scholarly journals A Novel DCL2-Dependent Micro-Like RNA Vm-PC-3p-92107_6 Affects Pathogenicity by Regulating the Expression of Vm-VPS10 in Valsa mali

2021 ◽  
Vol 12 ◽  
Author(s):  
Feiran Guo ◽  
Jiahao Liang ◽  
Ming Xu ◽  
Gao Zhang ◽  
Lili Huang ◽  
...  
Keyword(s):  
Small Rnas ◽  
Target Genes ◽  
Pathogenic Fungus ◽  
Small Interfering Rnas ◽  
Wild Type ◽  
Over Expression ◽  
In The Wild ◽  
Vacuolar Protein ◽  
And Function ◽  
Valsa Mali

Dicer proteins are mainly responsible for generating small RNAs (sRNAs), which are involved in gene silencing in most eukaryotes. In previous research, two DCL proteins in Valsa mali, the pathogenic fungus causing apple tree Valsa canker, were found associated with both the pathogenicity and generation of sRNAs. In this study, the differential expression of small interfering RNAs (siRNAs) and miRNA-like RNAs (milRNAs) was analyzed based on the deep sequencing of the wild type and Vm-DCL2 mutant, respectively. Overall, the generation of 40 siRNAs and 18 milRNAs was evidently associated with Vm-DCL2. The target genes of milRNAs were then identified using degradome sequencing; according to the prediction results, most candidate targets are related to pathogenicity. Further, expression of Vm-PC-3p-92107_6 was confirmed in the wild type but not in the Vm-DCL2 mutant. Moreover, the pathogenicity of Vm-PC-3p-92107_6 deletion mutants (ΔVm-PC-3p-92107_6) and the over-expression transformants (Vm-PC-3p-92107_6-OE) was significantly increased and decreased, respectively. Based on those degradome results, vacuolar protein sorting 10 (Vm-VPS10) was identified as the target of Vm-PC-3p-92107_6. Co-expression analysis in tobacco leaves further confirmed that Vm-PC-3p-92107_6 could suppress the expression of Vm-VPS10. Meanwhile, the expression levels of Vm-PC-3p-92107_6 and Vm-VPS10 displayed divergent trends in ΔVm-PC-3p-92107_6 and Vm-PC-3p-92107_6-OE, respectively. Perhaps most importantly, ΔVm-VPS10 featured a significant reduction in pathogenicity. Taken together, our results indicate that a DCL2-dependent milRNA Vm-PC-3p-92107_6 plays roles in pathogenicity by regulating the expression of Vm-VPS10. This study lays a foundation for the comprehensive analysis of pathogenic mechanisms of V. mali and deepens our understanding of the generation and function of fungal sRNA.

scholarly journals Small RNAs in parasitic nematodes – forms and functions

Parasitology ◽  
2019 ◽  
Vol 147 (8) ◽  
pp. 855-864
Author(s):  
Collette Britton ◽  
Roz Laing ◽  
Eileen Devaney
Keyword(s):  
Gene Expression ◽  
Small Rna ◽  
Small Rnas ◽  
Target Genes ◽  
Parasitic Nematodes ◽  
Small Rna Sequencing ◽  
Small Interfering Rnas ◽  
Rna Sequences ◽  
C Elegans

AbstractSmall RNAs are important regulators of gene expression. They were first identified in Caenorhabditis elegans, but it is now apparent that the main small RNA silencing pathways are functionally conserved across diverse organisms. Availability of genome data for an increasing number of parasitic nematodes has enabled bioinformatic identification of small RNA sequences. Expression of these in different lifecycle stages is revealed by small RNA sequencing and microarray analysis. In this review we describe what is known of the three main small RNA classes in parasitic nematodes – microRNAs (miRNAs), Piwi-interacting RNAs (piRNAs) and small interfering RNAs (siRNAs) – and their proposed functions. miRNAs regulate development in C. elegans and the temporal expression of parasitic nematode miRNAs suggest modulation of target gene levels as parasites develop within the host. miRNAs are also present in extracellular vesicles released by nematodes in vitro, and in plasma from infected hosts, suggesting potential regulation of host gene expression. Roles of piRNAs and siRNAs in suppressing target genes, including transposable elements, are also reviewed. Recent successes in RNAi-mediated gene silencing, and application of small RNA inhibitors and mimics will continue to advance understanding of small RNA functions within the parasite and at the host–parasite interface.

