Mutant analysis in the nonlegume
            Parasponia andersonii
            identifies NIN and NF‐YA1 transcription factors as a core genetic network in nitrogen‐fixing nodule symbioses

The cryoprotection of cell activity is a key determinant in frozen-dough technology. Although several factors that contribute to freezing tolerance have been reported, the mechanism underlying the manner in which yeast cells respond to freezing and thawing (FT) stress is not well established. Therefore, the present study demonstrated the relationship between DaMDHAR encoding monodehydroascorbate reductase from Antarctic hairgrass Deschampsia antarctica and stress tolerance to repeated FT cycles (FT2) in transgenic yeast Saccharomyces cerevisiae. DaMDHAR-expressing yeast (DM) cells identified by immunoblotting analysis showed high tolerance to FT stress conditions, thereby causing lower damage for yeast cells than wild-type (WT) cells with empty vector alone. To detect FT2 tolerance-associated genes, 3′-quant RNA sequencing was employed using mRNA isolated from DM and WT cells exposed to FT (FT2) conditions. Approximately 332 genes showed ≥2-fold changes in DM cells and were classified into various groups according to their gene expression. The expressions of the changed genes were further confirmed using western blot analysis and biochemical assay. The upregulated expression of 197 genes was associated with pentose phosphate pathway, NADP metabolic process, metal ion homeostasis, sulfate assimilation, β-alanine metabolism, glycerol synthesis, and integral component of mitochondrial and plasma membrane (PM) in DM cells under FT2 stress, whereas the expression of the remaining 135 genes was partially related to protein processing, selenocompound metabolism, cell cycle arrest, oxidative phosphorylation, and α-glucoside transport under the same condition. With regard to transcription factors in DM cells, MSN4 and CIN5 were activated, but MSN2 and MGA1 were not. Regarding antioxidant systems and protein kinases in DM cells under FT stress, CTT1, GTO, GEX1, and YOL024W were upregulated, whereas AIF1, COX2, and TRX3 were not. Gene activation represented by transcription factors and enzymatic antioxidants appears to be associated with FT2-stress tolerance in transgenic yeast cells. RCK1, MET14, and SIP18, but not YPK2, have been known to be involved in the protein kinase-mediated signalling pathway and glycogen synthesis. Moreover, SPI18 and HSP12 encoding hydrophilin in the PM were detected. Therefore, it was concluded that the genetic network via the change of gene expression levels of multiple genes contributing to the stabilization and functionality of the mitochondria and PM, not of a single gene, might be the crucial determinant for FT tolerance in DaMDAHR-expressing transgenic yeast. These findings provide a foundation for elucidating the DaMDHAR-dependent molecular mechanism of the complex functional resistance in the cellular response to FT stress.

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Chromatin reader Dido3 regulates the genetic network of B cell differentiation

10.1101/2021.02.23.432411 ◽

2021 ◽

Author(s):

Fernando Gutiérrez del Burgo ◽

Tirso Pons ◽

Enrique Vázquez de Luis ◽

Carlos Martínez-A ◽

Ricardo Villares

Keyword(s):

Transcription Factors ◽

Cell Differentiation ◽

B Cell ◽

Cell Lineage ◽

Genetic Network ◽

B Cell Differentiation ◽

Differentiation Markers ◽

Hematopoietic Stem ◽

Severe Deficiency ◽

The Impact

ABSTRACTThe development of hematopoietic lineages is based on a complex balance of transcription factors whose expression depends on the epigenetic signatures that characterize each differentiation step. The B cell lineage arises from hematopoietic stem cells through the stepwise silencing of stemness genes and balanced expression of mutually regulated transcription factors, as well as DNA rearrangement. Here we report the impact on B cell differentiation of the lack of DIDO3, a reader of chromatin status, in the mouse hematopoietic compartment. We found reduced DNA accessibility in hematopoietic precursors, leading to severe deficiency in the generation of successive B cell differentiation stages. The expression of essential transcription factors and differentiation markers is affected, as is the somatic recombination process.One Sentence Summary: Epigenetic control of early hematopoiesis

