scholarly journals Interactions between nascent proteins translated by adjacent ribosomes drive homomer assembly

Science ◽  
2020 ◽  
Vol 371 (6524) ◽  
pp. 57-64
Author(s):  
Matilde Bertolini ◽  
Kai Fenzl ◽  
Ilia Kats ◽  
Florian Wruck ◽  
Frank Tippmann ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Activity ◽  
Direct Interaction ◽  
Coiled Coils ◽  
General Mechanism ◽  
Dimer Formation ◽  
Oligomer Formation ◽  
Nuclear Lamin ◽  
Nascent Chain ◽  
Major Domain

Accurate assembly of newly synthesized proteins into functional oligomers is crucial for cell activity. In this study, we investigated whether direct interaction of two nascent proteins, emerging from nearby ribosomes (co-co assembly), constitutes a general mechanism for oligomer formation. We used proteome-wide screening to detect nascent chain–connected ribosome pairs and identified hundreds of homomer subunits that co-co assemble in human cells. Interactions are mediated by five major domain classes, among which N-terminal coiled coils are the most prevalent. We were able to reconstitute co-co assembly of nuclear lamin in Escherichia coli, demonstrating that dimer formation is independent of dedicated assembly machineries. Co-co assembly may thus represent an efficient way to limit protein aggregation risks posed by diffusion-driven assembly routes and ensure isoform-specific homomer formation.

scholarly journals Suppression of ΔbipA Phenotypes in Escherichia coli by Abolishment of Pseudouridylation at Specific Sites on the 23S rRNA

2008 ◽  
Vol 190 (23) ◽  
pp. 7675-7683 ◽  
Author(s):  
Karthik Krishnan ◽  
Ann M. Flower
Keyword(s):  
Escherichia Coli ◽  
Target Genes ◽  
Regulatory Protein ◽  
Elongation Factor ◽  
Direct Interaction ◽  
Cold Sensitivity ◽  
23S Rrna ◽  
Pseudouridine Synthase ◽  
Suppressor Effect ◽  
Cold Sensitive

ABSTRACT The BipA protein of Escherichia coli has intriguing similarities to the elongation factor subfamily of GTPases, including EF-Tu, EF-G, and LepA. In addition, phenotypes of a bipA deletion mutant suggest that BipA is involved in regulation of a variety of pathways. These two points have led to speculation that BipA may be a novel regulatory protein that affects efficient translation of target genes through direct interaction with the ribosome. We isolated and characterized suppressors of the cold-sensitive growth phenotype exhibited by ΔbipA strains and identified insertion mutations in rluC. The rluC gene encodes a pseudouridine synthase responsible for pseudouridine modification of 23S rRNA at three sites, all located near the peptidyl transferase center. Deletion of rluC not only suppressed cold sensitivity but also alleviated the decrease in capsule synthesis exhibited by bipA mutants, suggesting that the phenotypic effects of BipA are manifested through an effect on the ribosome. The suppressor effect is specific to rluC, as deletion of other rlu genes did not relieve cold sensitivity, and further, more than a single pseudouridine residue is involved, as alteration of single residues did not produce suppressors. These results are consistent with a role for BipA in either the structure or the function of the ribosome and imply that wild-type ribosomes are dependent on BipA for efficient expression of target mRNAs and that the lack of pseudouridylation at these three sites renders the ribosomes BipA independent.