scholarly journals Arabidopsis thaliana cbp80, c2h2, and flk Knockout Mutants Accumulate Increased Amounts of Circular RNAs

Cells ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 1937
Author(s):  
Anna Philips ◽  
Katarzyna Nowis ◽  
Michal Stelmaszczuk ◽  
Jan Podkowiński ◽  
Luiza Handschuh ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Circular Rnas ◽  
Wild Type ◽  
Proper Order ◽  
Knockout Mutants ◽  
In The Wild ◽  
And Function

Circular RNAs (circRNAs) are the products of the non-canonical splicing of pre-mRNAs. In contrast to humans and animals, our knowledge of the biogenesis and function of circRNAs in plants is very scarce. To identify proteins involved in plant circRNA generation, we characterized the transcriptomes of 18 Arabidopsis thaliana knockout mutants for genes related to splicing. The vast majority (>90%) of circRNAs were formed in more than one variant; only a small fraction of circRNAs was mutant-specific. Five times more circRNA types were identified in cbp80 and three times more in c2h2 mutants than in the wild-type. We also discovered that in cbp80, c2h2 and flk mutants, the accumulation of circRNAs was significantly increased. The increased accumulation of circular transcripts was not accompanied by corresponding changes in the accumulation of linear transcripts. Our results indicate that one of the roles of CBP80, C2H2 and FLK in splicing is to ensure the proper order of the exons. In the absence of one of the above-mentioned factors, the process might be altered, leading to the production of circular transcripts. This suggests that the transition toward circRNA production can be triggered by factors sequestering these proteins. Consequently, the expression of linear transcripts might be regulated through circRNA production.

scholarly journals The interplay between small RNA pathways shapes chromatin landscapes in C. elegans

2018 ◽  
Author(s):  
Ekaterina Gushchanskaia ◽  
Ruben Esse ◽  
Qicheng Ma ◽  
Nelson Lau ◽  
Alla Grishok
Keyword(s):  
Small Rnas ◽  
Target Genes ◽  
Noncoding Rnas ◽  
Small Interfering Rnas ◽  
Loss Of Function ◽  
Partial Loss ◽  
Rna Dependent Rna Polymerase ◽  
C Elegans ◽  
Highly Expressed Genes ◽  
Rna Pathways

ABSTRACTThe nematode C. elegans contains several types of endogenous small interfering RNAs (endo-siRNAs) produced by RNA-dependent RNA polymerase (RdRP) complexes. Both “silencing” siRNAs bound by Worm-specific Argonautes (WAGO) and “activating” siRNAs bound by the CSR-1 Argonaute require the DRH-3 helicase, an RdRP component. Here we show that, in the drh-3(ne4253) mutant deficient in RdRP-produced secondary endo-siRNAs, the silencing histone mark H3K9me3 is largely depleted, whereas in the csr-1 partial loss-of-function mutant this mark is ectopically deposited on CSR-1 target genes. Moreover, we observe ectopic H3K9me3 at enhancer elements in both drh-3 and csr-1 partial loss-of-function mutants and describe small RNAs matching enhancers. Finally, we detect accumulation of H3K27me3 at highly expressed genes in the drh-3(ne4253) mutant, which correlates with their reduced transcription. Our study shows that when abundant RdRP-produced siRNAs are depleted, there is ectopic elevation of noncoding RNAs linked to increase in silencing chromatin marks. Moreover, our results suggest that enhancer small RNAs may guide local H3K9 methylation.

scholarly journals A Modified Actin (Gly65Val Substitution) Expressed in Cotton Disrupts Polymerization of Actin Filaments Leading to the Phenotype of Ligon Lintless-1 (Li1) Mutant

2021 ◽  
Vol 22 (6) ◽  
pp. 3000
Author(s):  
Yuefen Cao ◽  
Hui Huang ◽  
Yanjun Yu ◽  
Huaqin Dai ◽  
Huanfeng Hao ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Actin Filaments ◽  
Textile Industry ◽  
Actin Polymerization ◽  
Natural Fibers ◽  
Wild Type ◽  
Over Expression ◽  
High Resolution Mapping ◽  
In The Wild ◽  
Resolution Mapping

