Role of Transcription Factor Kar4 in Regulating Downstream Events in the Saccharomyces cerevisiae Pheromone Response Pathway

ABSTRACT Yeast Kar4 is a putative transcription factor required for karyogamy (the fusion of haploid nuclei during mating) and possibly other functions. Previously known to be required only for the transcriptional induction of KAR3 and CIK1, microarray experiments identified many genes regulated by Kar4 in both mating and mitosis. Several gene clusters are positively or negatively regulated by mating pheromone in a Kar4-dependent manner. Chromatin immunoprecipitation and gel shift assays confirmed that Kar4 binds to regulatory DNA sequences upstream of KAR3. Together with one-hybrid experiments, these data support a model in which both Kar4 and Ste12 bind jointly to the KAR3 promoter. Analysis of the upstream regions of Kar4-induced genes identified a DNA sequence motif that may be a binding site for Kar4. Mutation within the motif upstream of KAR3 eliminated pheromone induction. Genes regulated by Kar4, on average, are delayed in their temporal expression and exhibit a more stringent dose response to pheromone. Furthermore, the induction of Kar4 by pheromone is necessary for the delayed temporal induction of KAR3 and PRM2, genes required for efficient nuclear fusion during mating. Accordingly, we propose that Kar4 plays a critical role in the choreography of the mating response.

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Borders of Cis-Regulatory DNA Sequences Preferentially Harbor the Divergent Transcription Factor Binding Motifs in the Human Genome

Frontiers in Genetics ◽

10.3389/fgene.2018.00571 ◽

2018 ◽

Vol 9 ◽

Author(s):

Jia-Hsin Huang ◽

Ryan Shun-Yuen Kwan ◽

Zing Tsung-Yeh Tsai ◽

Tzu-Chieh Lin ◽

Huai-Kuang Tsai

Keyword(s):

Transcription Factor ◽

Human Genome ◽

Dna Sequences ◽

Transcription Factor Binding ◽

Binding Motifs ◽

Factor Binding ◽

Regulatory Dna Sequences ◽

Transcription Factor Binding Motifs ◽

Divergent Transcription ◽

Regulatory Dna

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Down Regulation of SIRT2 Reduced ASS Induced NSCLC Apoptosis Through the Release of Autophagy Components via Exosomes

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2020.601953 ◽

2020 ◽

Vol 8 ◽

Author(s):

Lei Wang ◽

Pei Xu ◽

Xiao Xie ◽

Fengqing Hu ◽

Lianyong Jiang ◽

...

Keyword(s):

Transcription Factor ◽

Cell Apoptosis ◽

Critical Role ◽

Nsclc Cell ◽

Dependent Manner ◽

Cell Metastasis ◽

Rna Immunoprecipitation ◽

Transcription Factor Eb

Metastasis of cancer is the main cause of death in many types of cancer. Acute shear stress (ASS) is an important part of tumor micro-environment, it plays a crucial role in tumor invasion and spread. However, less is known about the role of ASS in tumorigenesis and metastasis of NSCLC. In this study, NSCLC cells were exposed to ASS (10 dyn/cm2) to explore the effect of ASS in regulation of autophagy and exosome mediated cell survival. Finally, the influence of SIRT2 on NSCLC cell metastasis was verified in vivo. Our data demonstrates that ASS promotes exosome and autophagy components releasing in a time dependent manner, inhibition of exosome release exacerbates ASS induced NSCLC cell apoptosis. Furthermore, we identified that this function was regulated by sirtuin 2 (SIRT2). And, RNA immunoprecipitation (RIP) assay suggested SIRT2 directly bound to the 3′UTR of transcription factor EB (TFEB) and facilitated its mRNA stability. TFEB is a key transcription factor involved in the regulation of many lysosome related genes and plays a critical role in the fusion of autophagosome and lysosome. Altogether, this data revealed that SIRT2 is a mechanical sensitive protein, and it regulates ASS induced cell apoptosis by modulating the release of exosomes and autophagy components, which provides a promising strategy for the treatment of NSCLCs.

