with the DICER–LIKE1 complex, which generates miRNAs from their primary transcripts and is required for efficient miRNA production. These results suggest that CDC5 may have dual roles in miRNA biogenesis: functioning as a positive transcription factor of MIR and/or acting as a component of the DICER–LIKE1 complex to enhance primary miRNA processing

Personalized Alert Notifications and Evacuation Routes in Indoor Environments

The preparedness phase is crucial in the emergency management process for reaching an adequate level of readiness to react to potential threats and hazards. During this phase, emergency plans are developed to establish, among other procedures, evacuation and emergency escape routes. Information and Communication Technologies (ICT) can support and improve these procedures providing appropriate, updated and accessible information to all people in the affected zone. Current emergency management and evacuation systems do not adapt information to the context and the profile of each person, so messages received in the emergency might be useless. In this paper, we propose a set of criteria that ICT-based systems could achieve in order to avoid this problem adapting emergency alerts and evacuation routes to different situations and people. Moreover, in order to prove the applicability of such criteria, we define a mechanism that can be used as a complement of traditional evacuation systems to provide personalized alerts and evacuation routes to all kinds of people during emergency situations in working places. This mechanism is composed by three main components: CAP-ONES for notifying emergency alerts, NERES for defining emergency plans and generating personalized evacuation routes, and iNeres as the interface to receive and visualize these routes on smartphones. The usability and understandability of proposed interface has been assessed through a user study performed in a fire simulation in an indoor environment. This evaluation demonstrated that users considered iNeres easy to understand, to learn and to use, and they also found very innovative the idea to use smartphones as a support for escaping instead of static signals on walls and doors

Mutational Analyses of a Fork Head Associated Domain Protein, DAWDLE, in \u3ci\u3eArabidopsis thaliana\u3c/i\u3e

DAWDLE (DDL) gene encodes a protein that contains an N-terminal arginine-rich domain and a C-terminal Fork Head Associated (FHA) domain in Arabidopsis thaliana. DDL protein is believed to function in microRNA biogenesis by mediating the recruitment of pri-microRNA to DICER-LIKE 1 and also stabilizing the microRNA. The aim of this study was to conduct a structure-function analysis to identify the regions in DDL that are of functional significance. Targeted Induced Local Lesions in Genome screen was performed in the Columbia erecta-105 background of Arabidopsis resulting in the identification of eight point mutations spanning DDL. The mutants were characterized by phenotypic and molecular analyses based on the prior knowledge on ddl knockout mutants. Height of the plant, hypocotyl and root length, and fertility were measured for phenotypic characterization, and microRNA172 levels were measured to assess the mutation effect at the molecular level. Phenotypic and molecular analyses of the mutants revealed effects resulting in ddl phenotypes of varying degrees in different organs and each mutant displayed at least one phenotype studied. Reduction in fertility and increase in stem length were two phenotypes that most of the mutants consistently displayed. Identification and characterization of several key residues in the arginine rich region and FHA domain will serve as an important tool for elucidation of DDL signaling pathway

Downregulation of \u3cem\u3eOrco\u3c/em\u3e and \u3cem\u3e5-HTT\u3c/em\u3e Alters Nestmate Discrimination in the Subterranean Termite \u3cem\u3eOndontotermes formosanus\u3c/em\u3e (Shiraki)

Nestmate discrimination allows social insects to recognize nestmates from non-nestmates using colony-specific chemosensory cues, which typically evoke aggressive behavior toward non-nestmates. Functional analysis of genes associated with nestmate discrimination has been primarily focused on inter-colonial discrimination in Hymenopterans, and parallel studies in termites, however, are grossly lacking. To fill this gap, we investigated the role of two genes, Orco and 5-HTT, associated with chemosensation and neurotransmission respectively, in nestmate discrimination in a highly eusocial subterranean termite, Odontotermes formosanus (Shiraki). We hypothesized that knocking down of these genes will compromise the nestmate recognition and lead to the antagonistic behavior. To test this hypothesis, we carried out (1) an in vivo RNAi to suppress the expression of Orco and 5-HTT, respectively, (2) a validation study to examine the knockdown efficiency, and finally, (3) a behavioral assay to document the phenotypic impacts/behavioral consequences. As expected, the suppression of either of these two genes elevated stress level (e.g., vibrations and retreats), and led to aggressive behaviors (e.g., biting) in O. formosanus workers toward their nestmates, suggesting both Orco and 5-HTT can modulate nestmate discrimination in termites. This research links chemosensation and neurotransmission with nestmate discrimination at the genetic basis, and lays the foundation for functional analyses of nestmate discrimination in termites

Methylation protects microRNAs from an AGO1- associated activity that uridylates 5′ RNA fragments generated by AGO1 cleavage

In plants, methylation catalyzed by HEN1 (small RNA methyl transferase) prevents microRNAs (miRNAs) from degradation triggered by uridylation. Howmethylation antagonizes uridylation of miRNAs in vivo is not well understood. In addition, 5′ RNA fragments (5′ fragments) produced by miRNA-mediated RNA cleavage can be uridylated in plants and animals. However, the biological significance of this modification is unknown, and enzymes uridylating 5′ fragments remain to be identified. Here, we report that in Arabidopsis, HEN1 suppressor 1 (HESO1, a miRNA nucleotidyl transferase) uridylates 5′ fragments to trigger their degradation.We also show that Argonaute 1 (AGO1), the effector protein of miRNAs, interacts with HESO1 through its Piwi/Argonaute/Zwille and PIWI domains, which bind the 3′ end of miRNA and cleave the target mRNAs, respectively. Furthermore, HESO1 is able to uridylate AGO1-bound miRNAs in vitro. miRNA uridylation in vivo requires a functional AGO1 in hen1, in which miRNA methylation is impaired, demonstrating that HESO1 can recognize its substrates in the AGO1 complex. On the basis of these results, we propose that methylation is required to protect miRNAs from AGO1-associated HESO1 activity that normally uridylates 5′ fragments

Methylation protects microRNAs from an AGO1- associated activity that uridylates 5′ RNA fragments generated by AGO1 cleavage

In plants, methylation catalyzed by HEN1 (small RNA methyl transferase) prevents microRNAs (miRNAs) from degradation triggered by uridylation. Howmethylation antagonizes uridylation of miRNAs in vivo is not well understood. In addition, 5′ RNA fragments (5′ fragments) produced by miRNA-mediated RNA cleavage can be uridylated in plants and animals. However, the biological significance of this modification is unknown, and enzymes uridylating 5′ fragments remain to be identified. Here, we report that in Arabidopsis, HEN1 suppressor 1 (HESO1, a miRNA nucleotidyl transferase) uridylates 5′ fragments to trigger their degradation.We also show that Argonaute 1 (AGO1), the effector protein of miRNAs, interacts with HESO1 through its Piwi/Argonaute/Zwille and PIWI domains, which bind the 3′ end of miRNA and cleave the target mRNAs, respectively. Furthermore, HESO1 is able to uridylate AGO1-bound miRNAs in vitro. miRNA uridylation in vivo requires a functional AGO1 in hen1, in which miRNA methylation is impaired, demonstrating that HESO1 can recognize its substrates in the AGO1 complex. On the basis of these results, we propose that methylation is required to protect miRNAs from AGO1-associated HESO1 activity that normally uridylates 5′ fragments