scholarly journals An Ash1-Like Protein MoKMT2H Null Mutant Is Delayed for Conidium Germination and Pathogenesis inMagnaporthe oryzae

2016 ◽  
Vol 2016 ◽  
pp. 1-9 ◽  
Author(s):  
Zhaojun Cao ◽  
Yue Yin ◽  
Xuan Sun ◽  
Jun Han ◽  
Qing peng Sun ◽  
...  
Keyword(s):  
Target Gene ◽  
Hyphal Growth ◽  
Gene Replacement ◽  
Phytopathogenic Fungus ◽  
Null Mutant ◽  
Lesion Formation ◽  
Target Gene Replacement ◽  
Conidium Germination ◽  
Rice Leaves ◽  
And Function

Ash1 is a known H3K36-specific histone demethylase that is required for normal Hox gene expression and fertility inDrosophilaand mammals. However, little is known about the expression and function of the fungal ortholog of Ash1 in phytopathogenic fungusMagnaporthe oryzae. Here we report that MoKMT2H, an Ash1-like protein, is required for conidium germination and virulence in rice. We obtainedMoKMT2Hnull mutant (ΔMoKMT2H) using a target gene replacement strategy. In theΔMoKMT2Hnull mutants, global histone methyltransferase modifications (H3K4me3, H3K9me3, H3K27me3, and H3K36me2/3) of the genome were unaffected. TheΔMoKMT2Hmutants showed no defect in vegetative hyphal growth, conidium morphology, conidiation, or disease lesion formation on rice leaves. However, theMoKMT2Hdeletion mutants were delayed for conidium germination and consequently had decreased virulence. Taken together, our results indicated that MoKMT2H plays an important role in conidium germination during appressorium formation in the rice blast fungus and perhaps other pathogenic plant fungi.

scholarly journals Glia-derived exosomal miR-274 targets Sprouty in trachea and synaptic boutons to modulate growth and responses to hypoxia

2019 ◽  
Vol 116 (49) ◽  
pp. 24651-24661 ◽  
Author(s):  
Yi-Wei Tsai ◽  
Hsin-Ho Sung ◽  
Jian-Chiuan Li ◽  
Chun-Yen Yeh ◽  
Pei-Yi Chen ◽  
...  
Keyword(s):  
Cell Growth ◽  
Target Cell ◽  
Target Gene ◽  
Mapk Signaling ◽  
Target Cells ◽  
Null Mutant ◽  
Specific Expression ◽  
Exosome Biogenesis ◽  
Synaptic Boutons ◽  
And Function

Secreted exosomal microRNAs (miRNAs) mediate interorgan/tissue communications by modulating target gene expression, thereby regulating developmental and physiological functions. However, the source, route, and function in target cells have not been formally established for specific miRNAs. Here, we show that glial miR-274 non-cell-autonomously modulates the growth of synaptic boutons and tracheal branches. Whereas the precursor form of miR-274 is expressed in glia, the mature form of miR-274 distributes broadly, including in synaptic boutons, muscle cells, and tracheal cells. Mature miR-274 is secreted from glia to the circulating hemolymph as an exosomal cargo, a process requiring ESCRT components in exosome biogenesis and Rab11 and Syx1A in exosome release. We further show that miR-274 can function in the neurons or tracheal cells to modulate the growth of synaptic boutons and tracheal branches, respectively. Also, miR-274 uptake into the target cells by AP-2–dependent mechanisms modulates target cell growth. In the target cells, miR-274 down-regulates Sprouty (Sty) through a targeting sequence at the sty 3′ untranslated region, thereby enhancing MAPK signaling and promoting cell growth. miR-274 expressed in glia of an mir-274 null mutant is released as an exosomal cargo in the circulating hemolymph, and such glial-specific expression resets normal levels of Sty and MAPK signaling and modulates target cell growth. mir-274 mutant larvae are hypersensitive to hypoxia, which is suppressed by miR-274 expression in glia or by increasing tracheal branches. Thus, glia-derived miR-274 coordinates growth of synaptic boutons and tracheal branches to modulate larval hypoxia responses.

scholarly journals Comparison and Validation of Putative Pathogenicity-Related Genes Identified by T-DNA Insertional Mutagenesis and Microarray Expression Profiling inMagnaporthe oryzae

2017 ◽  
Vol 2017 ◽  
pp. 1-7
Author(s):  
Ying Wang ◽  
Ying Wáng ◽  
Qi Tan ◽  
Ying Nv Gao ◽  
Yan Li ◽  
...  
Keyword(s):  
High Throughput ◽  
Expression Profiling ◽  
Insertional Mutagenesis ◽  
Target Gene ◽  
Gene Replacement ◽  
Knockout Mutants ◽  
Target Gene Replacement ◽  
Microarray Expression Profiling ◽  
Microarray Expression ◽  
Dna Insertional Mutagenesis