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The borders of cis-regulatory DNA sequences harbor the divergent transcription factor binding motifs in the human genome

10.1101/383182 ◽

2018 ◽

Author(s):

Jia-Hsin Huang ◽

Ryan Shun-Yuen Kwan ◽

Zing Tsung-Yeh Tsai ◽

Huai-Kuang Tsai

Keyword(s):

Transcription Factor ◽

Dna Sequences ◽

Dynamic Change ◽

Evolutionary Divergence ◽

Evolutionary Time ◽

Regulatory Regions ◽

Binding Motifs ◽

Regulatory Dna Sequences ◽

Hypersensitive Sites ◽

Regulatory Dna

AbstractChanges in the cis-regulatory DNA sequences and transcription factor (TF) repertoires provide major sources that shape the gene regulatory evolution in eukaryotes. However, it is currently unclear how dynamic change of DNA sequences introduce various divergence level of TF binding motifs in the genome over evolutionary time. Here, we estimated the evolutionary divergence level of the TF binding motifs, and quantified their occurrences in the DNase I hypersensitive sites. Results from our in silico motif scan and empirical TF-ChIP (chromatin immunoprecipitation) demonstrate that the divergent motifs tend to be introduced at the borders of the cis-regulatory regions, that are likely accompanied with the expansion through evolutionary time. Accordingly, we propose that an expansion by incorporating divergent motifs within the cis-regulatory regions provides a rationale for the evolutionary divergence of regulatory circuits.

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Quantifying the regulatory role of individual transcription factors in Escherichia coli

10.1101/2021.01.04.425191 ◽

2021 ◽

Author(s):

Sunil Guharajan ◽

Shivani Chhabra ◽

Vinuselvi Parisutham ◽

Robert C Brewster

Keyword(s):

Transcription Factors ◽

Dna Sequences ◽

Transcription Initiation ◽

Target Genes ◽

Regulatory Role ◽

E Coli ◽

Regulatory Dna Sequences ◽

The Individual ◽

Regulatory Dna

Transcription factors (TFs) modulate gene expression by binding to regulatory DNA sequences surrounding target genes. To isolate the fundamental regulatory interactions of E. coli TFs, we measure regulation of TFs acting on synthetic target genes that are designed to isolate the individual TF regulatory effect. This data is interpreted through a thermodynamic model that decouples the role of TF copy number and TF binding affinity from the interactions of the TF on RNA polymerase through two distinct mechanisms: (de)stabilization of the polymerase and (de)acceleration of transcription initiation. We find the contribution of each mechanism towards the observed regulation depends on TF identity and binding location; for the set of TFs studied here, regulation immediately downstream of the promoter is not sensitive to TF identity, however these same TFs regulate through distinct mechanisms at an upstream binding site. Furthermore, depending on binding location, these two mechanisms of regulation can act coherently, to reinforce the observed regulatory role (activation or repression), or incoherently, where the TF regulates two distinct steps with opposing effect.

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The repression of nuclear factor I/CCAAT transcription factor (NFI/CTF) transactivating domain by oxidative stress is mediated by a critical cysteine (Cys-427)

Biochemical Journal ◽

10.1042/bj3480235 ◽

2000 ◽

Vol 348 (1) ◽

pp. 235-240 ◽

Cited By ~ 11

Author(s):

Yannick MOREL ◽

Robert BAROUKI

Keyword(s):