The Escherichia coli RNA processing and degradation machinery is compartmentalized within an organized cellular network

2014 ◽  
Vol 458 (1) ◽  
pp. 11-22 ◽  
Author(s):  
Aziz Taghbalout ◽  
Qingfen Yang ◽  
Véronique Arluison
Keyword(s):  
Escherichia Coli ◽  
Rna Processing ◽  
Cellular Network ◽  
High Order ◽  
Cellular Structures ◽  
General Mechanism ◽  
Protein Networks ◽  
Macromolecular Assembly ◽  
Bacterial Rna

We have shown that the multiprotein network of the bacterial RNA processing and degradation is organized within high-order cellular structures. Macromolecular assembly of protein networks could provide a general mechanism to streamline specific pathways within the seemingly non-compartmentalized prokaryotic cytoplasm.

scholarly journals MazG, a Nucleoside Triphosphate Pyrophosphohydrolase, Interacts with Era, an Essential GTPase in Escherichia coli

2002 ◽  
Vol 184 (19) ◽  
pp. 5323-5329 ◽  
Author(s):  
Junjie Zhang ◽  
Masayori Inouye
Keyword(s):  
Escherichia Coli ◽  
Direct Interaction ◽  
Kanamycin Resistance ◽  
Nucleoside Triphosphate ◽  
Cellular Functions ◽  
Genomic Libraries ◽  
E Coli ◽  
Yeast Two Hybrid System ◽  
Nucleoside Triphosphate Pyrophosphohydrolase

ABSTRACT Era is an essential GTPase in Escherichia coli, and Era has been implicated in a number of cellular functions. Homologues of Era have been identified in various bacteria and some eukaryotes. Using the era gene as bait in the yeast two-hybrid system to screen E. coli genomic libraries, we discovered that Era interacts with MazG, a protein of unknown function which is highly conserved among bacteria. The direct interaction between Era and MazG was also confirmed in vitro, being stronger in the presence of GDP than in the presence of GTPγS. MazG was characterized as a nucleoside triphosphate pyrophosphohydrolase which can hydrolyze all eight of the canonical ribo- and deoxynucleoside triphosphates to their respective monophosphates and PPi, with a preference for deoxynucleotides. A mazG deletion strain of E. coli was constructed by replacing the mazG gene with a kanamycin resistance gene. Unlike mutT, a gene for another conserved nucleotide triphosphate pyrophosphohydrolase that functions as a mutator gene, the mazG deletion did not result in a mutator phenotype in E. coli.

scholarly journals The Bacteriophage T4 Inhibitor and Coactivator AsiA Inhibits Escherichia coli RNA Polymerase More Rapidly in the Absence of σ70 Region 1.1: Evidence that Region 1.1 Stabilizes the Interaction between σ70 and Core

2006 ◽  
Vol 188 (4) ◽  
pp. 1279-1285 ◽  
Author(s):  
Deborah M. Hinton ◽  
Srilatha Vuthoori ◽  
Rebecca Mulamba
Keyword(s):  
Escherichia Coli ◽  
Dna Binding ◽  
Rna Polymerase ◽  
Dynamic Equilibrium ◽  
Bacteriophage T4 ◽  
Direct Interaction ◽  
Binding Properties ◽  
E Coli ◽  
Dna Binding Properties ◽  
Two Hybrid

ABSTRACT The N-terminal region (region 1.1) of σ70, the primary σ subunit of Escherichia coli RNA polymerase, is a negatively charged domain that affects the DNA binding properties of σ70 regions 2 and 4. Region 1.1 prevents the interaction of free σ70 with DNA and modulates the formation of stable (open) polymerase/promoter complexes at certain promoters. The bacteriophage T4 AsiA protein is an inhibitor of σ70-dependent transcription from promoters that require an interaction between σ70 region 4 and the −35 DNA element and is the coactivator of transcription at T4 MotA-dependent promoters. Like AsiA, the T4 activator MotA also interacts with σ70 region 4. We have investigated the effect of region 1.1 on AsiA inhibition and MotA/AsiA activation. We show that σ70 region 1.1 is not required for MotA/AsiA activation at the T4 middle promoter P uvsX . However, the rate of AsiA inhibition and of MotA/AsiA activation of polymerase is significantly increased when region 1.1 is missing. We also find that RNA polymerase reconstituted with σ70 that lacks region 1.1 is less stable than polymerase with full-length σ70. Our previous work has demonstrated that the AsiA-inhibited polymerase is formed when AsiA binds to region 4 of free σ70 and then the AsiA/σ70 complex binds to core. Our results suggest that in the absence of region 1.1, there is a shift in the dynamic equilibrium between polymerase holoenzyme and free σ70 plus core, yielding more free σ70 at any given time. Thus, the rate of AsiA inhibition and AsiA/MotA activation increases when RNA polymerase lacks region 1.1 because of the increased availability of free σ70. Previous work has argued both for and against a direct interaction between regions 1.1 and 4. Using an E. coli two-hybrid assay, we do not detect an interaction between these regions. This result supports the idea that the ability of region 1.1 to prevent DNA binding by free σ70 arises through an indirect effect.

