scholarly journals Binding and Transcriptional Regulation by 14-3-3 (Bmh) Proteins Requires Residues Outside of the Canonical Motif

2013 ◽  
Vol 13 (1) ◽  
pp. 21-30 ◽  
Author(s):  
Pabitra K. Parua ◽  
Elton T. Young
Keyword(s):  
Regulation Of Transcription ◽  
Interacting Proteins ◽  
Regulatory Domain ◽  
Interaction Sites ◽  
Important Interaction ◽  
Proposed Model ◽  
Evolutionarily Conserved ◽  
Regulatory Mutations ◽  
Canonical Motif ◽  
Transcriptional Phenotype

ABSTRACTEvolutionarily conserved 14-3-3 proteins have important functions as dimers in numerous cellular signaling processes, including regulation of transcription. Yeast 14-3-3 proteins, known as Bmh, inhibit a post-DNA binding step in transcription activation by Adr1, a glucose-regulated transcription factor, by binding to its regulatory domain, residues 226 to 240. The domain was originally defined by regulatory mutations,ADR1calleles that alter activator-dependent gene expression. Here, we report thatADR1calleles and other mutations in the regulatory domain impair Bmh binding and abolish Bmh-dependent regulation both directly and indirectly. The indirect effect is caused by mutations that inhibit phosphorylation of Ser230 and thus inhibit Bmh binding, which requires phosphorylated Ser230. However, several mutations inhibit Bmh binding without inhibiting phosphorylation and thus define residues that provide important interaction sites between Adr1 and Bmh. Our proposed model of the Adr1 regulatory domain bound to Bmh suggests that residues Ser238 and Tyr239 could provide cross-dimer contacts to stabilize the complex and that this might explain the failure of a dimerization-deficient Bmh mutant to bind Adr1 and to inhibit its activity. A bioinformatics analysis of Bmh-interacting proteins suggests that residues outside the canonical 14-3-3 motif might be a general property of Bmh target proteins and might help explain the ability of 14-3-3 to distinguish target and nontarget proteins. Bmh binding to the Adr1 regulatory domain, and its failure to bind when mutations are present, explains at a molecular level the transcriptional phenotype ofADR1cmutants.

scholarly journals Elucidating the regulatory mechanism of Swi1 prion in global transcription and stress responses

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Zhiqiang Du ◽  
Jeniece Regan ◽  
Elizabeth Bartom ◽  
Wei-Sheng Wu ◽  
Li Zhang ◽  
...  
Keyword(s):  
Stress Responses ◽  
Regulation Of Transcription ◽  
Interacting Proteins ◽  
Environmental Stress Response ◽  
Key Factors ◽  
Cellular Functions ◽  
Beneficial Effects ◽  
Genome Wide ◽  
Genes Encoding ◽  
A Subunit

AbstractTranscriptional regulators are prevalent among identified prions in Saccharomyces cerevisiae, however, it is unclear how prions affect genome-wide transcription. We show here that the prion ([SWI+]) and mutant (swi1∆) forms of Swi1, a subunit of the SWI/SNF chromatin-remodeling complex, confer dramatically distinct transcriptomic profiles. In [SWI+] cells, genes encoding for 34 transcription factors (TFs) and 24 Swi1-interacting proteins can undergo transcriptional modifications. Several TFs show enhanced aggregation in [SWI+] cells. Further analyses suggest that such alterations are key factors in specifying the transcriptomic signatures of [SWI+] cells. Interestingly, swi1∆ and [SWI+] impose distinct and oftentimes opposite effects on cellular functions. Translation-associated activities, in particular, are significantly reduced in swi1∆ cells. Although both swi1∆ and [SWI+] cells are similarly sensitive to thermal, osmotic and drought stresses, harmful, neutral or beneficial effects were observed for a panel of tested chemical stressors. Further analyses suggest that the environmental stress response (ESR) is mechanistically different between swi1∆ and [SWI+] cells—stress-inducible ESR (iESR) are repressed by [SWI+] but unchanged by swi1∆ while stress-repressible ESR (rESR) are induced by [SWI+] but repressed by swi1∆. Our work thus demonstrates primarily gain-of-function outcomes through transcriptomic modifications by [SWI+] and highlights a prion-mediated regulation of transcription and phenotypes in yeast.

