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Interferon-gamma (IFNγ) is a pleiotropic cytokine produced by natural killer (NK) cells during the early infection response. IFNγ expression is tightly regulated to mount sterilizing immunity while preventing tissue pathology. Several post-transcriptional effectors dampen IFNγ expression through IFNG mRNA degradation. In this study, we identify mRNA splicing as a positive regulator of IFNγ production. While treatment with the combination of IL-12 and IL-2 causes synergistic induction of IFNG mRNA and protein, defying transcription-translation kinetics, we observe that NK cells treated with IL-12 alone transcribe IFNG with introns intact. When NK cells are treated with both IL-2 and IL-12, IFNG transcript is spliced to form mature mRNA with a concomitant increase in IFNγ protein. We find that IL-2-mediated intron splicing occurs independently of nascent transcription but relies upon NF-κB signaling. We propose that while IL-12 transcriptionally induces IFNG mRNA, IL-2 signaling stabilizes IFNG mRNA by splicing detained introns, allowing for rapid IFNγ protein production. This study uncovers a novel role for cytokine-induced splicing in regulating IFNγ through a mechanism potentially applicable to other inflammatory mediators. Synopsis: IL-2 stimulation post-transcriptionally enhances splicing of IFNG mRNA in NK cells. Further, NF-κB signaling downstream of the IL-2 receptor is required for IFNG mRNA processing. IL-12 and IL-2 signals synergize to induce robust IFNγ protein in NK cells. While IL-12 treatment induces transcription of IFNG mRNA, IL-2 treatment promotes stabilization and intronic splicing. NF-κB signaling as a result of IL-2 or PMA treatment is required for post-transcriptional processing of IFNG mRNA. IL-2 stimulation post-transcriptionally enhances splicing of IFNG mRNA in NK cells. Further, NF-κB signaling downstream of the IL-2 receptor is required for IFNG mRNA processing. [ABSTRACT FROM AUTHOR]

Transcription factors are key molecules involved in transcriptional and post-transcriptional regulation in plants and play an important regulatory role in resisting biological stress. In this study, we identified a regulatory factor, OsZF8, mediating rice response to Rhizoctonia solani (R. solani) AG1-IA infection. The expression of OsZF8 affects R. solani rice infection. OsZF8 knockout and overexpressed rice plants were constructed, and the phenotypes of mutant and wild-type (WT) plants showed that OsZF8 negatively regulated rice resistance to rice sheath blight. However, it was speculated that OsZF8 plays a regulatory role at the protein level. The interacting protein PRB1 of OsZF8 was screened using the yeast two-hybrid and bimolecular fluorescence complementation test. The results showed that OsZF8 effectively inhibited PRB1-induced cell death in tobacco cells, and molecular docking results showed that PRB1 had a strong binding effect with OsZF8. Further, the binding ability of OsZF8-PRB1 to ergosterol was significantly reduced when compared with the PRB1 protein. These findings provide new insights into elucidating the mechanism of rice resistance to rice sheath blight. [ABSTRACT FROM AUTHOR]

The circadian clock system acts as an endogenous timing reference that coordinates many metabolic and physiological processes in plants. Previous studies have shown that the application of osmotic stress delays circadian rhythms via 3ʹ-phospho-adenosine 5ʹ-phosphate (PAP), a retrograde signalling metabolite that is produced in response to redox stress within organelles. PAP accumulation leads to the inhibition of exoribonucleases (XRNs), which are responsible for RNA degradation. Interestingly, we are now able to demonstrate that post-transcriptional processing is crucial for the circadian response to osmotic stress. Our data show that osmotic stress increases the stability of specific circadian RNAs, suggesting that RNA metabolism plays a vital role in circadian clock coordination during drought. Inactivation of XRN4 is sufficient to extend circadian rhythms as part of this response, with PRR7 and LWD1 identified as transcripts that are post-transcriptionally regulated to delay circadian progression. [ABSTRACT FROM AUTHOR]

Purpose: The abnormal regulation of lncRNA CARMN has been proved to be a tumor suppressor gene of cervical cancer (CC). However, its role in CC is still elusive. The regulation of CARMN post-transcriptional level by m6A modification and miRNA has not been studied. This study aims to analyze the molecular mechanism of m6A modification and miRNA on the abnormal expression of CARMN in CC cells, so as to provide a new theoretical basis for the diagnosis and treatment of CC. Methods: MeRIP-seq was used to identify the differential m6A-modified genes between tumor and normal cervical tissues. RT-qPCR assay was used to detect gene expression levels in tissues or cells. The m6A modification sites of CARMN was predicted by bioinformatics, and the modification of m6A and its regulatory effect on CARMN were analyzed by MeRIP-qPCR, Actinomycin D assay and RIP assay. RIP-microarray combined with bioinformatics methods to screen miRNAs that may target CARMN. The regulation mechanism between miRNA and CARMN was verified by RT-qPCR, nucleo-plasmic separation assay, mRNA stability assay, dual-luciferase reporter assay, and in vivo experiments. Results: MeRIP-seq found that CARMN is a significant different gene in the abundance of m6A in CC, and the modification level of m6A in CC tissues was higher than that in normal cervical tissues. Further, this study verified that m6A reader YTHDF2 could recognize m6A-modified CARMN and promote its degradation in CC cells. miR-21-5p was proved to be the downstream target gene of CARMN, and miR-21-5p could negatively regulate the expression of CARMN. Further experiments showed that miR-21-5p could directly bind to CARMN and lead to the degradation of CARMN. The in vivo experimental results indicated that the level of miR-21-5p in the overexpressed CARMN group was significantly lower than that in the control group. Conclusion: m6A modification and miR-21-5p play important roles in promoting the occurrence and development of tumors by regulating CARMN, provide new potential targets for the treatment of CC. [ABSTRACT FROM AUTHOR]

More than 170 post‐transcriptional modifications of RNAs have currently been identified. Detailed biophysical investigations of these modifications have been limited since large RNAs containing these post‐transcriptional modifications are difficult to produce. Further, adequate readout of spectroscopic fingerprints are important, necessitating additional labeling procedures beyond the naturally occurring RNA modifications. Here, we report the chemo‐enzymatic synthesis of RNA modifications and several structurally similar fluorine‐modified analogs further optimizing a recently developed methodology.[1] This chemo‐enzymatic method allows synthesis of also large RNAs. We were able to incorporate 16 modified nucleotides and 6 19F‐labeled nucleotides. To showcase the applicability of such modified large RNAs, we incorporated a 19F‐labeled cytidine into the aptamer domain of the 2'dG sensing riboswitch (2'dG‐sw) from Mesoplasma florum, enabling characterizing RNA fold, ligand binding and kinetics. Thanks to the large chemical shift dispersion of 19F, we can detect conformational heterogeneity in the apo state of the riboswitch. [ABSTRACT FROM AUTHOR]