Dynamic remodeling of the actin cytoskeleton plays a central role in the elongation of cotton fibers, which are the most important natural fibers in the global textile industry. Here, a high-resolution mapping approach combined with comparative sequencing and a transgenic method revealed that a G65V substitution in the cotton actin Gh_D04G0865 (GhACT17D in the wild-type) is responsible for the Gossypium hirsutum Ligon lintless-1 (Li1) mutant (GhACT17DM). In the mutant GhACT17DM from Li1 plant, Gly65 is substituted with valine on the lip of the nucleotide-binding domain of GhACT17D, which probably affects the polymerization of F-actin. Over-expression of GhACT17DM, but not GhACT17D, driven by either a CaMV35 promoter or a fiber-specific promoter in cotton produced a Li1-like phenotype. Compared with the wild-type control, actin filaments in Li1 fibers showed higher growth and shrinkage rates, decreased filament skewness and parallelness, and increased filament density. Taken together, our results indicate that the incorporation of GhACT17DM during actin polymerization disrupts the establishment and dynamics of the actin cytoskeleton, resulting in defective fiber elongation and the overall dwarf and twisted phenotype of the Li1 mutant.

scholarly journals Construction of afurnull mutant and RNA-sequencing provide deeper global understanding of theAliivibrio salmonicidaFur regulon

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3461 ◽  
Author(s):  
Sunniva Katharina Thode ◽  
Cecilie Bækkedal ◽  
Jenny Johansson Söderberg ◽  
Erik Hjerde ◽  
Hilde Hansen ◽  
...  
Keyword(s):  
Rna Sequencing ◽  
Iron Homeostasis ◽  
Target Genes ◽  
Iron Acquisition ◽  
Null Mutant ◽  
Wild Type ◽  
Knock Out ◽  
In The Wild ◽  
Global Understanding ◽  
Main Regulator

BackgroundThe ferric uptake regulator (Fur) is a transcription factor and the main regulator of iron acquisition in prokaryotes. When bound to ferric iron, Fur recognizes its DNA binding site and generally executes its function by repressing transcription of its target genes. Due to its importance in virulence, the Fur regulon is well studied for several model bacteria. In our previous work, we used computational predictions and microarray to gain insights into Fur-regulation inAliivibrio salmonicida, and have identified a number of genes and operons that appear to be under direct control of Fur. To provide a more accurate and deeper global understanding of the biological role of Fur we have now generated anA. salmonicida furknock-out strain and used RNA-sequencing to compare gene expression between the wild-type andfurnull mutant strains.ResultsAnA. salmonicida furnull mutant strain was constructed. Biological assays demonstrate that deletion offurresults in loss of fitness, with reduced growth rates, and reduced abilities to withstand low-iron conditions, and oxidative stress. When comparing expression levels in the wild-type and thefurnull mutant we retrieved 296 differentially expressed genes distributed among 18 of 21 functional classes of genes. A gene cluster encoding biosynthesis of the siderophore bisucaberin represented the highest up-regulated genes in thefurnull mutant. Other highly up-regulated genes all encode proteins important for iron acquisition. Potential targets for the RyhB sRNA was predicted from the list of down-regulated genes, and significant complementarities were found between RyhB and mRNAs of thefur,sodB,cysNand VSAL_I0422 genes. Other sRNAs with potential functions in iron homeostasis were identified.ConclusionThe present work provides by far the most comprehensive and deepest understanding of the Fur regulon inA. salmonicidato date. Our data also contribute to a better understanding of how Fur plays a key role in iron homeostasis in bacteria in general, and help to show how Fur orchestrates iron uptake when iron levels are extremely low.

scholarly journals HIF1α Protein Stability Is Increased by Acetylation at Lysine 709

2012 ◽  
Vol 287 (42) ◽  
pp. 35496-35505 ◽  
Author(s):  
Hao Geng ◽  
Qiong Liu ◽  
Changhui Xue ◽  
Larry L. David ◽  
Tomasz M. Beer ◽  
...  
Keyword(s):  
Protein Stability ◽  
Cancer Cells ◽  
Target Genes ◽  
Protein A ◽  
Growth Arrest ◽  
Lysine Acetylation ◽  
Protein Acetylation ◽  
Wild Type ◽  
Transcriptional Complex ◽  
And Function