scholarly journals Microsatellites as Agents of Adaptive Change: An RNA-seq based Comparative Study of Transcriptomes from Five Helianthus Species

2021 ◽  
Author(s):  
Chathurani Ranathunge ◽  
Sreepriya Pramod ◽  
Sébastien Renaut ◽  
Gregory Wheeler ◽  
Andy Perkins ◽  
...  
Keyword(s):  
Gene Expression ◽  
Differentially Expressed ◽  
Regulation Of Transcription ◽  
Expression Divergence ◽  
Rna Seq ◽  
Gene Expression Divergence ◽  
Coding Regions ◽  
Regulatory Mutations ◽  
Helianthus Species

Mutations that provide environment dependent selective advantages drive adaptive divergence among species. Many phenotypic differences among related species are more likely to result from gene expression divergence rather than from non-synonymous mutations. In this regard, cis-regulatory mutations play an important part in generating functionally significant variation. Some proposed mechanisms that explore the role of cis-regulatory mutations in gene expression divergence involve microsatellites. Microsatellites exhibit high mutation rates and are abundant in both coding and non-coding regions and could influence gene function and products. Here we tested the hypothesis that microsatellites contribute to gene expression divergence among species with 50 individuals from nine closely related Helianthus species using an RNA-seq approach. Differential expression analyses of the transcriptomes revealed that genes containing microsatellites in non-coding regions (UTRs and introns) are more likely to be differentially expressed among species when compared to genes with microsatellites in the coding regions and transcripts lacking microsatellites. We detected a greater proportion of shared microsatellites in 5’UTRs and coding regions compared to 3’UTRs and non-coding transcripts among Helianthus spp. Further, allele frequency differences measured by pairwise FST at single nucleotide polymorphisms (SNPs), indicate greater genetic divergence in transcripts containing microsatellites compared to those lacking microsatellites. A gene ontology (GO) analysis revealed that microsatellite-containing differentially expressed genes are significantly enriched for GO terms associated with regulation of transcription and transcription factor activity. Collectively, our study provides compelling evidence to support the role of microsatellites in gene expression divergence.

scholarly journals Regulation of Pseudomonas aeruginosa virulence by the ExsACDE Signaling Cascade

2014 ◽  
Vol 70 (a1) ◽  
pp. C1393-C1393
Author(s):  
Robert Bernhards ◽  
Xiao Yi ◽  
Manisha Shrestha ◽  
Florian Schubot
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Crystal Structures ◽  
Conformational Changes ◽  
Type Iii Secretion ◽  
Negative Regulator ◽  
Signaling Cascade ◽  
Regulation Of Transcription ◽  
Regulatory Domain ◽  
Type Iii ◽  
Dimerization Interface

Pseudomonas aeruginosa requires its type III secretion system (T3SS) to facilitate acute infections. T3SS-related gene expression is controlled by the AraC-type transcriptional activator ExsA. A signaling cascade involving ExsA and three additional proteins -ExsC, ExsD, and ExsE directly ties the up-regulation of transcription to the activation of the type III secretion apparatus. In order to characterize the molecular interactions underlying the signaling process the crystal structures of the unique T3SS chaperone ExsC in complex with its cognate effector ExsE, the structure of the negative regulator ExsD and that of the regulatory domain of ExsA have been determined. The ExsC-ExsE structure revealed two Arg-X-Val-X-Arg motifs in ExsE that form identical interactions along opposite sides of the ExsC dimer. The structure of ExsD not only provided insights into the interactions of how ExsD is sequestered by ExsC but also revealed surprising similarities between ExsD and DNA binding proteins. Based on these findings, a new model for the ExsC-ExsD complex is proposed to explain its distinctive 2:2 stoichiometry and why ExsC displays a weaker affinity for ExsD than for ExsE. Lastly, we have determined the structure of ExsA regulatory domain revealing the protein's dimerization interface. The position of the interface appears to postulate interesting conformational changes upon binding of ExsA to DNA. The crystal structures of ExsD and the ExsA domain also serve as road maps for determining the interface for the critical interactions between these two proteins at the bottom of this unique signaling cascade.