SUMMARY: Salt stress is one of the major causes of reduced crop production, limiting agricultural development globally. Plants have evolved with complex systems to maintain the balance between growth and stress responses, where signaling pathways such as hormone signaling play key roles. Recent studies revealed that hormones are modulated by microRNAs (miRNAs). Previously, two sweet sorghum (Sorghum bicolor) inbred lines with different salt tolerance were identified: the salt‐tolerant M‐81E and the salt‐sensitive Roma. The levels of endogenous hormones in M‐81E and Roma varied differently under salt stress, showing a different balance between growth and stress responses. miRNA and degradome sequencing showed that the expression of many upstream transcription factors regulating signal transduction and hormone‐responsive genes was directly induced by differentially expressed miRNAs, whose levels were very different between the two sweet sorghum lines. Furthermore, the effects of representative miRNAs on salt tolerance in sorghum were verified through a transformation system mediated by Agrobacterium rhizogenes. Also, miR‐6225‐5p reduced the level of Ca2+ in the miR‐6225‐5p‐overexpressing line by inhibiting the expression of the Ca2+ uptake gene SbGLR3.1 in the root epidermis and affected salt tolerance in sorghum. This study provides evidence for miRNA‐mediated growth and stress responses in sweet sorghum. Significance Statement: The interactions between plant hormones, miRNAs, and key transcription factors form a complex response network that balances the growth and stress responses in sweet sorghum (Sorghum bicolor). [ABSTRACT FROM AUTHOR]

Main conclusion: Circadian clock components exhibit structural variations in different plant systems, and functional variations during various abiotic stresses. These variations bear relevance for plant fitness and could be important evolutionarily. All organisms on earth have the innate ability to measure time as diurnal rhythms that occur due to the earth's rotations in a 24-h cycle. Circadian oscillations arising from the circadian clock abide by its fundamental properties of periodicity, entrainment, temperature compensation, and oscillator mechanism, which is central to its function. Despite the fact that a myriad of research in Arabidopsis thaliana illuminated many detailed aspects of the circadian clock, many more variations in clock components’ organizations and functions remain to get deciphered. These variations are crucial for sustainability and adaptation in different plant systems in the varied environmental conditions in which they grow. Together with these variations, circadian clock functions differ drastically even during various abiotic and biotic stress conditions. The present review discusses variations in the organization of clock components and their role in different plant systems and abiotic stresses. We briefly introduce the clock components, entrainment, and rhythmicity, followed by the variants of the circadian clock in different plant types, starting from lower non-flowering plants, marine plants, dicots to the monocot crop plants. Furthermore, we discuss the interaction of the circadian clock with components of various abiotic stress pathways, such as temperature, light, water stress, salinity, and nutrient deficiency with implications for the reprogramming during these stresses. We also update on recent advances in clock regulations due to post-transcriptional, post-translation, non-coding, and micro-RNAs. Finally, we end this review by summarizing the points of applicability, a remark on the future perspectives, and the experiments that could clear major enigmas in this area of research. [ABSTRACT FROM AUTHOR]

To sense the transition from environment to host, bacteria use a range of environmental cues to control expression of virulence genes. Iron is tightly sequestered in host tissues and in the human pathogen enterohemorrhagic Escherichia coli (EHEC) iron-limitation induces transcription of the outer membrane haem transporter encoded by chuAS. ChuA expression is post-transcriptionally activated at 37°C by a FourU RNA thermometer ensuring that the haem receptor is only expressed under low iron, high temperature conditions that indicate the host. Here we demonstrate that expression of chuA is also independently regulated by the cAMP-responsive small RNA (sRNA) CyaR and transcriptional terminator Rho. These results indicate that chuAS expression is regulated at the transcription initiation, transcript elongation, and translational level. We speculate that additional sensing of the gluconeogenic environment allows further precision in determining when EHEC is at the gastrointestinal epithelium of the host. With previous studies, it appears that the chuAS transcript is controlled by eight regulatory inputs that control expression through six different transcriptional and post-transcriptional mechanisms. The results highlight the ability of regulatory sRNAs to integrate multiple environmental signals into a layered hierarchy of signal input. [ABSTRACT FROM AUTHOR]

SUMMARY: Plants need to rapidly and flexibly adjust their metabolism to changes of their immediate environment. Since this necessity results from the sessile lifestyle of land plants, key mechanisms for orchestrating central metabolic acclimation are likely to have evolved early. Here, we explore the role of lysine acetylation as a post‐translational modification to directly modulate metabolic function. We generated a lysine acetylome of the moss Physcomitrium patens and identified 638 lysine acetylation sites, mostly found in mitochondrial and plastidial proteins. A comparison with available angiosperm data pinpointed lysine acetylation as a conserved regulatory strategy in land plants. Focusing on mitochondrial central metabolism, we functionally analyzed acetylation of mitochondrial malate dehydrogenase (mMDH), which acts as a hub of plant metabolic flexibility. In P. patens mMDH1, we detected a single acetylated lysine located next to one of the four acetylation sites detected in Arabidopsis thaliana mMDH1. We assessed the kinetic behavior of recombinant A. thaliana and P. patens mMDH1 with site‐specifically incorporated acetyl‐lysines. Acetylation of A. thaliana mMDH1 at K169, K170, and K334 decreases its oxaloacetate reduction activity, while acetylation of P. patens mMDH1 at K172 increases this activity. We found modulation of the malate oxidation activity only in A. thaliana mMDH1, where acetylation of K334 strongly activated it. Comparative homology modeling of MDH proteins revealed that evolutionarily conserved lysines serve as hotspots of acetylation. Our combined analyses indicate lysine acetylation as a common strategy to fine‐tune the activity of central metabolic enzymes with likely impact on plant acclimation capacity. Significance Statement: We explore the role of lysine acetylation as a mechanism to directly modulate mitochondrial metabolism in land plants by generating the lysine acetylome of the moss Physcomitrium patens and comparing it with available angiosperm data. We found acetylation of evolutionarily conserved lysines as a strategy to fine‐tune the activity of mitochondrial malate dehydrogenase in a species‐dependent molecular context. [ABSTRACT FROM AUTHOR]

The endogenous circadian timekeeping system drives ~24-h rhythms in gene expression and rhythmically coordinates the physiology, metabolism and behavior in a wide range of organisms. Regulation at various levels is important for the accurate functioning of this circadian timing system. The core circadian oscillator consists of an interlocked transcriptional-translational negative feedback loop (TTFL) that imposes a substantial delay between the accumulation of clock gene mRNA and its protein to generate 24-h oscillations. This TTFL mediated daily oscillation of clock proteins is further fine-tuned by post-translational modifications that regulate the clock protein stability, interaction with other proteins and subcellular localization. Emerging evidence from various studies indicates that besides TTFL and post-translational modifications, post-transcriptional regulation plays a key role in shaping the rhythmicity of mRNAs and to delay the accumulation of clock proteins in relation to their mRNAs. In this review, we summarize the current knowledge on the importance of post-transcriptional regulatory mechanisms such as splicing, polyadenylation, the role of RNA-binding proteins, RNA methylation and microRNAs in the context of shaping the circadian rhythmicity in Drosophila and mammals. In particular, we discuss microRNAs, an important player in post-transcriptional regulation of core-clock machinery, circadian neural circuit, clock input, and output pathways. Furthermore, we provide an overview of the microRNAs that exhibit diurnal rhythm in expression and their role in mediating rhythmic physiological processes. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are noncoding, small RNAs of 20–24 nucleotides (nt) and function critically at the post-transcriptional level to regulate gene expression through cleaving mRNA targets or interfering with translation of the target mRNAs. They are broadly involved in many biological processes in plants. The miR397 family in plants contains several conserved members either in 21-nt or in 22-nt that mainly target the laccase (LAC) genes functioning in lignin synthesis and are involved in the development of plants under various conditions. Recent findings showed that miR397b in Arabidopsis could also target to Casein Kinase II Subunit Beta 3 (CKB3) and mediate circadian regulation and plant flowering. This review aims to summarise recent updates on miR397 and provides the available basis for understanding the functional mechanisms of miR397 in plant growth and development regulation and in response to external adverse stimulation. MiR397 is involved in regulation of plant development under various conditions. This review provides concise and precise updates on the latest progress on the research of a conserved microRNA, miR397. It will be helpful in understanding the function of miR397 and divulging its application prospects for increasing agricultural production as well as its ability to enhance plant adaptation to abiotic stresses. [ABSTRACT FROM AUTHOR]

