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Biomedical subjects

Evgeny Nudler

Publications and source records attributed to Evgeny Nudler.

At least 19 recordsLinked to original sources
PubMed2006

Gene control by large noncoding RNAs.

Large noncoding RNAs (lncRNAs) have emerged as key players in regulating various fundamental cellular processes. Recent reports identify a functional lncRNA, Evf-2, that operates during development to control the expression of specific homeodomain proteins, and they provide important insights into the mechanism of cooperation between a newly discovered nuclear receptor co-repressor protein (SLIRP) and steroid receptor activator RNA. Evf-2 is the first example of lncRNA directly involved in organogenesis in vertebrates.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Extensive functional overlap between sigma factors in Escherichia coli.

Bacterial core RNA polymerase (RNAP) must associate with a sigma factor to recognize promoter sequences. Escherichia coli encodes seven sigma factors, each believed to be specific for a largely distinct subset of promoters. Using microarrays representing the entire E. coli genome, we identify 87 in vivo targets of sigma32, the heat-shock sigma factor, and estimate that there are 120-150 sigma32 promoters in total. Unexpectedly, 25% of these sigma32 targets are located within coding regions, suggesting novel regulatory roles for sigma32. The majority of sigma32 promoter targets overlap with those of sigma70, the housekeeping sigma factor. Furthermore, their DNA sequence motifs are often interdigitated, with RNAPsigma70 and RNAPsigma32 initiating transcription in vitro with similar efficiency and from identical positions. SigmaE-regulated promoters also overlap extensively with those for sigma70. These results suggest that extensive functional overlap between sigma factors is an important phenomenon.

Binding Sites↗
PubMed2006

Flipping riboswitches.

Riboswitches are common cis-acting regulatory elements in bacteria. They are made of nascent RNA that changes its conformation in response to direct binding of cognate metabolites. The publication of five high-resolution crystal structures provides a comprehensive view of how riboswitches sense their ligands and points to new challenges in this emerging field.

Bacteria↗
PubMed2006

Transcription regulatory elements are punctuation marks for DNA replication.

Collisions between DNA replication and transcription significantly affect genome organization, regulation, and stability. Previous studies have described collisions between replication forks and elongating RNA polymerases. Although replication collisions with the transcription-initiation or -termination complexes are potentially even more important because most genes are not actively transcribed during DNA replication, their existence and mechanisms remained unproven. To address this matter, we have designed a bacterial promoter that binds RNA polymerase and maintains it in the initiating mode by precluding the transition into the elongation mode. By using electrophoretic analysis of replication intermediates, we have found that this steadfast transcription-initiation complex inhibits replication fork progression in an orientation-dependent manner during head-on collisions. Transcription terminators also appeared to attenuate DNA replication, but in the opposite, codirectional orientation. Thus, transcription regulatory signals may serve as "punctuation marks" for DNA replication in vivo.

AT Rich Sequence↗
PubMed2006

RNA-mediated response to heat shock in mammalian cells.

The heat-shock transcription factor 1 (HSF1) has an important role in the heat-shock response in vertebrates by inducing the expression of heat-shock proteins (HSPs) and other cytoprotective proteins. HSF1 is present in unstressed cells in an inactive monomeric form and becomes activated by heat and other stress stimuli. HSF1 activation involves trimerization and acquisition of a site-specific DNA-binding activity, which is negatively regulated by interaction with certain HSPs. Here we show that HSF1 activation by heat shock is an active process that is mediated by a ribonucleoprotein complex containing translation elongation factor eEF1A and a previously unknown non-coding RNA that we term HSR1 (heat shock RNA-1). HSR1 is constitutively expressed in human and rodent cells and its homologues are functionally interchangeable. Both HSR1 and eEF1A are required for HSF1 activation in vitro; antisense oligonucleotides or short interfering (si)RNA against HSR1 impair the heat-shock response in vivo, rendering cells thermosensitive. The central role of HSR1 during heat shock implies that targeting this RNA could serve as a new therapeutic model for cancer, inflammation and other conditions associated with HSF1 deregulation.

Animals↗
PubMed2006

Thermodynamic and kinetic modeling of transcriptional pausing.