scholarly journals Phosphoacceptor Site S173 in the Regulatory Domain of Epstein-Barr Virus ZEBRA Protein Is Required for Lytic DNA Replication but Not for Activation of Viral Early Genes

2007 ◽  
Vol 81 (7) ◽  
pp. 3303-3316 ◽  
Author(s):  
Ayman El-Guindy ◽  
Lee Heston ◽  
Henri-Jacques Delecluse ◽  
George Miller
Keyword(s):  
Dna Binding ◽  
Viral Replication ◽  
Epstein Barr Virus ◽  
Lytic Replication ◽  
Lytic Cycle ◽  
Regulation Of Transcription ◽  
Regulatory Domain ◽  
Barr Virus ◽  
Early Genes ◽  
Epstein Barr

ABSTRACT The Epstein-Barr virus ZEBRA protein controls the viral lytic cycle. ZEBRA activates the transcription of viral genes required for replication. ZEBRA also binds to oriLyt and interacts with components of the viral replication machinery. The mechanism that differentiates the roles of ZEBRA in regulation of transcription and initiation of lytic replication is unknown. Here we show that S173, a residue in the regulatory domain, is obligatory for ZEBRA to function as an origin binding protein but is dispensable for its role as a transcriptional activator of early genes. Serine-to-alanine substitution of this residue, which prevents phosphorylation of S173, resulted in a threefold reduction in the DNA binding affinity of ZEBRA for oriLyt, as assessed by chromatin immunoprecipitation. An independent assay based on ZEBRA solubility demonstrated a marked defect in DNA binding by the Z(S173A) mutant. The phenotype of a phosphomimetic mutant, the Z(S173D) mutant, was similar to that of wild-type ZEBRA. Our findings suggest that phosphorylation of S173 promotes viral replication by enhancing ZEBRA's affinity for DNA. The results imply that stronger DNA binding is required for ZEBRA to activate replication than that required to activate transcription.

scholarly journals Cohesin Irr1/Scc3 is likely to influence transcription in Saccharomyces cerevisiae via interaction with Mediator complex.

2013 ◽  
Vol 60 (2) ◽  
Author(s):  
Agata Cena ◽  
Marek Skoneczny ◽  
Anna Chełstowska ◽  
Piotr Kowalec ◽  
Renata Natorff ◽  
...  
Keyword(s):  
Gene Expression ◽  
Saccharomyces Cerevisiae ◽  
Mediator Complex ◽  
Regulation Of Transcription ◽  
Transcriptional Coactivator ◽  
Diploid Strain ◽  
The Core ◽  
Chromatid Cohesion ◽  
Evolutionarily Conserved

The evolutionarily conserved proteins forming sister chromatid cohesion complex are also involved in the regulation of gene transcription. The participation of SA2p (mammalian ortholog of yeast Irr1p, associated with the core of the complex) in the regulation of transcription is already described. Here we analyzed microarray profiles of gene expression of a Saccharomyces cerevisiae irr1-1/IRR1 heterozygous diploid strain. We report that expression of 33 genes is affected by the presence of the mutated Irr1-1p and identify those genes. This supports the suggested role of Irr1p in the regulation of transcription. We also indicate that Irr1p may interact with elements of transcriptional coactivator Mediator.