High-throughput technologies of functional genomics such as T-DNA insertional mutagenesis and microarray expression profiling have been employed to identify genes related to pathogenicity inMagnaporthe oryzae. However, validation of the functions of individual genes identified by these high-throughput approaches is laborious. In this study, we compared two published lists of genes putatively related to pathogenicity inM. oryzaeidentified by T-DNA insertional mutagenesis (comprising 1024 genes) and microarray expression profiling (comprising 236 genes), respectively, and then validated the functions of some overlapped genes between the two lists by knocking them out using the method of target gene replacement. Surprisingly, only 13 genes were overlapped between the two lists, and none of the four genes selected from the overlapped genes exhibited visible phenotypic changes on vegetative growth, asexual reproduction, and infection ability in their knockout mutants. Our results suggest that both of the lists might contain large proportions of unrelated genes to pathogenicity and therefore comparing the two gene lists is hardly helpful for the identification of genes that are more likely to be involved in pathogenicity as we initially expected.

scholarly journals CmAim24 Is Essential for Mitochondrial Morphology, Conidiogenesis, and Mycoparasitism in Coniothyrium minitans

2019 ◽  
Vol 86 (5) ◽  
Author(s):  
Xiaoxiang Yang ◽  
Huizhang Zhao ◽  
Chenwei Luo ◽  
Lei Du ◽  
Jiasen Cheng ◽  
...  
Keyword(s):  
Cellular Localization ◽  
Sequence Similarity ◽  
Critical Role ◽  
Gene Replacement ◽  
Phytopathogenic Fungus ◽  
Mitochondrial Morphology ◽  
Coniothyrium Minitans ◽  
High Sequence Similarity ◽  
Content Type ◽  
And Function

ABSTRACT Coniothyrium minitans is an important mycoparasite of the notorious phytopathogenic fungus Sclerotinia sclerotiorum. The mycoparasitism system of C. minitans-S. sclerotiorum is unique and important in probing fungi and fungal interactions. Here, we report a conidiation-deficient mutant, ZS-1TN1961, which was screened from a transfer DNA (T-DNA) insertional library of C. minitans. A single-copy gene, encoding a protein with high sequence similarity to Aim24 (altered inheritance of mitochondria protein 24) in Saccharomyces cerevisiae, was disrupted by T-DNA insertion in this mutant. Gene replacement and complementation experiments confirmed that mutants lacking CmAim24 exhibited significantly reduced conidial production and germination as well as reduced sclerotial mycoparasitic ability. Furthermore, cellular localization assays showed that CmAim24 localized to mitochondria, and abnormal mitochondria were observed in the ΔCmAim24 mutant. The ΔCmAim24 mutant exhibited significant accumulation of reactive oxygen species (ROS) and a reduced ATP content in mycelia. In summary, our results suggest that CmAim24 plays a key role in mitochondrial architecture and function, conidiogenesis, and mycoparasitism in C. minitans. IMPORTANCE Aim24 proteins are involved in mitochondrial biogenesis and accumulate between the two membranes of a mitochondrion. Their function in prokaryotes and filamentous fungi is as yet unknown. In the present study, we characterized an Aim24 protein, CmAim24, in the mycoparasite Coniothyrium minitans and proved its critical role in mitochondrial morphology and function, conidiogenesis, conidial germination, and mycoparasitism to S. sclerotiorum.

scholarly journals RecET recombination system driving chromosomal target gene replacement in Zymomonas mobilis

2017 ◽  
Vol 30 ◽  
pp. 118-124 ◽  
Author(s):  
Yan Wu ◽  
Tao Li ◽  
Qinghua Cao ◽  
Xuedan Li ◽  
Yizheng Zhang ◽  
...  
Keyword(s):  
Zymomonas Mobilis ◽  
Target Gene ◽  
Gene Replacement ◽  
Recombination System ◽  
Target Gene Replacement

Gene replacement with one-sided homologous recombination

1992 ◽  
Vol 12 (1) ◽  
pp. 360-367
Author(s):  
N Berinstein ◽  
N Pennell ◽  
C A Ottaway ◽  
M J Shulman
Keyword(s):  
Homologous Recombination ◽  
Dna Sequences ◽  
Mammalian Cells ◽  
Target Gene ◽  
Gene Replacement ◽  
Immunoglobulin Genes ◽  
Nonhomologous Recombination ◽  
Immunoglobulin Locus ◽  
Chromosomal Sequences ◽  
Dna Vectors