We present a statistical mechanics approach for the prediction of backtracked pauses in bacterial transcription elongation derived from structural models of the transcription elongation complex (EC). Our algorithm is based on the thermodynamic stability of the EC along the DNA template calculated from the sequence-dependent free energy of DNA-DNA, DNA-RNA, and RNA-RNA base pairing associated with (i) the translocational and size fluctuations of the transcription bubble; (ii) changes in the associated DNA-RNA hybrid; and (iii) changes in the cotranscriptional RNA secondary structure upstream of the RNA exit channel. The calculations involve no adjustable parameters except for a cutoff used to discriminate paused from nonpaused complexes. When applied to 100 experimental pauses in transcription elongation by Escherichia coli RNA polymerase on 10 DNA templates, the approach produces statistically significant results. We also present a kinetic model for the rate of recovery of backtracked paused complexes. A crucial ingredient of our model is the incorporation of kinetic barriers to backtracking resulting from steric clashes of EC with the cotranscriptionally generated RNA secondary structure, an aspect not included explicitly in previous attempts at modeling the transcription elongation process.

Computational Biology↗
PubMed2005

NO-mediated cytoprotection: instant adaptation to oxidative stress in bacteria.

Numerous sophisticated systems have been described that protect bacteria from increased levels of reactive oxygen species. Although indispensable during prolonged oxidative stress, these response systems depend on newly synthesized proteins, and are hence both time and energy consuming. Here, we describe an "express" cytoprotective system in Bacillus subtilis which depends on nitric oxide (NO). We show that NO immediately protects bacterial cells from reactive oxygen species by two independent mechanisms. NO transiently suppresses the enzymatic reduction of free cysteine that fuels the damaging Fenton reaction. In addition, NO directly reactivates catalase, a major antioxidant enzyme that has been inhibited in vivo by endogenous cysteine. Our data also reveal a critical role for bacterial NO-synthase in adaptation to oxidative stress associated with fast metabolic changes, and suggest a possible role for NO in defending pathogens against immune oxidative attack.

Adaptation, Physiological↗
PubMed2005

A ratchet mechanism of transcription elongation and its control.

RNA chain elongation is a highly processive and accurate process that is finely regulated by numerous intrinsic and extrinsic signals. Here we describe a general mechanism that governs RNA polymerase (RNAP) movement and response to regulatory inputs such as pauses, terminators, and elongation factors. We show that E.coli RNAP moves by a complex Brownian ratchet mechanism, which acts prior to phosphodiester bond formation. The incoming substrate and the flexible F bridge domain of the catalytic center serve as two separate ratchet devices that function in concert to drive forward translocation. The adjacent G loop domain controls F bridge motion, thus keeping the proper balance between productive and inactive states of the elongation complex. This balance is critical for cell viability since it determines the rate, processivity, and fidelity of transcription.

Cross-Linking Reagents↗
PubMed2004

Control of plasma nitric oxide bioactivity by perfluorocarbons: physiological mechanisms and clinical implications.

BACKGROUND: Perfluorocarbons (PFCs) are promising blood substitutes because of their chemical inertness and unparalleled ability to transport and upload O2 and CO2. Here, we report that PFC emulsions also efficiently absorb and transport nitric oxide (NO). METHODS AND RESULTS: Accumulation of NO and O2 in PFC micelles results in rapid NO oxidation and generation of reactive NO(x) species. Such micellar catalysis of NO oxidation leads to formation of vasoactive S-nitrosothiols (RSNO) in vitro and in vivo as detected electrochemically. The efficiency of PFC-mediated S-nitrosation depends on the amount of PFC in aqueous solution. The optimal PFC concentration that produced the maximum level of RSNO was approximately 1% (vol/vol). Larger PFC amounts were progressively less efficient in generating RSNO and functioned simply as NO sink. These results explain the characteristic hemodynamic effects of PFCs. Intravenous bolus application of PFC (0.14 g/kg, approximately 1% vol/vol) to Wistar-Kyoto rats decreased mean arterial pressure significantly (-10 mm Hg over 40 minutes). PFC-induced hypotension could be further stimulated (-17 mm Hg over 140 minutes) by exogenous thiols (cysteine and glutathione). In contrast, a larger amount of PFC (1 g/kg, approximately 7% vol/vol) exhibited a strong hypertensive effect (11 mm Hg over 40 minutes). CONCLUSIONS: The present study reveals a physiologically significant pool of endogenous plasma NO and underscores the crucial role of the circulating hydrophobic phase in modulating its bioactivity. The results also establish PFC as a conceptually new pharmacological tool for various cardiovascular complications associated with NO imbalance.

Animals↗
PubMed2004

The riboswitch control of bacterial metabolism.