Lysine acetylation regulates protein stability and function. p300 is a component of the HIF-1 transcriptional complex and positively regulates the transactivation of HIF-1. Here, we show a novel molecular mechanism by which p300 facilitates HIF-1 activity. p300 increases HIF-1α (HIF1α) protein acetylation and stability. The regulation can be opposed by HDAC1, but not by HDAC3, and is abrogated by disrupting HIF1α-p300 interaction. Mechanistically, p300 specifically acetylates HIF1α at Lys-709, which increases the protein stability and decreases polyubiquitination in both normoxia and hypoxia. Compared with the wild-type protein, a HIF1α K709A mutant protein is more stable, less polyubiquitinated, and less dependent on p300. Overexpression of the HIF1α wild-type or K709A mutant in cancer cells lacking the endogenous HIF1α shows that the K709A mutant is transcriptionally more active toward the HIF-1 reporter and some endogenous target genes. Cancer cells containing the K709A mutant are less sensitive to hypoxia-induced growth arrest than the cells containing the HIF1α wild-type. Taken together, these data demonstrate a novel biological consequence upon HIF1α-p300 interaction, in which HIF1α can be stabilized by p300 via Lys-709 acetylation.

Suppression of myeloid transcription factors and induction of STAT response genes by AML-specific Flt3 mutations

Blood ◽  
2003 ◽  
Vol 101 (8) ◽  
pp. 3164-3173 ◽  
Author(s):  
Masao Mizuki ◽  
Joachim Schwäble ◽  
Claudia Steur ◽  
Chunaram Choudhary ◽  
Shuchi Agrawal ◽  
...  
Keyword(s):  
Target Genes ◽  
Wild Type ◽  
Transcriptional Program ◽  
Serine Threonine Kinase ◽  
Myeloid Progenitor ◽  
Genome Wide ◽  
Flt3 Mutations ◽  
Myeloid Progenitor Cells ◽  
And Function ◽  
Microarray Expression

Abstract The receptor tyrosine kinase Flt3 is expressed and functionally important in early myeloid progenitor cells and in the majority of acute myeloid leukemia (AML) blasts. Internal tandem duplications (ITDs) in the juxtamembrane domain of the receptor occur in 25% of AML cases. Previously, we have shown that these mutations activate the receptor and induce leukemic transformation. In this study, we performed genome-wide parallel expression analyses of 32Dcl3 cells stably transfected with either wild-type or 3 different ITD isoforms of Flt3. Comparison of microarray expression analyses revealed that 767 of 6586 genes differed in expression between FLT3-WT– and FLT3-ITD–expressing cell lines. The target genes of mutationally activated Flt3 resembled more closely those of the interleukin 3 (IL-3) receptor than those of ligand-activated Flt3. The serine-threonine kinase Pim-2 was up-regulated on the mRNA and the protein level in Flt3-ITD–expressing cells. Further experiments indicated that Pim-2 function was important for clonal growth of 32D cells. Several genes repressed by the mutations were found to be involved in myeloid gene regulation. Pu.1 and C/EBPα, both induced by ligand-activation of wild-type Flt3, were suppressed in their expression and function by the Flt3 mutations. In conclusion, internal tandem duplication mutations of Flt3 activate transcriptional programs that partially mimic IL-3 activity. Interestingly, other parts of the transcriptional program involve novel, IL-3–independent pathways that antagonize differentiation-inducing effects of wild-type Flt3. The identification of the transcriptional program induced by ITD mutations should ease the development of specific therapies.

scholarly journals Ester-Producing Mechanism of Ethanol O-acyltransferaseEHT1Gene inPichia pastorisfrom Shanxi Aged Vinegar

2019 ◽  
Vol 2019 ◽  
pp. 1-9
Author(s):  
Jia Chen ◽  
Ruipeng Nan ◽  
Rufu Wang ◽  
Lixin Zhang ◽  
Junfeng Shi
Keyword(s):  
Fatty Acids ◽  
Esterase Activity ◽  
Volatile Fatty Acids ◽  
The Other ◽  
Fatty Acid Esters ◽  
Wild Type ◽  
Fermentation Products ◽  
Over Expression ◽  
In The Wild ◽  
Methyl Nonanoate