‘PIPs’ in DNA polymerase: PCNA interaction affairs

2020 ◽  
Vol 48 (6) ◽  
pp. 2811-2822
Author(s):  
Narottam Acharya ◽  
Shraddheya Kumar Patel ◽  
Satya Ranjan Sahu ◽  
Premlata Kumari
Keyword(s):  
Dna Synthesis ◽  
Dna Polymerase ◽  
Dna Polymerases ◽  
Interacting Proteins ◽  
Short Sequence ◽  
Hydrophobic Pocket ◽  
Interaction Sites ◽  
Binding Partners ◽  
Eukaryotic Dna ◽  
Multiple Ligands

Interaction of PCNA with DNA polymerase is vital to efficient and processive DNA synthesis. PCNA being a homotrimeric ring possesses three hydrophobic pockets mostly involved in an interaction with its binding partners. PCNA interacting proteins contain a short sequence of eight amino acids, popularly coined as PIP motif, which snuggly fits into the hydrophobic pocket of PCNA to stabilize the interaction. In the last two decades, several PIP motifs have been mapped or predicted in eukaryotic DNA polymerases. In this review, we summarize our understandings of DNA polymerase-PCNA interaction, the function of such interaction during DNA synthesis, and emphasize the lacunae that persist. Because of the presence of multiple ligands in the replisome complex and due to many interaction sites in DNA polymerases, we also propose two modes of DNA polymerase positioning on PCNA required for DNA synthesis to rationalize the tool-belt model of DNA replication.

scholarly journals Custom Distinctions in the Interaction of G-protein β Subunits with N-type (CaV2.2) Versus P/Q-type (CaV2.1) Calcium Channels

2003 ◽  
Vol 121 (6) ◽  
pp. 495-510 ◽  
Author(s):  
Heather L. Agler ◽  
Jenafer Evans ◽  
Henry M. Colecraft ◽  
David T. Yue
Keyword(s):  
Calcium Channels ◽  
G Protein ◽  
Molecular Mechanisms ◽  
Single Point ◽  
Binding Pocket ◽  
Appreciable Effect ◽  
Voltage Range ◽  
Interaction Sites ◽  
Important Interaction ◽  
Binding Surface

Inhibition of N- (Cav2.2) and P/Q-type (Cav2.1) calcium channels by G-proteins contribute importantly to presynaptic inhibition as well as to the effects of opiates and cannabinoids. Accordingly, elucidating the molecular mechanisms underlying G-protein inhibition of voltage-gated calcium channels has been a major research focus. So far, inhibition is thought to result from the interaction of multiple proposed sites with the Gβγ complex (Gβγ). Far less is known about the important interaction sites on Gβγ itself. Here, we developed a novel electrophysiological paradigm, “compound-state willing-reluctant analysis,” to describe Gβγ interaction with N- and P/Q-type channels, and to provide a sensitive and efficient screen for changes in modulatory behavior over a broad range of potentials. The analysis confirmed that the apparent (un)binding kinetics of Gβγ with N-type are twofold slower than with P/Q-type at the voltage extremes, and emphasized that the kinetic discrepancy increases up to ten-fold in the mid-voltage range. To further investigate apparent differences in modulatory behavior, we screened both channels for the effects of single point alanine mutations within four regions of Gβ1, at residues known to interact with Gα. These residues might thereby be expected to interact with channel effectors. Of eight mutations studied, six affected G-protein modulation of both N- and P/Q-type channels to varying degrees, and one had no appreciable effect on either channel. The remaining mutation was remarkable for selective attenuation of effects on P/Q-, but not N-type channels. Surprisingly, this mutation decreased the (un)binding rates without affecting its overall affinity. The latter mutation suggests that the binding surface on Gβγ for N- and P/Q-type channels are different. Also, the manner in which this last mutation affected P/Q-type channels suggests that some residues may be important for “steering” or guiding the protein into the binding pocket, whereas others are important for simply binding to the channel.

scholarly journals The Ribosomal Protein L2 Interacts with the RNA Polymerase α Subunit and Acts as a Transcription Modulator in Escherichia coli