The plant RNA degradome was defined as an aggregate of the RNA fragments degraded from various biochemical pathways, such as RNA turnover, maturation and quality surveillance. In recent years, the degradome sequencing (degradome-seq) libraries became a rich storehouse for researchers to study on RNA processing and regulation. Here, we provided a brief overview of the uses of degradome-seq data in plant RNA biology, especially on non-coding RNA processing and small RNA-guided target cleavages. Some novel applications in RNA research area, such as in vivo mapping of the endoribonucleolytic cleavage sites, identification of conserved motifs at the 5ʹ ends of the uncapped RNA fragments, and searching for the protein-binding regions on the transcripts, were also mentioned. More importantly, we proposed a model for the biologists to deduce the contributions of transcriptional and/or post-transcriptional regulation to gene differential expression based on degradome-seq data. Finally, we hope that the degradome-based analytical methods could be widely applied for the studies on RNA biology in eukaryotes. [ABSTRACT FROM AUTHOR]

The central dogma of molecular biology illustrates the importance of mRNAs as critical mediators between genetic information encoded at the DNA level and proteomes/metabolomes that determine the diverse functional outcome at the cellular and organ levels. Although the total number of protein-producing (coding) genes in the mammalian genome is ~20,000, it is evident that the intricate processes of cardiac development and the highly regulated physiological regulation in the normal heart, as well as the complex manifestation of pathological remodeling in a diseased heart, would require a much higher degree of complexity at the transcriptome level and beyond. Indeed, in addition to an extensive regulatory scheme implemented at the level of transcription, the complexity of transcript processing following transcription is dramatically increased. RNA processing includes post-transcriptional modification, alternative splicing, editing and transportation, ribosomal loading, and degradation. While transcriptional control of cardiac genes has been a major focus of investigation in recent decades, a great deal of progress has recently been made in our understanding of how post-transcriptional regulation of mRNA contributes to transcriptome complexity. In this review, we highlight some of the key molecular processes and major players in RNA maturation and post-transcriptional regulation. In addition, we provide an update to the recent progress made in the discovery of RNA processing regulators implicated in cardiac development and disease. While post-transcriptional modulation is a complex and challenging problem to study, recent technological advancements are paving the way for a new era of exciting discoveries and potential clinical translation in the context of cardiac biology and heart disease. [ABSTRACT FROM AUTHOR]

An inflammatory response is essential for combating invading pathogens. Several effector components, as well as immune cell populations, are involved in mounting an immune response, thereby destroying pathogenic organisms such as bacteria, fungi, viruses, and parasites. In the past decade, microRNAs (miRNAs), a group of noncoding small RNAs, have emerged as functionally significant regulatory molecules with the significant capability of fine-tuning biological processes. The important role of miRNAs in inflammation and immune responses is highlighted by studies in which the regulation of miRNAs in the host was shown to be related to infectious diseases and associated with the eradication or susceptibility of the infection. Here, we review the biological aspects of microRNAs, focusing on their roles as regulators of gene expression during pathogen–host interactions and their implications in the immune response against Leishmania, Trypanosoma, Toxoplasma, and Plasmodium infectious diseases. [ABSTRACT FROM AUTHOR]

Bicaudal-C (Bicc1) is a conserved RNA-binding protein that represses the translation of selected mRNAs to control development. In Xenopus embryos, Bicc1 binds and represses specific maternal mRNAs to control anterior-posterior cell fates. However, it is not known how Bicc1 binds its RNA targets or how binding affects Bicc1-dependent embryogenesis. Focusing on the KH domains, we analyzed Bicc1 mutants for their ability to bind RNA substrates in vivo and in vitro. Analyses of these Bicc1 mutants demonstrated that a single KH domain, KH2, was crucial for RNA binding in vivo and in vitro, while the KH1 and KH3 domains contributed minimally. The Bicc1 mutants were also assayed for their ability to repress translation, and results mirrored the RNA-binding data, with KH2 being the only domain essential for repression. Finally, maternal knockdown and rescue experiments indicated that the KH domains were essential for the regulation of embryogenesis by Bicc1. These data advance our understanding of how Bicc1 selects target mRNAs and provide the first direct evidence that the RNA binding functions of Bicc1 are essential for both Bicc1-dependent translational repression and maternal vertebrate development. [ABSTRACT FROM AUTHOR]

Maintaining the diversity and constant numbers of naïve T cells throughout the organism's lifetime is necessary for efficient immune responses. Naïve T cell homeostasis, which consists of prolonged survival, occasional proliferation and enforcement of quiescence, is tightly regulated by multiple signaling pathways which are in turn controlled by various transcription factors. However, full understanding of the molecular mechanisms underlying the maintenance of the peripheral T cell pool has not been achieved. In the present study, we demonstrate that T cell-specific deficiency in let-7 miRNAs results in peripheral T cell lymphopenia resembling that of Dicer1 knockout mice. Deletion of let-7 leads to profound T cell apoptosis while overexpression prevents it. We further show that in the absence of let-7, T cells cannot sustain optimal levels of the pro-survival factor Bcl2 in spite of the intact IL-7 signaling, and re-expression of Bcl2 in let-7 deficient T cells completely rescues the survival defect. Thus, we have uncovered a novel let-7-dependent mechanism of post-transcriptional regulation of naïve T cell survival in vivo. [ABSTRACT FROM AUTHOR]

AU-rich elements (AREs) are cis-acting instability and translation inhibition elements that are present in the 3ʹUTR of most inducible inflammatory mRNAs such as TNF and Cxcl2. mRNAs that contain AREs are, by default, repressed and only transiently expressed in response to stimuli. They are targeted by the inducible RNA-binding protein Tristetraprolin (TTP) which blocks their translation and facilitates their decay, thereby contributing to the quick termination of their expression. The exogenous over-expression of TTP in HEK293 cells can unexpectedly lead to the upregulation and extended expression of a nanoLuciferase reporter that contains the ARE of TNF. Here we show that, a moderate downregulation of the highly expressed endogenous TTP after LPS induction by siRNA in macrophages can lead to a reduction in the release of TNF and Cxcl2. We propose that, in contrast to their canonical function, very high levels of induced TTP at the onset of the inflammatory response can enhance the expression of ARE-mRNAs at the post-transcriptional level, independently of phosphorylation status. As the inflammatory response progresses, TTP levels diminish but they continuously regain their ability to reduce the expression of ARE-mRNAs to reach a turning point of 'optimal TTP level' with a maximum ability to repress ARE-mRNA expression. Below this level, a further reduction in TTP levels now leads to the loss of canonical-TTP function resulting in increased ARE-mRNA expression. These novel findings should contribute to the understanding of feedback loops that control the kinetics of the inflammatory response. [ABSTRACT FROM AUTHOR]