scholarly journals Roles of AtpI and Two YidC-Type Proteins from Alkaliphilic Bacillus pseudofirmus OF4 in ATP Synthase Assembly and Nonfermentative Growth

2012 ◽  
Vol 195 (2) ◽  
pp. 220-230 ◽  
Author(s):  
Jun Liu ◽  
David B. Hicks ◽  
Terry A. Krulwich
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Atpase Activity ◽  
Atp Synthase ◽  
Membrane Association ◽  
Functional Specialization ◽  
Chromosomal Deletion ◽  
Content Type ◽  
Oligomer Formation ◽  
Alkaliphilic Bacillus

ABSTRACTAtpI, a membrane protein encoded by many bacterialatpoperons, is reported to be necessary forc-ring oligomer formation during assembly of some ATP synthase complexes. We investigated chaperone functions of AtpI and compared them to those of AtpZ, a protein encoded by a gene upstream ofatpIthat has a role in magnesium acquisition at near-neutral pH, and of SpoIIIJ and YqjG, two YidC/OxaI/Alb3 family proteins, in alkaliphilicBacillus pseudofirmusOF4. A strain with a chromosomal deletion ofatpIgrew nonfermentatively, and its purified ATP synthase had ac-ring of normal size, indicating that AtpI is not absolutely required for ATP synthase function. However, deletion ofatpI, but notatpZ, led to reduced stability of the ATP synthase rotor, reduced membrane association of the F1domain, reduced ATPase activity, and modestly reduced nonfermentative growth on malate at both pH 7.5 and 10.5. BothspoIIIJandyqjG, but notatpIoratpZ, complemented a YidC-depletedEscherichia colistrain. Consistent with such overlapping functions, single deletions ofspoIIIJoryqjGin the alkaliphile did not affect membrane ATP synthase levels or activities, but functional specialization was indicated by YqjG and SpoIIIJ showing respectively greater roles in malate growth at pH 7.5 and 10.5. Expression ofyqjGwas elevated at pH 7.5 relative to that at pH 10.5 and in ΔspoIIIJstrains, but it was lower than constitutivespoIIIJexpression. Deletion ofatpZcaused the largest increase among the mutants in magnesium concentrations needed for pH 7.5 growth. The basis for this phenotype is not yet resolved.

scholarly journals Transcriptional regulation of the fad regulon genes of Escherichia coli by ArcA

Microbiology ◽  
2006 ◽  
Vol 152 (8) ◽  
pp. 2207-2219 ◽  
Author(s):  
Byung-Kwan Cho ◽  
Eric M. Knight ◽  
Bernhard Ø. Palsson
Keyword(s):  
Escherichia Coli ◽  
Transcription Factor ◽  
Transcriptional Regulation ◽  
Expression Profiles ◽  
Direct Interaction ◽  
Anaerobic Growth ◽  
Binding Motif ◽  
Promoter Regions ◽  
Chip Analysis