Oxidative Stress ◽

Transcription Factor ◽

Transcription Factors ◽

Critical Role ◽

Point Mutations ◽

Dependent Manner ◽

Nuclear Factor ◽

Factor I ◽

Nuclear Factor I

The activity of the nuclear factor I/CCAAT transcription factor (NFI/CTF) is negatively regulated by oxidative stress. The addition of relatively high (millimolar) H2O2 concentrations inactivates cellular NFI DNA-binding activity whereas lower concentrations can repress NFI/CTF transactivating function. We have investigated the mechanism of this regulation using Gal4 fusion proteins and transfection assays. We show that micromolar H2O2 concentrations repress the transactivating domain of NFI/CTF in a dose-dependent manner and are less or not active on other transcription factors' transactivating domains. Studies using deletions and point mutations pointed to the critical role of Cys-427. Indeed, when this cysteine is mutated into a serine, the repression by H2O2 is totally blunted. Mutation of other cysteine, serine and tyrosine residues within the transactivating domain had no clear effect on the repression by H2O2. Finally, treatment of cells with the thiol-alkylating reagent N-ethylmaleimide leads to a decrease in the transactivating function, which is dependent on Cys-427. This study shows that transactivating domains of transcription factors can constitute very sensitive targets of oxidative stress and highlights the critical role of these domains.

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The binding of NCAM to FGFR1 induces a specific cellular response mediated by receptor trafficking

The Journal of Cell Biology ◽

10.1083/jcb.200903030 ◽

2009 ◽

Vol 187 (7) ◽

pp. 1101-1116 ◽

Cited By ~ 78

Author(s):

Chiara Francavilla ◽

Paola Cattaneo ◽

Vladimir Berezin ◽

Elisabeth Bock ◽

Diletta Ami ◽

...

Keyword(s):

Cell Migration ◽

Cellular Response ◽

Critical Role ◽

Biological Significance ◽

Receptor Activation ◽

Dependent Manner ◽

Fgf Receptor ◽

Neural Cell Adhesion ◽

Sustained Activation

Neural cell adhesion molecule (NCAM) associates with fibroblast growth factor (FGF) receptor-1 (FGFR1). However, the biological significance of this interaction remains largely elusive. In this study, we show that NCAM induces a specific, FGFR1-mediated cellular response that is remarkably different from that elicited by FGF-2. In contrast to FGF-induced degradation of endocytic FGFR1, NCAM promotes the stabilization of the receptor, which is recycled to the cell surface in a Rab11- and Src-dependent manner. In turn, FGFR1 recycling is required for NCAM-induced sustained activation of various effectors. Furthermore, NCAM, but not FGF-2, promotes cell migration, and this response depends on FGFR1 recycling and sustained Src activation. Our results implicate NCAM as a nonconventional ligand for FGFR1 that exerts a peculiar control on the intracellular trafficking of the receptor, resulting in a specific cellular response. Besides introducing a further level of complexity in the regulation of FGFR1 function, our findings highlight the link of FGFR recycling with sustained signaling and cell migration and the critical role of these events in dictating the cellular response evoked by receptor activation.

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The P-element-induced silencing effect of KP transposons is dose dependent in Drosophila melanogaster

Genome ◽

10.1139/g11-023 ◽

2011 ◽

Vol 54 (9) ◽

pp. 752-762 ◽

Cited By ~ 7

Author(s):

Alireza Sameny ◽

John Locke

Keyword(s):

Drosophila Melanogaster ◽

Critical Role ◽

Mutagenic Effect ◽

P Element ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

P Elements ◽

Single Well ◽

Dose Dependent

Transposable elements are found in the genomes of all eukaryotes and play a critical role in altering gene expression and genome organization. In Drosophila melanogaster, transposable P elements are responsible for the phenomenon of hybrid dysgenesis. KP elements, a deletion-derivative of the complete P element, can suppress this mutagenic effect. KP elements can also silence the expression of certain other P-element-mediated transgenes in a process called P-element-dependent silencing (PDS), which is thought to involve the recruitment of heterochromatin proteins. To explore the mechanism of this silencing, we have mobilized KP elements to create a series of strains that contain single, well-defined KP insertions that show PDS. To understand the quantitative role of KP elements in PDS, these single inserts were combined in a series of crosses to obtain genotypes with zero, one, or two KP elements, from which we could examine the effect of KP gene dose. The extent of PDS in these genotypes was shown to be dose dependent in a logarithmic rather than linear fashion. A logarithmic dose dependency is consistent with the KP products interacting with heterochromatic proteins in a concentration-dependent manner such that two molecules are needed to induce gene silencing.