Homologous recombination is now routinely used in mammalian cells to replace endogenous chromosomal sequences with transferred DNA. Vectors for this purpose are traditionally constructed so that the replacement segment is flanked on both sides by DNA sequences which are identical to sequences in the chromosomal target gene. To test the importance of bilateral regions of homology, we measured recombination between transferred and chromosomal immunoglobulin genes when the transferred segment was homologous to the chromosomal gene only on the 3' side. In each of the four recombinants analyzed, the 5' junction was unique, suggesting that it was formed by nonhomologous, i.e., random or illegitimate, recombination. In two of the recombinants, the 3' junction was apparently formed by homologous recombination, while in the other two recombinants, the 3' junction as well as the 5' junction might have involved a nonhomologous crossover. As reported previously, we found that the frequency of gene targeting increases monotonically with the length of the region of homology. Our results also indicate that targeting with fragments bearing one-sided homology can be as efficient as with fragments with bilateral homology, provided that the overall length of homology is comparable. The frequency of these events suggests that the immunoglobulin locus is particularly susceptible to nonhomologous recombination. Vectors designed for one-sided homologous recombination might be advantageous for some applications in genetic engineering.

scholarly journals PIN1 Is an E2F Target Gene Essential for Neu/Ras-Induced Transformation of Mammary Epithelial Cells

2002 ◽  
Vol 22 (15) ◽  
pp. 5281-5295 ◽  
Author(s):  
Akihide Ryo ◽  
Yih-Cherng Liou ◽  
Gerburg Wulf ◽  
Masafumi Nakamura ◽  
Sam W. Lee ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cyclin D1 ◽  
Transcriptional Activation ◽  
Target Gene ◽  
Mammary Epithelial Cells ◽  
Cell Transformation ◽  
Mammary Tumorigenesis ◽  
Mammary Epithelial ◽  
Oncogenic Signaling ◽  
And Function

ABSTRACT Oncogenes Neu/HER2/ErbB2 and Ras can induce mammary tumorigenesis via upregulation of cyclin D1. One major regulatory mechanism in these oncogenic signaling pathways is phosphorylation of serines or threonines preceding proline (pSer/Thr-Pro). Interestingly, the pSer/Thr-Pro motifs in proteins exist in two completely distinct cis and trans conformations, whose conversion is catalyzed specifically by the essential prolyl isomerase Pin1. By isomerizing pSer/Thr-Pro bonds, Pin1 can regulate the conformation and function of certain phosphorylated proteins. We have previously shown that Pin1 is overexpressed in breast tumors and positively regulates cyclin D1 by transcriptional activation and posttranslational stabilization. Moreover, in Pin1 knockout mice, mammary epithelial cells fail to undergo massive proliferation during pregnancy, as is the case in cyclin D1 null mice. These results indicate that Pin1 is upregulated in breast cancer and may be involved in mammary tumors. However, the mechanism of Pin1 overexpression in cancer and its significance in cell transformation remain largely unknown. Here we demonstrate that PIN1 expression is mediated by the transcription factor E2F and enhanced by c-Neu and Ha-Ras via E2F. Furthermore, overexpression of Pin1 not only confers transforming properties on mammary epithelial cells but also enhances the transformed phenotypes of Neu/Ras-transformed mammary epithelial cells. In contrast, inhibition of Pin1 suppresses Neu- and Ras-induced transformed phenotypes, which can be fully rescued by overexpression of a constitutively active cyclin D1 mutant that is refractory to the Pin1 inhibition. Thus, Pin1 is an E2F target gene that is essential for the Neu/Ras-induced transformation of mammary epithelial cells through activation of cyclin D1.

scholarly journals H3K9 Promotes Under-Replication of Pericentromeric Heterochromatin in Drosophila Salivary Gland Polytene Chromosomes

Genes ◽  
2019 ◽  
Vol 10 (2) ◽  
pp. 93 ◽  
Author(s):  
Robin Armstrong ◽  
Taylor Penke ◽  
Samuel Chao ◽  
Gabrielle Gentile ◽  
Brian Strahl ◽  
...  
Keyword(s):  
Salivary Gland ◽  
Dna Replication ◽  
Polytene Chromosomes ◽  
Cell Types ◽  
Gene Replacement ◽  
Pericentric Heterochromatin ◽  
Replication Initiation ◽  
H3k9 Methylation ◽  
Diploid Cells ◽  
And Function