2010 ◽  
Vol 192 (7) ◽  
pp. 1882-1889 ◽  
Author(s):  
Valentina Rippa ◽  
Claudia Cirulli ◽  
Benedetta Di Palo ◽  
Nunzianna Doti ◽  
Angela Amoresano ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Ribosomal Protein ◽  
Transcription Initiation ◽  
Direct Interaction ◽  
Regulation Of Transcription ◽  
Interacting Proteins ◽  
Α Subunit ◽  
Gene Reporter Assay ◽  
Ribosomal Protein L2

ABSTRACT Identification of interacting proteins in stable complexes is essential to understand the mechanisms that regulate cellular processes at the molecular level. Transcription initiation in prokaryotes requires coordinated protein-protein and protein-DNA interactions that often involve one or more transcription factors in addition to RNA polymerase (RNAP) subunits. The RNAP α subunit (RNAPα) is a key regulatory element in gene transcription and functions through direct interaction with other proteins to control all stages of this process. A clear description of the RNAPα protein partners should greatly increase our understanding of transcription modulation. A functional proteomics approach was employed to investigate protein components that specifically interact with RNAPα. A tagged form of Escherichia coli RNAPα was used as bait to determine the molecular partners of this subunit in a whole-cell extract. Among other interacting proteins, 50S ribosomal protein L2 (RPL2) was clearly identified by mass spectrometry. The direct interaction between RNAPα and RPL2 was confirmed both in vivo and in vitro by performing coimmunoprecipitation and bacterial two-hybrid experiments. The functional role of this interaction was also investigated in the presence of a ribosomal promoter by using a β-galactosidase gene reporter assay. The results clearly demonstrated that RPL2 was able to increase β-galactosidase expression only in the presence of a specific ribosomal promoter, whereas it was inactive when it was assayed with an unrelated promoter. Interestingly, other ribosomal proteins (L1, L3, L20, and L27) did not have any effect on rRNA expression. The findings reported here strongly suggest that in addition to its role in ribosome assembly the highly conserved RPL2 protein plays a specific and direct role in regulation of transcription.

scholarly journals Critical Impact of Different Conserved Endoplasmic Retention Motifs and Dopamine Receptor Interacting Proteins (DRIPs) on Intracellular Localization and Trafficking of the D2 Dopamine Receptor (D2-R) Isoforms

Biomolecules ◽  
2020 ◽  
Vol 10 (10) ◽  
pp. 1355
Author(s):  
Kaja Blagotinšek Cokan ◽  
Maša Mavri ◽  
Catrin Sian Rutland ◽  
Sanja Glišić ◽  
Milan Senćanski ◽  
...  
Keyword(s):  
Dopamine Receptor ◽  
Intracellular Localization ◽  
Interacting Proteins ◽  
D2 Dopamine Receptor ◽  
Dopamine Receptor D2 ◽  
Evolutionarily Conserved ◽  
Interaction Partners ◽  
Receptor Interacting Proteins ◽  
G Protein Coupled

The type 2 dopamine receptor D2 (D2-R), member of the G protein-coupled receptor (GPCR) superfamily, exists in two isoforms, short (D2S-R) and long (D2L-R). They differ by an additional 29 amino acids (AA) in the third cytoplasmic loop (ICL3) of the D2L-R. These isoforms differ in their intracellular localization and trafficking functionality, as D2L-R possesses a larger intracellular pool, mostly in the endoplasmic reticulum (ER). This review focuses on the evolutionarily conserved motifs in the ICL3 of the D2-R and proteins interacting with the ICL3 of both isoforms, specifically with the 29 AA insert. These motifs might be involved in D2-R exit from the ER and have an impact on cell-surface and intracellular localization and, therefore, also play a role in the function of dopamine receptor signaling, ligand binding and possible homo/heterodimerization. Our recent bioinformatic data on potential new interaction partners for the ICL3 of D2-Rs are also presented. Both are highly relevant, and have clinical impacts on the pathophysiology of several diseases such as Parkinson’s disease, schizophrenia, Tourette’s syndrome, Huntington’s disease, manic depression, and others, as they are connected to a variety of essential motifs and differences in communication with interaction partners.

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