Aptamers are artificial nucleic acids that selectively bind small molecules. In the past two years, it has become clear that nature has already devised its own aptamers that play important regulatory roles. RNA sensors have been discovered in both Gram-positive and Gram-negative bacteria that function as molecular switches in response to direct binding of structurally diverse metabolites. These natural RNA aptamers, called 'riboswitches', are imbedded in the leader sequences of numerous metabolic genes. Riboswitches are able to repress or activate their cognate genes at both transcriptional and translational levels. Here, we summarize the recent progress in the identification and characterization of riboswitches and discuss their evolution and distribution.

Bacteria↗
PubMed2004

STR2: a structure to string approach for locating G-box riboswitch shapes in pre-selected genes.

Traditional sequence-based search methods such as BLAST and FASTA can be used to identify sequence similarities. Recently, there is a growing interest in performing RNA shape similarity searches inside selected genes to locate RNA structure motifs that are known to possess functionally important roles. For example, in the newly discovered RNA genetic control elements called "riboswitches", the box domain is known to be highly conserved among various bacterial species in both its nucleotide composition and shape. However, in non-bacterial species, shape conservation is likely to become more important than sequence conservation when searching for riboswitch patterns. For this purpose, we present an approach tailored for detecting RNA shape similarities. We extend the Structure to String (ST R2) method that was initially proposed to locate shape similarities in proteins to identify predicted secondary structures of RNAs. The ST R2 for RNAs is a translation of a secondary structure to a string of characters, after which known sequence-based search algorithms with an efficient implementation are being used. We validate that the ST R2 succeeds to locate G-box riboswitches in prokaryotes, as expected. Subsequently we show running examples when attempting to detect G-box riboswitch candidates in eukaryotes.

Algorithms↗
PubMed2003

Transcription through the roadblocks: the role of RNA polymerase cooperation.

During transcription, cellular RNA polymerases (RNAP) have to deal with numerous potential roadblocks imposed by various DNA binding proteins. Many such proteins partially or completely interrupt a single round of RNA chain elongation in vitro. Here we demonstrate that Escherichia coli RNAP can effectively read through the site-specific DNA-binding proteins in vitro and in vivo if more than one RNAP molecule is allowed to initiate from the same promoter. The anti-roadblock activity of the trailing RNAP does not require transcript cleavage activity but relies on forward translocation of roadblocked complexes. These results support a cooperation model of transcription whereby RNAP molecules behave as 'partners' helping one another to traverse intrinsic and extrinsic obstacles.

Base Sequence↗
PubMed2003

Cooperation between RNA polymerase molecules in transcription elongation.

Transcription elongation is responsible for rapid synthesis of RNA chains of thousands of nucleotides in vivo. In contrast, a single round of transcription performed in vitro is frequently interrupted by pauses and arrests that drastically reduce the elongation rate and the yield of the full-length transcript. Here we demonstrate that most transcriptional delays disappear if more than one RNA polymerase (RNAP) molecule initiates from the same promoter. Anti-arrest and anti-pause effects of trailing RNAP are due to forward translocation of leading (backtracked) complexes. Such cooperation between RNAP molecules links the rate of elongation to the rate of initiation and explains why elongation is still fast and processive in vivo even without anti-arrest factors.

Adenosine Triphosphate↗
PubMed2003

The riboswitch-mediated control of sulfur metabolism in bacteria.

Many operons in Gram-positive bacteria that are involved in methionine (Met) and cysteine (Cys) biosynthesis possess an evolutionarily conserved regulatory leader sequence (S-box) that positively controls these genes in response to methionine starvation. Here, we demonstrate that a feed-back regulation mechanism utilizes S-adenosyl-methionine as an effector. S-adenosyl-methionine directly and specifically binds to the nascent S-box RNA, causing an intrinsic terminator to form and interrupt transcription prematurely. The S-box leader RNA thus expands the family of newly discovered riboswitches, i.e., natural regulatory RNA aptamers that seem to sense small molecules ranging from amino acid derivatives to vitamins.

Base Sequence↗
PubMed2003

RNA polymerase holoenzyme: structure, function and biological implications.

The past three years have marked the breakthrough in our understanding of the structural and functional organization of RNA polymerase. The latest major advance was the high-resolution structures of bacterial RNA polymerase holoenzyme and the holoenzyme in complex with promoter DNA. Together with an array of genetic, biochemical and biophysical data accumulated to date, the structures provide a comprehensive view of dynamic interactions between the major components of transcription machinery during the early stages of the transcription cycle. They include the binding of sigma factor to the core enzyme, and the recognition of promoter sequences and DNA melting by holoenzyme, transcription initiation and promoter clearance.

Bacteria↗