When plants are subjected to unfavorable environmental conditions, overall gene expression in stressed cells is altered from a programmed pattern for normal development to an adaptive pattern for survival. Rapid changes in plant gene expression include production of stress responsive proteins for protection as well as reduction of irrelevant proteins to minimize energy consumption during growth. In addition to the many established mechanisms known to modulate gene expression in eukaryotes, a novel strategy involving tRNA-derived fragments (tRFs) was recently reported to control gene expression. In animals, tRFs are shown to play a certain role in infected or cancer cells. However, tRFs are expected to function in the regulation of gene expression against abiotic stress conditions in plants. Moreover, the underlying mechanism linking up-regulation of tRFs under stress conditions with the stress tolerant response remains unknown. In this review, the biogenesis and putative function of diverse tRFs in abiotic stress signaling are discussed with a focus on tRFs as a transcriptional/post-transcriptional/translational regulator. [ABSTRACT FROM AUTHOR]

Post-transcriptional regulation of gene expression plays a crucial role in many processes. In cells, it is mediated by diverse RNA-binding proteins. These proteins can influence mRNA stability, translation, and localization. The PUF protein family (Pumilio and FBF) is composed of RNA-binding proteins highly conserved among most eukaryotic organisms. Previous investigations indicated that they could be involved in many processes by binding corresponding motifs in the 30UTR or by interacting with other proteins. To date, most of the investigations on PUF proteins have been focused on Caenorhabditis elegans, Drosophila melanogaster, and Saccharomyces cerevisiae, while only a few have been conducted on Arabidopsis thaliana. The present article provides an overview of the PUF protein family. It addresses their RNA-binding motifs, biological functions, and post-transcriptional control mechanisms in Caenorhabditis elegans, Drosophila melanogaster, Saccharomyces cerevisiae, and Arabidopsis thaliana. These items of knowledge open onto new investigations into the relevance of PUF proteins in specific plant developmental processes.Post-transcriptional regulation of gene expression plays a crucial role in many processes. In cells, it is mediated by diverse RNA-binding proteins. These proteins can influence mRNA stability, translation, and localization. The PUF protein family (Pumilio and FBF) is composed of RNA-binding proteins highly conserved among most eukaryotic organisms. Previous investigations indicated that they could be involved in many processes by binding corresponding motifs in the 3'UTR or by interacting with other proteins. To date, most of the investigations on PUF proteins have been focused on Caenorhabditis elegans, Drosophila melanogaster, and Saccharomyces cerevisiae, while only a few have been conducted on Arabidopsis thaliana. The present article provides an overview of the PUF protein family. It addresses their RNA-binding motifs, biological functions, and post-transcriptional control mechanisms in Caenorhabditis elegans, Drosophila melanogaster, Saccharomyces cerevisiae, and Arabidopsis thaliana. These items of knowledge open onto new investigations into the relevance of PUF proteins in specific plant developmental processes. [ABSTRACT FROM AUTHOR]

Contents Gamete and embryo development are indispensable processes for successful reproduction. Cells involved in these processes acquire pluripotency, the ability to differentiate into multiple different cell types, through a series of events known as reprogramming that lead to profound changes in histone and DNA methylation. While essential for pluripotency, this epigenetic remodelling removes constraints that normally limit the expression of genomic sequences known as transposable elements ( TEs). Unconstrained TE expression can lead to many deleterious consequences including infertility, so organisms have evolved complex and potent mechanistic arsenals to target and suppress TE expression during reprogramming. This review will focus on the control of transposable elements in gametes and embryos, and one important TE suppressing system known as the PIWI pathway. This broadly conserved, small RNA-targeted silencing mechanism appears critical for fertility in many species and may participate in multiple aspects of gene regulation in reproduction and other contexts. [ABSTRACT FROM AUTHOR]

To explore if the regulation at post-transcriptional level of follicular phase (Fols) to luteal phase (Luts) transition occurs in the ovaries of Anhuai goats, the differentially expressed microRNAs (miRNAs) of ovaries in the Fols and Luts were analyzed using Solexa sequencing in the study. In total, 320 known miRNAs were co-expressed in the two phases, 339 and 353 known miRNAs were expressed in the ovary in the Fols and Luts, respectively. In addition, 45 novel miRNAs were co-expressed in the two phases, 70 and 94 novel miRNAs were expressed in the ovary in the Fols and Luts, respectively. Let-7f was the highest expressed significantly different known miRNA in the two phases, and mir-159 was the highest expressed significantly different novel miRNA in the two phases, which may participate in the follicular-luteal transition of Anhuai goats. GO annotation and KEGG pathway analysis were applied to analyze the target genes of differentially expressed miRNAs detected in the two phases. The results will help to further understand the role of miRNAs in the regulation of follicular to luteal transition in goat ovaries. [ABSTRACT FROM AUTHOR]

Cellular senescence is a complex stress response that leads to an irreversible state of cell growth arrest. Senescence may be induced by various stimuli such as telomere shortening, DNA damage or oncogenic insult, among others. Senescent cells are metabolically highly active, producing a wealth of cytokines and chemokines that, depending on the context, may have a beneficial or deleterious effect on the organism. Senescence is considered a tightly regulated stress response that is largely governed by the p53/p21 and p16/Rb pathways. Many molecules have been identified as regulators of these two networks, such as transcription factors, chromatin modifiers and non-coding RNAs. The expression level of several long non-coding RNAs is affected during different types of senescence; however, which of these are important for the biological function remains poorly understood. Here we review our current knowledge of the mechanistic roles of lncRNAs affecting the main senescence pathways, and discuss the importance of identifying new regulators. [ABSTRACT FROM AUTHOR]

Cardiovascular diseases such as atherosclerosis are one of the leading causes of morbidity and mortality worldwide. The clinical manifestations of atherosclerosis, which include heart attack and stroke, occur several decades after initiation of the disease and become more severe with age. Inflammation of blood vessels plays a prominent role in atherogenesis. Activation of the endothelium by inflammatory mediators leads to the recruitment of circulating inflammatory cells, which drives atherosclerotic plaque formation and progression. Inflammatory signaling within the endothelium is driven predominantly by the pro-inflammatory transcription factor, NF-κB. Interestingly, activation of NF-κB is enhanced during the normal aging process and this may contribute to the development of cardiovascular disease. Importantly, studies utilizing mouse models of vascular inflammation and atherosclerosis are uncovering a network of noncoding RNAs, particularly microRNAs, which impinge on the NF-κB signaling pathway. Herewe summarize the literature regarding the control of vascular inflammation by microRNAs, and provide insight into how these microRNA-based pathways might be harnessed for therapeutic treatment of disease. We also discuss emerging areas of endothelial cell biology, including the involvement of long noncoding RNAs and circulating microRNAs in the control of vascular inflammation. [ABSTRACT FROM AUTHOR]