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The role of homedomain-containing transcription factors in the control of organogenesis of nitrogen fixing nodules

Abstract book of the 2nd International Scientific Conference "Plants and Microbes: the Future of Biotechnology" PLAMIC2020 ◽

10.28983/plamic2020.062 ◽

2020 ◽

Author(s):

A. V. Dolgikh ◽

E. A. Dolgikh

Keyword(s):

Transcription Factors ◽

Nitrogen Fixing

Interconnection between key transcription factors of symbiosis regulation and the homeodomain-containing transcription factors KNOXs and BELLs in the development of nodules in pea was investigated.

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Morphogenesis at criticality

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1324186111 ◽

2014 ◽

Vol 111 (10) ◽

pp. 3683-3688 ◽

Cited By ~ 63

Author(s):

Dmitry Krotov ◽

Julien O. Dubuis ◽

Thomas Gregor ◽

William Bialek

Keyword(s):

Transcription Factors ◽

Spatial Patterns ◽

Gaussian Distribution ◽

Gene Network ◽

Fruit Fly ◽

Genetic Network ◽

Strong Correlations ◽

Expression Levels ◽

Gap Gene ◽

Gap Genes

Spatial patterns in the early fruit fly embryo emerge from a network of interactions among transcription factors, the gap genes, driven by maternal inputs. Such networks can exhibit many qualitatively different behaviors, separated by critical surfaces. At criticality, we should observe strong correlations in the fluctuations of different genes around their mean expression levels, a slowing of the dynamics along some but not all directions in the space of possible expression levels, correlations of expression fluctuations over long distances in the embryo, and departures from a Gaussian distribution of these fluctuations. Analysis of recent experiments on the gap gene network shows that all these signatures are observed, and that the different signatures are related in ways predicted by theory. Although there might be other explanations for these individual phenomena, the confluence of evidence suggests that this genetic network is tuned to criticality.

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Epigenetic Modifying Drugs Inhibit MDS/AML Cell Growth through Selective Disruption of the Interactions Between Lineage-Determining Transcription Factors and DNA/Histone Modifiers

Blood ◽

10.1182/blood.v126.23.3853.3853 ◽

2015 ◽

Vol 126 (23) ◽

pp. 3853-3853

Author(s):

Yuan Li ◽

Ming Yue ◽

John Anastasi ◽

Jeffery Q Shen ◽

Yali Dou ◽

...

Keyword(s):