ArcA is a global transcription factor required for optimal growth of Escherichia coli during anaerobic growth. In this study, the role of ArcA on the transcriptional regulatory subnetwork of the fad regulon was investigated. Gene expression profiles of deletion mutants (ΔarcA, ΔfadR and ΔarcA/ΔfadR) indicated that (i) ArcA is a major transcription factor for the transcriptional regulation of fatty acid metabolism in the absence of oxygen, and (ii) ArcA and FadR cooperatively regulate the fad regulon under anaerobic conditions. To determine the direct interaction between ArcA and the promoters of the fad regulon genes, chromatin immunoprecipitation (ChIP) analysis was performed. ChIP analysis suggested that ArcA directly binds to the promoter regions of the fad regulon genes in vivo. An ArcA-binding motif was identified from known binding sequences and predicted putative binding sites in the promoter regions of the fad regulon genes. These results indicate that ArcA directly represses the expression of fad regulon genes during anaerobic growth.

scholarly journals Phospholipase D Activity Regulates Integrin-mediated Cell Spreading and Migration by Inducing GTP-Rac Translocation to the Plasma Membrane

2008 ◽  
Vol 19 (7) ◽  
pp. 3111-3123 ◽  
Author(s):  
Young Chan Chae ◽  
Jung Hwan Kim ◽  
Kyung Lock Kim ◽  
Hyun Wook Kim ◽  
Hye Young Lee ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Phospholipase D ◽  
Molecular Mechanisms ◽  
Direct Interaction ◽  
Cell Spreading ◽  
Small Gtpase ◽  
General Mechanism ◽  
Downstream Target ◽  
P21 Activated Kinase ◽  
And Migration

Small GTPase Rac is a crucial regulator of actin cytoskeletal rearrangement, and it plays an important role in cell spreading, migration, mitogenesis, phagocytosis, superoxide generation, and axonal growth. It is generally accepted that Rac activity is regulated by the guanosine triphosphate (GTP)/guanosine diphosphate (GDP) cycle. But, it is suggested that in addition to Rac-GTP loading, membrane localization is required for the initiation of downstream effector signaling. However, the molecular mechanisms that control the targeting of GTP-Rac to the plasma membrane remain largely unknown. Here, we have uncovered a signaling pathway linking phospholipase D (PLD) to the localized functions of Rac1. We show that PLD product phosphatidic acid (PA) acts as a membrane anchor of Rac1. The C-terminal polybasic motif of Rac1 is responsible for direct interaction with PA, and Rac1 mutated in this region is incapable of translocating to the plasma membrane and of activating downstream target p21-activated kinase upon integrin activation. Finally, we show that PA induces dissociation of Rho-guanine nucleotide dissociation inhibitor from Rac1 and that PA-mediated Rac1 localization is important for integrin-mediated lamellipodia formation, cell spreading, and migration. These results provide a novel molecular mechanism for the GTP-Rac1 localization through the elevating PLD activity, and they suggest a general mechanism for diverse cellular functions that is required localized Rac activation.

Expression of a truncated guanylate cyclase (GC-C), a receptor for heat-stable enterotoxin of enterotoxigenic Escherichia coli, and its dimer formation in COS-7 cells

1993 ◽  
Vol 15 (4) ◽  
pp. 283-291 ◽  
Author(s):  
Toshiya Hirayama ◽  
Akihiro Wada ◽  
Yuji Hidaka ◽  
Jun-ichi Fujisawa ◽  
Yoshifumi Takeda ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Guanylate Cyclase ◽  
Enterotoxigenic Escherichia Coli ◽  
Dimer Formation ◽  
Heat Stable

scholarly journals RppH-dependent pyrophosphohydrolysis of mRNAs is regulated by direct interaction with DapF in Escherichia coli

2014 ◽  
Vol 42 (20) ◽  
pp. 12746-12757 ◽  
Author(s):  
Chang-Ro Lee ◽  
Miri Kim ◽  
Young-Ha Park ◽  
Yeon-Ran Kim ◽  
Yeong-Jae Seok
Keyword(s):  
Escherichia Coli ◽  
Direct Interaction
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