The morphology and the connectivity of neuronal structures formed during early development must be actively maintained as the brain matures. Although impaired axon stability is associated with the progression of various neurological diseases, relatively little is known about the factors controlling this process. We identified Brain tumor (Brat), a conserved member of the TRIM-NHL family of proteins, as a new regulator of axon maintenance in Drosophila CNS. Brat function is dispensable for the initial growth of Mushroom Body axons, but is required for the stabilization of axon bundles. We found that Brat represses the translation of src64B, an upstream regulator of a conserved Rho-dependent pathway previously shown to promote axon retraction. Furthermore, brat phenotypes are phenocopied by src64B overexpression, and partially suppressed by reducing the levels of src64B or components of the Rho pathway, suggesting that brat promotes axon maintenance by downregulating the levels of Src64B. Finally, Brat regulates brain connectivity via its NHL domain, but independently of its previously described partners Nanos, Pumilio, and d4EHP. Thus, our results uncover a novel post-transcriptional regulatory mechanism that controls the maintenance of neuronal architecture by tuning the levels of a conserved rho-dependent signaling pathway. [ABSTRACT FROM AUTHOR]

Persistent and relapsing infections, despite apparently adequate antibiotic therapy, occur frequently with many pathogens, but it is an especially prominent problem with Staphylococcus aureus infections. For the purposes of this review, persistence will encompass both of the concepts of long term survival within the host, including colonization, and the concept of resisting antibiotic therapy even when susceptible in the clinical microbiology laboratory. Over the past two decades, the mechanisms whereby bacteria achieve persistence are slowly being unraveled. S. aureus small colony variants (SCVs) are linked to chronic, recurrent, and antibiotic-resistant infections, and the study of SCVs has contributed significantly to understanding of persistence. In our earlier work, defects in electron transport and thymidylate biosynthesis were linked to the development of the SCV phenotype (reviewed in 2006), thus this work will be discussed only briefly. Since 2006, it has been found that persistent organisms including SCVs are part of the normal life cycle of bacteria, and often they arise in response to harsh conditions, e.g., antibiotics, starvation, host cationic peptides. Many of the changes found in these early SCVs have provided a map for the discovery mechanisms (pathways) for the development of persistent organisms. For example, changes in RNA processing, stringent response, toxin-antitoxin, ribosome protein L6 (RplF), and cold shock protein B (CspB) found in SCVs are also found in other persisters. In addition, many classic persister organisms also show slow growth, hence SCVs. Recent work on S. aureus USA300 has elucidated the impact of aerobic expression of arginine deiminase genes on its ability to chronically colonize the skin and survive in abscesses. S. aureus SCVs also express arginine deiminase genes aerobically as well. Thus, many pathways found activated in electron transport type of SCVs are also increased in persisters that have intact electron transport. Many of these changes in metabolism result in slow growth; hence, small colonies are formed. Another common theme is that slow growth is also associated with reduced expression of virulence factors and enhanced uptake/survival within host cells. These adaptations to survive within the host are rooted in responses that were required for organisms to survive in a harsh environment long before they were mammals on the earth. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression. For over a decade the deluge of research describing the biogenesis and activity of miRNAs has lead researchers to postulate rules to help make sense of the enormous amount of data produced. These rules are repeated in miRNA research papers and reviews. While these rules have been helpful one must be conscious of their limitations or risk missing future breakthroughs. Here we describe some of the most commonly stated rules, the reasoning behind their formation, their uses, and limitations. [ABSTRACT FROM AUTHOR]

Short Interspersed Nuclear Elements (SINEs) are non-autonomous retrotransposons that comprise a large fraction of the human genome. SINEs are demethylated in human disease, but whether SINEs become transcriptionally induced and how the resulting transcripts may affect the expression of protein coding genes is unknown. Here, we show that downregulation of the mRNA of the tumor suppressor gene BRCA1 is associated with increased transcription of SINEs and production of sense and antisense SINE small RNAs. We find that BRCA1 mRNA is post-transcriptionally down-regulated in a Dicer and Drosha dependent manner and that expression of a SINE inverted repeat with sequence identity to a BRCA1 intron is sufficient for downregulation of BRCA1 mRNA. These observations suggest that transcriptional activation of SINEs could contribute to a novel mechanism of RNA mediated post-transcriptional silencing of human genes. [ABSTRACT FROM AUTHOR]

ABSTRACT When bound to the vitamin D receptor (VDR), the active form of vitamin D, 1,25-dihydroxyvitamin D (1,25D) is a potent regulator of osteoblast transcription. Less clear is the impact of 1,25D on posttranscriptional events in osteoblasts, such as the generation and action of microRNAs (miRNAs). Microarray analysis using replicate ( n = 3) primary cultures of human osteoblasts (HOBs) identified human miRNAs that were differentially regulated by >1.5-fold following treatment with 1,25D (10 nM, 6 hours), which included miRNAs 637 and 1228. Quantitative reverse transcription PCR analyses showed that the host gene for miR-1228, low-density lipoprotein receptor-related protein 1 ( LRP1), was coinduced with miR-1228 in a dose-dependent fashion following treatment with 1,25D (0.1-10 nM, 6 hours). By contrast, the endogenous host gene for miR-637, death-associated protein kinase 3 ( DAPK3), was transcriptionally repressed by following treatment with 1,25D. Analysis of two potential targets for miR-637 and miR-1228 in HOB, type IV collagen (COL4A1) and bone morphogenic protein 2 kinase (BMP2K), respectively, showed that 1,25D-mediates suppression of these targets via distinct mechanisms. In the case of miR-637, suppression of COL4A1 appears to occur via decreased levels of COL4A1 mRNA. By contrast, suppression of BMP2K by miR-1228 appears to occur by inhibition of protein translation. In mature HOBs, small interfering RNA (siRNA) inactivation of miR-1228 alone was sufficient to abrogate 1,25D-mediated downregulation of BMP2K protein expression. This was associated with suppression of prodifferentiation responses to 1,25D in HOB, as represented by parallel decrease in osteocalcin and alkaline phosphatase expression. These data show for the first time that the effects of 1,25D on human bone cells are not restricted to classical VDR-mediated transcriptional responses but also involve miRNA-directed posttranscriptional mechanisms. [ABSTRACT FROM AUTHOR]

Prostate cancer is a leading cause of cancer mortality in men. One of the distinct characteristics of prostate cancer is over-expression of the ERG proto-oncogene. The TMPRSS2-ERG gene fusion, the most common gene fusion, is found in approximately 50% of prostate cancer cases. We show that certain micro RNAs are extensively deregulated in prostate cancer cell lines and primary clinical cancer samples. Micro RNAs are capable of modulating post-transcriptional gene expression via inhibition of protein synthesis. Independent target prediction methods have indicated that the 3′ untranslated region of the ERG m RNA is a potential target of miR-145. miR-145 is consistently down-regulated in prostate cancer. Here we show that the ERG 3′ untranslated region is a regulative target of miR-145 in vitro. Ectopic expression of miR-145 led to a reduction in expression of the ERG protein. We analyzed 26 prostate cancer samples and corresponding normal tissue. ERG protein expression was found to be elevated in the tumor samples, together with increased expression of several ERG isoforms. We identified ERG proteins of 35 and 24 kDa, which may represent unknown ERG splice variants. Analyses of miR-145 and ERG m RNA expression revealed a general down-regulation of miR-145 irrespective of the presence or absence of translocations involving ERG. This observation indicates that down-regulation of miR-145 may contribute to the increased expression of most ERG splice variants sharing the miR-145 target sequence in their 3′ untranslated region. [ABSTRACT FROM AUTHOR]