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Functional modulation of PTH1R activation and signalling by RAMP2

10.1101/2021.12.08.471790 ◽

2021 ◽

Author(s):

Katarina Nemec ◽

Hannes Schihada ◽

Gunnar Kleinau ◽

Ulrike Zabel ◽

Eugene O. Grushevskyi ◽

...

Keyword(s):

Homology Modelling ◽

Critical Role ◽

Ion Homeostasis ◽

Optical Biosensors ◽

Dependent Manner ◽

Activated State ◽

Class B ◽

Receptor Activity Modifying Proteins ◽

G Protein Coupled

Receptor-activity-modifying proteins (RAMPs) are ubiquitously expressed membrane proteins that associate with different G protein-coupled receptors (GPCRs) including the parathyroid hormone 1 receptor (PTH1R), a class B GPCR, and an important modulator of mineral ion homeostasis and bone metabolism. However, it is unknown whether and how RAMP proteins may affect PTH1R function. Using different optical biosensors to measure the activation of PTH1R and its downstream signalling, we describe here that RAMP2 acts as a specific allosteric modulator of PTH1R, shifting PTH1R to a unique pre-activated state that permits faster activation in a ligand-specific manner. Moreover, RAMP2 modulates PTH1R downstream signalling in an agonist-dependent manner, most notably increasing the PTH-mediated Gi3 signalling sensitivity. Additionally, RAMP2 increases both PTH- and PTHrP-triggered β-arrestin2 recruitment to PTH1R. Employing homology modelling we describe the putative structural molecular basis underlying our functional findings. These data uncover a critical role of RAMPs in the activation and signalling of a GPCR that may provide a new venue for highly specific modulation of GPCR function and advanced drug design.

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Investigating the role of noncoding regulatory DNA in plasmid development for Yarrowia lipolytica

10.22541/au.160691063.38058320/v1 ◽

2020 ◽

Author(s):

Carmen Lopez ◽

Mingfeng Cao ◽

Zhanyi Yao ◽

Zengyi Shao

Keyword(s):

Yarrowia Lipolytica ◽

Plasmid Stability ◽

Critical Role ◽

Fold Increase ◽

Microbial Cell Factories ◽

Cell Factories ◽

Expression Platform ◽

Regulatory Dna ◽

Selection Of

Production of industrially relevant compounds in microbial cell factories can employ either genomes or plasmids as an expression platform. Selection of plasmids as pathway carriers is advantageous for rapid demonstration but poses a challenge of stability. Yarrowia lipolytica has attracted great attention in the past decade for the biosynthesis of chemicals related to fatty acids at titers attractive to industry, and many genetic tools have been developed to explore its oleaginous potential. Our recent studies on the autonomously replicating sequences (ARSs) of nonconventional yeasts revealed that the ARSs from Y. lipolytica showcase a unique structure that includes a previously unannotated sequence (spacer) linking the origin of replication (ORI) and the centromeric (CEN) element and plays a critical role in modulating plasmid behavior. Maintaining a native 645-bp spacer yielded a 4.5-fold increase in gene expression and higher plasmid stability compared to a more universally employed minimized ARS. Testing the modularity of the ARS sub-elements indicated that plasmid stability exhibits a pronounced cargo dependency. Instability caused both plasmid loss and intramolecular rearrangements. Altogether, our work clarifies the appropriate application of various ARSs for the scientific community and sheds light on a previously unexplored DNA element as a potential target for engineering Y. lipolytica.

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