Chromatin structure and its organization contributes to the proper regulation and timing of DNA replication. Yet, the precise mechanism by which chromatin contributes to DNA replication remains incompletely understood. This is particularly true for cell types that rely on polyploidization as a developmental strategy for growth and high biosynthetic capacity. During Drosophila larval development, cells of the salivary gland undergo endoreplication, repetitive rounds of DNA synthesis without intervening cell division, resulting in ploidy values of ~1350C. S phase of these endocycles displays a reproducible pattern of early and late replicating regions of the genome resulting from the activity of the same replication initiation factors that are used in diploid cells. However, unlike diploid cells, the latest replicating regions of polyploid salivary gland genomes, composed primarily of pericentric heterochromatic enriched in H3K9 methylation, are not replicated each endocycle, resulting in under-replicated domains with reduced ploidy. Here, we employ a histone gene replacement strategy in Drosophila to demonstrate that mutation of a histone residue important for heterochromatin organization and function (H3K9) but not mutation of a histone residue important for euchromatin function (H4K16), disrupts proper endoreplication in Drosophila salivary gland polyploid genomes thereby leading to DNA copy gain in pericentric heterochromatin. These findings reveal that H3K9 is necessary for normal levels of under-replication of pericentric heterochromatin and suggest that under-replication at pericentric heterochromatin is mediated through H3K9 methylation.

scholarly journals Threonine-11, Phosphorylated by Rad3 and ATM In Vitro, Is Required for Activation of Fission Yeast Checkpoint Kinase Cds1

2001 ◽  
Vol 21 (10) ◽  
pp. 3398-3404 ◽  
Author(s):  
Katsunori Tanaka ◽  
Michael N. Boddy ◽  
Xiao-Bo Chen ◽  
Clare H. McGowan ◽  
Paul Russell
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Fission Yeast ◽  
Ataxia Telangiectasia ◽  
Null Mutant ◽  
Ataxia Telangiectasia Mutated ◽  
Tolerance Mechanisms ◽  
Checkpoint Kinase ◽  
And Function

ABSTRACT Fission yeast Cds1 is phosphorylated and activated when DNA replication is interrupted by nucleotide starvation or DNA damage. Cds1 enforces the S-M checkpoint that couples mitosis (M) to the completion of DNA synthesis (S). Cds1 also controls replicational stress tolerance mechanisms. Cds1 is regulated by a group of proteins that includes Rad3, a kinase related to human checkpoint kinase ATM (ataxia telangiectasia mutated). ATM phosphorylates serine or threonine followed by glutamine (SQ or TQ). Here we show that in vitro, Rad3 and ATM phosphorylate the N-terminal domain of Cds1 at the motif T11Q12. Substitution of threonine-11 with alanine (T11A) abolished Cds1 activation that occurs when DNA replication is inhibited by hydroxyurea (HU) treatment. Thecds1-T11A mutant was profoundly sensitive to HU, although not quite as sensitive as a cds1− null mutant. Cds1T11A was unable to enforce the S-M checkpoint. These results strongly suggest that Rad3-dependent phosphorylation of Cds1 at threonine-11 is required for Cds1 activation and function.

scholarly journals MyoD induced enhancer RNA interacts with hnRNPL to activate target gene transcription during myogenic differentiation

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Yu Zhao ◽  
Jiajian Zhou ◽  
Liangqiang He ◽  
Yuying Li ◽  
Jie Yuan ◽  
...  
Keyword(s):  
Target Gene ◽  
Myogenic Differentiation ◽  
Myoblast Differentiation ◽  
Enhancer Rna ◽  
Super Enhancer ◽  
And Function ◽  
Nuclear Ribonucleoprotein ◽  
Multiple Cells

AbstractEmerging evidence supports roles of enhancer RNAs (eRNAs) in regulating target gene. Here, we study eRNA regulation and function during skeletal myoblast differentiation. We provide a panoramic view of enhancer transcription and categorization of eRNAs. Master transcription factor MyoD is crucial in activating eRNA production. Super enhancer (se) generated seRNA-1 and -2 promote myogenic differentiation in vitro and in vivo. seRNA-1 regulates expression levels of two nearby genes, myoglobin (Mb) and apolipoprotein L6 (Apol6), by binding to heterogeneous nuclear ribonucleoprotein L (hnRNPL). A CAAA tract on seRNA-1 is essential in mediating seRNA-1/hnRNPL binding and function. Disruption of seRNA-1-hnRNPL interaction attenuates Pol II and H3K36me3 deposition at the Mb locus, in coincidence with the reduction of its transcription. Furthermore, analyses of hnRNPL binding transcriptome-wide reveal its association with eRNAs is a general phenomenon in multiple cells. Collectively, we propose that eRNA-hnRNPL interaction represents a mechanism contributing to target mRNA activation.

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