I.II.III.IV.V.VI.VII.References Summary: Infection by phytopathogenic bacteria triggers massive changes in plant gene expression, which are thought to be mostly a result of transcriptional reprogramming. However, evidence is accumulating that plants additionally use post‐transcriptional regulation of immune‐responsive mRNAs as a strategic weapon to shape the defense‐related transcriptome. Cellular RNA‐binding proteins regulate RNA stability, splicing or mRNA export of immune‐response transcripts. In particular, mutants defective in alternative splicing of resistance genes exhibit compromised disease resistance. Furthermore, detection of bacterial pathogens induces the differential expression of small non‐coding RNAs including microRNAs that impact the host defense transcriptome. Phytopathogenic bacteria in turn have evolved effector proteins to inhibit biogenesis and/or activity of cellular microRNAs. Whereas RNA silencing has long been known as an antiviral defense response, recent findings also reveal a major role of this process in antibacterial defense. Here we review the function of RNA‐binding proteins and small RNA‐directed post‐transcriptional regulation in antibacterial defense. We mainly focus on studies that used the model system Arabidopsis thaliana and also discuss selected examples from other plants. [ABSTRACT FROM AUTHOR]

RNA interference (RNAi) technology has become a novel tool for silencing gene expression in cells or organisms. RNA interference is the process that double-stranded RNA induces the homology-dependent degradation of cognate mRNA mediated by 21-23 nucleotide short interfering RNA (siRNA). RNA interference is a powerful mechanism of gene silencing that underlies many aspects of eukaryotic biology. On the molecular level, RNAi is mediated by a family of ribonucleoprotein (RNP) complexes called RNA-Induced Silencing Complexes (RISCs), which can be programmed to target virtually any nucleic acid sequence for silencing. The ability of RISC to locate target RNAs been co-opted by evolution many times to generate a broad spectrum of gene silencing pathways. The study about the Silencing of gene expression by siRNA is rapidly becoming a powerful tool for genetic analysis and represents a potential strategy for therapeutic product development. In this study, the applications of siRNA expressing recombinant adenovirus system in plants, animals and in cancer gene therapy are given importance with its modifications. [ABSTRACT FROM AUTHOR]

Chinese hamster ovary (CHO) cell lines are frequently used as hosts for the production of recombinant therapeutics, such as monoclonal antibodies, due to their ability to perform correct post-translational modifications. A potential issue when utilizing CHO cells for therapeutic protein production is the selection of cell lines that do not retain stable protein expression during long-term culture (LTC). Instability of expression impairs process yields, effective usage of time and money, and regulatory approval for the desired therapeutic. In this study, we investigated a model unstable GS-CHO cell line over a continuous period of approximately 100 generations to determine markers of mechanisms that underlie instability. In this cell line, stability of expression was retained for 40-50 generations after which time a 40% loss in antibody production was detected. The instability observed within the cell line was not due to a loss in recombinant gene copy number or decreased expression of mRNA encoding for recombinant antibody H or L chain, but was associated with lower cumulative cell time values and an apparent increased sensitivity to cellular stress (exemplified by increased mRNA expression of the stress-inducible gene GADD153). Changes were also noted in cellular metabolism during LTC (alterations to extracellular alanine accumulation, and enhanced rates of glucose and lactate utilization, during the exponential and decline phase of batch culture, respectively). Our data indicates the breadth of changes that may occur to recombinant CHO cells during LTC ranging from instability of recombinant target production at a post-mRNA level to metabolic events. Definition of the mechanisms, regulatory events, and linkages underpinning cellular phenotype changes require further detailed analysis at a molecular level. Biotechnol. Bioeng. 2012; 109:2093-2103. © 2012 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

The formation of putrescine by ornithine decarboxylase (ODC) is a key regulatory step in polyamine biosynthesis in metazoa and fungi. Excess polyamines post-transcriptionally induce the synthesis of a unique non-competitive protein inhibitor of ODC, termed antizyme. Binding of antizyme to an ODC monomer subunit results in enzymatic inhibition, rapid ubiquitin-independent degradation of ODC by the 26S proteasome and recycling of antizyme. Plants possess an additional route for synthesizing putrescine via arginine decarboxylase (ADC). No homologue of ODC antizyme has been detected in plant genomes but several biochemical studies have reported plant ODC antizyme proteins of 9 and 16 kDa. Here we show that plant cells grown in liquid culture do not exhibit any substantial post-transcriptional, polyamine-responsive feedback regulation of ODC or ADC. However, using the yeast two hybrid system, a plant ODC-binding polypeptide was detected: the C-terminal 84-87 amino acids of cytosolic ribosomal protein (rp) S15. The Arabidopsis rpS15 polypeptide interacted specifically with plant ODC but not with human or Saccharomyces cerevisiae ODCs. Co-expression of either the full length or C-terminal rpS15 polypeptides with a plant ODC in yeast did not reduce ODC enzymatic activity. Only the full length mRNA encoding rpS15 was detected in Arabidopsis cells, suggesting that the C-terminal rpS15 polypeptide is encoded by a low abundance mRNA or the polypeptide is not physiologically relevant in plants. These results confirm the primacy of S-adenosylmethionine decarboxylase as the key regulatory enzyme in plant polyamine biosynthesis. [ABSTRACT FROM AUTHOR]

Thrombospondin-1 (TSP-1) is a matricellular protein that participates in numerous normal and pathological tissue processes and is rapidly modulated by different stimuli. The presence of 8 highly-conserved AU rich elements (AREs) within the 3′-untranslated region (3′UTR) of the TSP-1 mRNA suggests that post-transcriptional regulation is likely to represent one mechanism by which TSP-1 gene expression is regulated. We investigated the roles of these AREs, and proteins which bind to them, in the control of TSP-1 mRNA stability. The endogenous TSP-1 mRNA half-life is approximately 2.0 hours in HEK293 cells. Luciferase reporter mRNAs containing the TSP-1 3′UTR show a similar rate of decay, suggesting that the 3′UTR influences the decay rate. Site-directed mutagenesis of individual and adjacent AREs prolonged reporter mRNA halflife to between 2.2 and 4.4 hours. Mutation of all AREs increased mRNA half life to 8.8 hours, suggesting that all AREs have some effect, but that specific AREs may have key roles in stability regulation. A labeled RNA oligonucleotide derived from the most influential ARE was utilized to purify TSP-1 AREbinding proteins. The AU-binding protein AUF1 was shown to associate with this motif. These studies reveal that AREs in the 3′UTR control TSP-1 mRNA stability and that the RNA binding protein AUF1 participates in this control. These studies suggest that ARE-dependent control of TSP-1 mRNA stability may represent an important component in the control of TSP-1 gene expression. [ABSTRACT FROM AUTHOR]

n endogenous timing mechanism, the circadian clock, causes rhythmic expression of a considerable fraction of the genome of most organisms to optimally align physiology and behavior with their environment. Circadian clocks are self-sustained oscillators primarily based on transcriptional feedback loops and post-translational modification of clock proteins. It is increasingly becoming clear that regulation at the RNA level strongly impacts the cellular circadian transcriptome and proteome as well as the oscillator mechanism itself. This review focuses on posttranscriptional events, discussing RNA-binding proteins that, by influencing the timing of pre-mRNA splicing, polyadenylation and RNA decay, shape rhythmic expression profiles. Furthermore, recent findings on the contribution of microRNAs to orchestrating circadian rhythms are summarized. [ABSTRACT FROM AUTHOR]