The ethanol O-acyltransferaseEHT1is an important element of key signaling pathways and is widely expressed in yeast strains. In this study, we investigated the expression ofEHT1 in the overexpression lines or knockout system ofPichia pastorisusing qRT-PCR and western blotting. The amount of total protein was determined using the Bradford method; the esterase activity was determined using p-nitrophenyl acetate as a substrate, and the production of volatile fatty acids in wild-type, knockout, and over-expression systems was detected using SPME GC-MS. The esterase activity ofEHT1-knockoutP. pastoriswas significantly lower than that in wild type (P<0.01), and the activities of esterase in threeEHT1-overexpressing strains—OE-1, OE-2, and OE-3—were significantly higher than those in wild type (P<0.01). In theEHT1-knockout strain products, the contents of nine volatile fatty acids were significantly lower than those in wild type (P<0.01), and the relative percentages of three fatty acids, methyl nonanoate, methyl decanoate, and ethyl caprate, were significantly lower than those in the other six species in the wild-type and knockout groups (P<0.05). The nine volatile fatty acids in the fermentation products of the overexpressedEHT1 gene were significantly higher than those in the wild-type group (P<0.01). The relative percentages of the three fatty acid esters, methyl nonanoate, methyl caprate, and ethyl caprate, were significantly higher than those in the other six species (P<0.05).EHT1 plays an important regulatory role in esterase activity and the production of medium-chain volatile fatty acids.

scholarly journals Regulation of Polyhydroxybutyrate Accumulation in Sinorhizobium meliloti by the Trans-Encoded Small RNA MmgR

2017 ◽  
Vol 199 (8) ◽  
Author(s):  
Antonio Lagares ◽  
Germán Ceizel Borella ◽  
Uwe Linne ◽  
Anke Becker ◽  
Claudio Valverde
Keyword(s):  
Small Rnas ◽  
Sinorhizobium Meliloti ◽  
Reducing Power ◽  
Negative Regulator ◽  
Fine Tuning ◽  
Biological Role ◽  
Wild Type ◽  
Content Type ◽  
In The Wild

ABSTRACT Riboregulation has a major role in the fine-tuning of multiple bacterial processes. Among the RNA players, trans-encoded untranslated small RNAs (sRNAs) regulate complex metabolic networks by tuning expression from multiple target genes in response to numerous signals. In Sinorhizobium meliloti, over 400 sRNAs are expressed under different stimuli. The sRNA MmgR (standing for Makes more granules Regulator) has been of particular interest to us since its sequence and structure are highly conserved among the alphaproteobacteria and its expression is regulated by the amount and quality of the bacterium's available nitrogen source. In this work, we explored the biological role of MmgR in S. meliloti 2011 by characterizing the effect of a deletion of the internal conserved core of mmgR (mmgR Δ33–51). This mutation resulted in larger amounts of polyhydroxybutyrate (PHB) distributed into more intracellular granules than are found in the wild-type strain. This phenotype was expressed upon cessation of balanced growth owing to nitrogen depletion in the presence of surplus carbon (i.e., at a carbon/nitrogen molar ratio greater than 10). The normal PHB accumulation was complemented with a wild-type mmgR copy but not with unrelated sRNA genes. Furthermore, the expression of mmgR limited PHB accumulation in the wild type, regardless of the magnitude of the C surplus. Quantitative proteomic profiling and quantitative reverse transcription-PCR (qRT-PCR) revealed that the absence of MmgR results in a posttranscriptional overexpression of both PHB phasin proteins (PhaP1 and PhaP2). Together, our results indicate that the widely conserved alphaproteobacterial MmgR sRNA fine-tunes the regulation of PHB storage in S. meliloti. IMPORTANCE High-throughput RNA sequencing has recently uncovered an overwhelming number of trans-encoded small RNAs (sRNAs) in diverse prokaryotes. In the nitrogen-fixing alphaproteobacterial symbiont of alfalfa root nodules Sinorhizobium meliloti, only four out of hundreds of identified sRNA genes have been functionally characterized. Thus, uncovering the biological role of sRNAs currently represents a major issue and one that is particularly challenging because of the usually subtle quantitative regulation contributed by most characterized sRNAs. Here, we have characterized the function of the broadly conserved alphaproteobacterial sRNA gene mmgR in S. meliloti. Our results strongly suggest that mmgR encodes a negative regulator of the accumulation of polyhydroxybutyrate, the major carbon and reducing power storage polymer in S. meliloti cells growing under conditions of C/N overbalance.

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