Transcription Factors ◽

Genetic Network ◽

Drug Action ◽

Leukemia Cells ◽

Monocytic Leukemia ◽

Lineage Differentiation ◽

Epigenetic Modifiers ◽

Genome Wide ◽

Histone Modifiers ◽

Distinct Lineage

Abstract Normal hematopoiesis is controlled by a well-connected genetic network composed of several transcription factors (TFs) including PU.1 and GATA1. It has been postulated that both transcription factors and epigenetic modifiers work collaboratively to regulate hematopoietic stem cell differentiation and lineage specification as well as leukemogenesis. However, it is unclear about how the interplay between genetic network and epigenetic regulatory modifiers regulates locus-specific chromatin modifications and gene expression in normal hematopoiesis and hematologic malignancies such as myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Drugs targeting epigenetic modifiers including DNA methyltransferases (DNMTs), histone methyltransferases (HMTs) and histone deacetylases (HDACs) have been shown to be effective in a small portion of patients with MDS/AML, but the mechanisms underlying the efficacy and selectivity of different epigenetic modifying drugs are unknown. In this study, we performed growth-inhibition experiments with several epigenetic modifying drugs in multiple AML cell lines and identified two distinct lineage/differentiation-associated growth-inhibition patterns. Monocytic leukemia cells, but not erythroid leukemia cells, were sensitive to H3K4 HMT inhibitors, whereas both erythroid and monocytic leukemia cells were hypersensitive to DNMT and H3K27 HMT inhibitors. Importantly, co-immunoprecipitation experiments demonstrated lineage-specific interactions between the lineage-determining TFs (PU.1/SPI1 and GATA1) and the DNA/histone modifiers (DNMT1, DNMT3A/3B, TET2 and EZH2). Specifically, SPI1/PU.1 interacts with DNMT1 and EZH2, while GATA1 interacts with TET2 and DNMT3A/3B in MDS-derived erythroid leukaemia. In monocytic leukemia, SPI1/PU.1 interacts with TET2. Epigenetic modifying drugs such as azacytidine and 3-deazaneplanocin efficiently disrupted the interactions between the lineage-determining TFs and the DNA/histone modifiers without changing the expression of these proteins. We developed a new method, crosslink-assisted DNA modification immunoprecipitation assay (CDMIA), to simultaneously measure 5-methylcytosine (5-mC) and hydroxymethylcytosine (5-hmC). CDMIAs revealed significant drug-responsive changes in 5-mC/5-hmC at the promoters of differentiation/lineage-controlling genes such as PU.1/SPI1, but not at the global 5-mC/5-hmC. Sequential-ChIP and chromatin conformation capture (3C) showed that PU.1/SPI1 recruited polymerase II (pol-II) and the DNA/histone modifying complexes to PU.1/SPI1 toform distinct chromatin structures in a lineage-specific manner. We have selected azacytidine-resistant clones and established azacytidine-resistant cell lines from the previously azacytine-sensitive erythroid and monocytic leukemia cells. Strikingly, azacytine at the same concentrations failed to disrupt the interactions between the lineage-determining transcription factors and the DNA/histone modifiers in these drug-resistant leukemia cells. Genome-wide sequencing revealed novel mutations in TET2, TET3, DNMT3L and PU.1/SP1 that were confirmed by Sanger sequencing. These mutations correlated with the altered interactions between PU.1/SPI1 and the DNA/histone modifying complexes and predicted the responses to epigenetic modifying drugs. Examination of clinical specimens from patients with MDS/AML confirmed the presence of distinct lineage/differentiation-specific chromatin structure with a high-level recruitment of DNA/histone modifiers. Our genome-wide epigenetic analysis demonstrates the statistically significant enrichment of the SPI1/PU.1, TP53 and MYB DNA-binding motifs in hyper-H3K27 trimethylated DNA sequences in erythroid-predominant MDS. These results demonstrate the presence of locus-specific, drug-sensitive chromatin structures in MDS/AML subtypes. Our data revealed a novel epigenetic modifying drug action model that involves selective disruption of the disease-specific interactions between the lineage-determining factors and DNA/histone modifiers. Such drug action models may provide new insights into the mechanisms underlying the efficacy and selectivity of epigenetic modifying drugs. Disclosures Larson: Novartis: Consultancy, Research Funding; Pfizer: Consultancy; Ariad: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy.

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Stochastic dynamics of genetic broadcasting networks

10.1101/074401 ◽

2016 ◽

Author(s):

Davit A. Potoyan ◽

Peter G. Wolynes

Keyword(s):

Transcription Factors ◽

Time Scale ◽

Stochastic Dynamics ◽

Genetic Network ◽

Large Network ◽

Residence Times ◽

Release Times ◽

Genetic Switches ◽

Number Of Binding Sites ◽

Genomic Locations

AbstractThe complex genetic programs of eukaryotic cells are often regulated by key transcription factors occupying or clearing out of a large number of genomic locations. Orchestrating the residence times of these factors is therefore important for the well organized functioning of a large network. The classic models of genetic switches sidestep this timing issue by assuming the binding of transcription factors to be governed entirely by thermodynamic protein-DNA affinities. Here we show that relying on passive thermodynamics and random release times can lead to a “time-scale crisis” of master genes that broadcast their signals to large number of binding sites. We demonstrate that this “time-scale crisis” can be resolved by actively regulating residence times through molecular stripping. We illustrate these ideas by studying the stochastic dynamics of the genetic network of the central eukaryotic master regulator NFκB which broadcasts its signals to many downstream genes that regulate immune response, apoptosis etc.