RNA-binding proteins are key regulators of plant gene expression. Consistent with this, the Arabidopsis genome encodes many RNA-binding proteins that are genetically required for normal development and for responding to environmental changes. However, the direct RNA targets and RNA processing events that these RNA-binding proteins control are poorly understood. In order to facilitate the functional characterization of RNA-binding proteins, we have applied the RNA immunoprecipitation assay to Arabidopsis. Working with the U2B″–U2 snRNA interaction as a model experimental system, we show that treatment of intact plants with formaldehyde allows immunocapture of U2 snRNA using antibodies that recognize U2B″ fused to the generic GFP tag. When coupled with recent developments in whole-genome tiling arrays and high-throughput next-generation sequencing, this combination of procedures and technology has the potential to allow systematic functional analysis of plant RNA-binding proteins. [ABSTRACT FROM AUTHOR]

Background and purpose: Bepridil is an anti-arrhythmic agent with anti-electrical remodelling effects that target many cardiac ion channels, including the voltage-gated Na+ channel. However, long-term effects of bepridil on the Na+ channel remain unclear. We explored the long-term effect of bepridil on the Na+ channel in isolated neonatal rat cardiomyocytes and in a heterologous expression system of human Nav1.5 channel. Experimental approach: Na+ currents were recorded by whole-cell voltage-clamp technique. Na+ channel message and protein were evaluated by real-time RT-PCR and Western blot analysis. Key results: Treatment of cardiomyocytes with 10 µmol·L−1 bepridil for 24 h augmented Na+ channel current ( INa) in a dose- and time-dependent manner. This long-term effect of bepridil was mimicked or masked by application of W-7, a calmodulin inhibitor, but not KN93 [2-[N-(2-hydroxyethyl)-N-(4-methoxy benzenesulphonyl)]-amino-N-(4-chlorocinnamyl)-N-methylbenzylamine], a Ca2+/calmodulin-dependent kinase inhibitor. During inhibition of protein synthesis by cycloheximide, the INa increase due to bepridil was larger than the increase without cycloheximide. Bepridil and W-7 significantly slowed the time course of Nav1.5 protein degradation in neonatal cardiomyocytes, although the mRNA levels of Nav1.5 were not modified. Bepridil and W-7 did not increase INa any further in the presence of the proteasome inhibitor MG132 [N-[(phenylmethoxy)carbonyl]-L-leucyl-N-[(1S)-1-formyl-3-methylbutyl]-L-leucinamide]. Bepridil, W-7 and MG132 but not KN93 significantly decreased 20S proteasome activity in a concentration-dependent manner. Conclusions and implications: We conclude that long-term exposure of cardiomyocytes to bepridil at therapeutic concentrations inhibits calmodulin action, which decreased degradation of the Nav1.5 α-subunit, which in turn increased Na+ current. [ABSTRACT FROM AUTHOR]

Antibody-mediated immunity is initiated by B cell differentiation into multiple cell subsets, including plasmablast, memory, and germinal center (GC) cells. B cell differentiation trajectories are determined by transcription factors, yet very few mechanisms that specifically determine early B cell fates have been described. Here, we report a post-transcriptional mechanism that suppresses the plasmablast genetic program and promotes GC B cell fate commitment. Single-cell RNA-sequencing analysis reveals that antigen-specific B cell precursors at the pre-GC stage upregulate YTHDF2, which enhances the decay of methylated transcripts. Ythdf2-deficient B cells exhibit intact proliferation and activation, whereas differentiation into GC B cells is blocked. Mechanistically, B cells require YTHDF2 to attenuate the plasmablast genetic program during GC seeding, and transcripts of key plasmablast-regulating genes are methylated and bound by YTHDF2. Collectively, this study reveals how post-transcriptional suppression of gene expression directs appropriate B cell fate commitment during initiation of the adaptive immune response. [Display omitted] • scRNA-seq of antigen-specific B cells reveals differentiation trajectories • YTHDF2 is expressed by early-responding B cells and facilitates germinal center seeding • YTHDF2 binds mRNAs of plasma cell-associated genes and suppresses their expression • Germinal center formation does not depend on YTHDF1 and YTHDF3 Using scRNA-seq and transgenic mice, Grenov et al. demonstrate that YTHDF2 is expressed by early-responding antigen-specific B cells and promotes their differentiation into germinal center B cells through the repression of genes associated with plasma cell formation, while other YTHDF paralogs are dispensable for the humoral immune response. [ABSTRACT FROM AUTHOR]

There is a universal requirement for post-translational regulatory mechanisms in circadian clock systems. Previous work in Drosophila has identified several kinases, phosphatases, and an E3 ligase that are critical for determining the nuclear translocation and/or stability of clock proteins. The present study evaluated the function of p90 ribosomal S6 kinase (RSK) in the Drosophila circadian system. In mammals, RSK1 is a light- and clock-regulated kinase known to be activated by the mitogen-activated protein kinase pathway, but there is no direct evidence that it functions as a component of the circadian system. Here, we show that Drosophila S6KIIRNAdisplays rhythms in abundance, indicative of circadian control. Importantly, an S6KII null mutant exhibits a short-period circadian phenotype that can be rescued by expression of the wild-type gene in clock neurons, indicating a role for S6KII in the molecular oscillator. Peak PER clock protein expression is elevated in the mutant, indicative of enhanced stability, whereas per mRNA level is decreased, consistent with enhanced feedback repression. Gene reporter assays show that decreased S6KII is associated with increased PER repression. Surprisingly, we demonstrate a physical interaction between S6KII and the casein kinase 2 regulatory subunit (CK2β), suggesting a functional relationship between the two kinases. In support of such a relationship, there are genetic interactions between S6KII and CK2 mutations, in vivo, which indicate that CK2 activity is required for S6KII action. We propose that the two kinases cooperate within clock neurons to fine-tune circadian period, improving the precision of the clock mechanism. [ABSTRACT FROM AUTHOR]

Dietary zinc deficiency in mice is accompanied by enhanced expression of the zinc uptake transporter Slc39a4 ( Zip4) and repressed expression of Slc39a5 ( Zip5) in tissues which regulate zinc homeostasis (intestine, pancreas and visceral yolk sac). Herein, mechanisms controlling this differential expression were investigated. The induction of Zip4 mRNA during zinc deficiency, and its repression in response to zinc repletion were found to reflect changes in Zip4 mRNA stability and not changes in the relative rate of transcription of this gene. During zinc deficiency, ZIP4 protein levels are increased and this protein is localized on the apical membranes. Administration of an oral gavage of zinc caused ZIP4 internalization and degradation in enterocytes and visceral endoderm cells. Similarly, ZIP4 is induced by zinc deficiency in cultured mouse Hepa cells and is rapidly degraded in response to added zinc. Zip5 mRNA abundance does not change in response to zinc, but the translation of this mRNA was found to be zinc-responsive. During zinc deficiency, Zip5 mRNA remains associated with polysomes, while the protein is internalized and degraded in enterocytes, acinar cells and endoderm cells. After zinc-gavage, ZIP5 is rapidly resynthesized and targeted to the basolateral membranes of these cell types. [ABSTRACT FROM AUTHOR]

Abstract: The mechanisms regulating expression of breast cancer 1 (BRCA1) are not fully characterised. By studying regulation of endogenous BRCA1 in human epithelial cell lines and during cell cycle progression, we provide evidence to suggest BRCA1 is regulated post-transcriptionally at the level of messenger RNA stability. We also show that RNA-binding proteins associate with an AU-rich, cis-active sequence of the BRCA1 3′untranslated region in a cell cycle-dependent manner. Our data identify a new post-transcriptional regulatory axis and a novel mechanism for modulating the levels of BRCA1 protein, with possible implications for understanding the mechanisms underlying BRCA1 repression in breast cancer. [Copyright &y& Elsevier]

This review addresses the scope of influence of mRNA decay on cellular functions and its potential role in normal and malignant hematopoiesis. Evidence is emerging that leukemic oncogenes and hematopoietic cytokines interact with mRNA decay pathways. These pathways can co-regulate functionally related genes through specific motifs in the 3′-untranslated region of targeted transcripts. The steps that link external stimuli to transcript turnover are not fully understood, but include subcellular relocalization or post-transcriptional modification of specific transcript-stabilizing or -destabilizing proteins. Improper functioning of these regulators of mRNA turnover can impede normal cellular differentiation or promote cancers. By delineating how subsets of transcripts decay in synchrony during normal hematopoiesis, it may be possible to determine whether this post-transcriptional regulatory pathway is hijacked in leukemogenesis.Leukemia (2007) 21, 1158–1171. doi:10.1038/sj.leu.2404656; published online 29 March 2007 [ABSTRACT FROM AUTHOR]

Transforming growth factor beta (TGF-β) is a biologically multipotent regulatory protein implicated in functions that include the regulation of cellular growth, differentiation, extracellular matrix formation, and wound healing. It also plays a role in the pathologies of Alzheimer's disease, cancer and autoimmune disorders. TGF-β modulates gene expression by affecting transcriptional activation and mRNA turnover rate. Steady-state mRNA levels depend on both the transcriptional activity and mRNA half-life. The stability of mRNA can be modified by the binding of trans -acting factors to cis -elements on the message. These can protect the mRNA from cleavage by RNAses, or they may promote mRNA cleavage. Changes in mRNA stability can lead to changes in the proteome and subsequently in cellular metabolism. The SMAD family of proteins has been implicated in the transduction of the TGF-β signal, where they regulate transcriptional activity. This review attempts to provide new insights into the role played by TGF-β in the regulation of mRNA turnover. [ABSTRACT FROM AUTHOR]

Expression of the pea plastocyanin gene ( PetE) is regulated by light and plastid signals. Previous work indicated that light and plastid regulation of pea PetE operates post-transcriptionally in transgenic tobacco, and requires the correct 5′ terminus of the PetE transcript and the PetE-coding region. The post-transcriptional light and plastid regulation of pea PetE has now been demonstrated to operate in transgenic Arabidopsis, where in contrast the endogenous PETE gene is regulated transcriptionally. Transgenic tobacco seedlings containing constructs with progressive 3′ deletions of the PetE-coding region fused to the luciferase ( Luc) reporter gene demonstrate that the first 60 nucleotides of the coding region are sufficient for regulated accumulation of Luc transcripts by light and plastid signalling pathways affected by treatment with norflurazon and lincomycin. PetE constructs containing premature stop codons were generated to investigate whether translation has a role in light or plastid regulation. Insertion of a stop codon in place of the second codon of the PetE-coding region diminished both light and plastid regulation of PetE transcripts, whereas stop codons inserted later in the transcript had no effect on light or plastid regulation. These experiments indicate that the 5′ end of the plastocyanin-coding region contains sequences important for regulation by light and plastid signals. [ABSTRACT FROM AUTHOR]

The 5' untranslated region (5'UTR) of the psbA mRNA (psbA encodes the PSII reaction center protein, D1) is a key site for RNA—protein interactions in the post-transcriptional regulation of gene expression. In this study, we mapped the major psbA mRNA 5'-terminus at -77 nt, and two minor termini clusters centered at -48 and -64 nt, upstream from the psbA translational start codon of Arabidopsis thaliana. RNA mobility shift, RNase protection and UV-crosslinking assays were used to characterize the interaction of chloroplast proteins with the RNA 5'UTR. RNA—protein interactions depended upon a thermolabile secondary structure and specific sequences in a 35 nt region of the 5'UTR, which were 80% conserved with the psbA 5'UTRs from five other plants. Major and minor proteins of 43- and 30-kDa, respectively, were detected by UV-crosslinking to RNA. Oxidizing conditions abolished the association of the proteins with the 5'UTR, while RNA-binding activity was recovered upon incubation with a reductant. Based on these findings, we hypothesize that post-transcriptional regulation of psbA gene expression in chloroplasts of vascular plants involves redox-dependent interactions between specific sequences in the 5'UTR and 43- and 30-kDa RNA-binding proteins. [ABSTRACT FROM AUTHOR]

Plastid gene expression involves many post-transcriptional maturation steps resulting in a complex transcriptome composed of multiple isoforms. Although short-read RNA-Seq has considerably improved our understanding of the molecular mechanisms controlling these processes, it is unable to sequence full-length transcripts. This information is crucial, however, when it comes to understanding the interplay between the various steps of plastid gene expression. Here, we describe a protocol to study the plastid transcriptome using nanopore sequencing. In the leaf of Arabidopsis thaliana, with about 1.5 million strand-specific reads mapped to the chloroplast genome, we could recapitulate most of the complexity of the plastid transcriptome (polygenic transcripts, multiple isoforms associated with post-transcriptional processing) using virtual Northern blots. Even if the transcripts longer than about 2500 nucleotides were missing, the study of the co-occurrence of editing and splicing events identified 42 pairs of events that were not occurring independently. This study also highlighted a preferential chronology of maturation events with splicing happening after most sites were edited. [ABSTRACT FROM AUTHOR]

The level of the heat inducible hsp70 protein (hsp72) has been shown to be elevated in the peripheral blood mononuclear cells of a subset of SLE patients. We show that this increased level of hsp70 is not dependent on enhanced transcription of the hsp70 gene or elevated levels of the hsp70 mRNA, neither of which are observed in SLE patients with enhanced protein levels. This indicates that post-transcriptional processes involving either improved translatability of the hsp70 mRNA or increased protein stability are responsible for the observed increase in protein levels in these patients. [ABSTRACT FROM PUBLISHER]