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Chromatin Reader Dido3 Regulates the Genetic Network of B Cell Differentiation

10.21203/rs.3.rs-1031396/v1 ◽

2021 ◽

Author(s):

Fernando Gutiérrez del Burgo ◽

Tirso Pons ◽

Enrique Vázquez de Luis ◽

Carlos Martínez-A ◽

Ricardo Villares

Keyword(s):

Transcription Factors ◽

Cell Differentiation ◽

B Cell ◽

Specific Activity ◽

Cell Lineage ◽

Genetic Network ◽

B Cell Differentiation ◽

Differentiation Markers ◽

Hematopoietic Stem ◽

The Impact

Abstract BACKGROUNDThe development of hematopoietic lineages is based on a complex balance of transcription factors whose expression depends on the epigenetic signatures that characterize each differentiation step. The B cell lineage arises from hematopoietic stem cells through the stepwise silencing of stemness genes and balanced expression of mutually regulated transcription factors, as well as DNA rearrangement, in a complex process involving epigenetic remodeling.RESULTSHere we report the impact on B cell differentiation of the lack of DIDO3, a reader of histone post-translational modifications, in the mouse hematopoietic compartment. We found reduced DNA accessibility in hematopoietic precursors, leading to a severe deficiency specifically in the generation of successive stages of B-cell differentiation. The expression of essential transcription factors and differentiation markers is impaired, as is the process of somatic recombination. DIDO3-deficient cells show transcriptional alterations of a number of polycomb repressive complex 2, suggesting the involvement of DIDO3 in determining the specific activity of PRC2 in the B cell lineage, including VH-DJH rearrangement. CONCLUSIONSTaken together, our data suggest that DIDO3 is an epigenetic reader involved in the specific differentiation of B cell precursors in the hematopoietic compartment of mice.

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Dmrt5 Controls Corticotrope and Gonadotrope Differentiation in the Zebrafish Pituitary

Molecular Endocrinology ◽

10.1210/me.2014-1176 ◽

2015 ◽

Vol 29 (2) ◽

pp. 187-199 ◽

Cited By ~ 7

Author(s):

Martin Graf ◽

Elizabeth-Raye Teo Qi-Wen ◽

Menachem Viktor Sarusie ◽

Flora Rajaei ◽

Christoph Winkler

Keyword(s):

Transcription Factors ◽

Cell Differentiation ◽

Reproductive System ◽

Anterior Pituitary ◽

Endocrine Cells ◽

Terminal Differentiation ◽

Pars Distalis ◽

Vertebrate Species ◽

Lineage Specification ◽

Mutant Analysis

Abstract Dmrt transcription factors control sex determination or sex-specific differentiation across all invertebrate and vertebrate species, in which they have been studied so far. In addition to important functions in the reproductive system, also nongonadal roles have been assigned to several dmrt family members. One example is dmrt5, which was shown to guide neurogenesis in the forebrain of some vertebrates including fish. Here we show that in zebrafish, dmrt5 is also expressed adjacent to the pituitary anlage and later in the anterior pars distalis in which it organizes differentiation of endocrine cells. We find that pituitary induction, cell survival, proliferation, and early lineage specification in the pituitary is independent of dmrt5. Instead, dmrt5 is required for terminal differentiation of corticotropes and gonadotropes. Gene knockdown and mutant analysis revealed that dmrt5 promotes corticotrope differentiation via tbx19 expression, whereas it prevents gonadotrope differentiation in the anterior pars distalis. In dmrt5 morphants and mutants, reduced corticotrope numbers may result in irregular positioning and reduced maintenance of lactotropes. In conclusion, our study establishes a novel function for dmrt5 for cell differentiation in the anterior pituitary. Intriguingly, its effect on gonadotrope numbers defines a first nongonadal role for a dmrt family member that appears crucial for the activity of the reproductive system.

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