scholarly journals Degradation of Lon in Caulobacter crescentus

2020 ◽  
Author(s):  
Benjamin B Barros ◽  
Samar A Mahmoud ◽  
Peter Chien ◽  
Rilee D. Zeinert
Keyword(s):  
Translational Control ◽  
Caulobacter Crescentus ◽  
Feedback Mechanism ◽  
Transcriptional Level ◽  
Lon Protease ◽  
Low Levels ◽  
Critical Machines ◽  
Carboxy Terminal ◽  
Replication Initiator ◽  
Proteolytic Capacity

AbstractProtein degradation is an essential process in all organisms. This process is irreversible and energetically costly; therefore, protein destruction must be tightly controlled. While environmental stresses often lead to upregulation of proteases at the transcriptional level, little is known about post-translational control of these critical machines. In this study we show that in Caulobacter crescentus levels of the Lon protease are controlled through proteolysis. Lon turnover requires active Lon and ClpAP proteases. We show that specific determinants dictate Lon stability with a key carboxy-terminal histidine residue driving recognition. Expression of stabilized Lon variants results in toxic levels of protease that deplete normal Lon substrates such as the replication initiator DnaA to lethally low levels. Taken together, this work demonstrates a feedback mechanism in which ClpAP and Lon collaborate to tune Lon proteolytic capacity for the cell.ImportanceProteases are essential, but unrestrained activity can also kill cells by degrading essential proteins. The quality control protease Lon must degrade many misfolded and native substrates. We show that Lon is itself controlled through proteolysis and that bypassing this control results in toxic consequences for the cell.

scholarly journals Degradation of Lon in Caulobacter crescentus

2020 ◽  
Vol 203 (1) ◽  
Author(s):  
Benjamin B. Barros ◽  
Samar A. Mahmoud ◽  
Peter Chien ◽  
Rilee D. Zeinert
Keyword(s):  
Caulobacter Crescentus ◽  
Feedback Mechanism ◽  
Transcriptional Level ◽  
Lon Protease ◽  
Content Type ◽  
Low Levels ◽  
Critical Machines ◽  
Carboxy Terminal ◽  
Replication Initiator ◽  
Proteolytic Capacity

ABSTRACT Protein degradation is an essential process in all organisms. This process is irreversible and energetically costly; therefore, protein destruction must be tightly controlled. While environmental stresses often lead to upregulation of proteases at the transcriptional level, little is known about posttranslational control of these critical machines. In this study, we show that in Caulobacter crescentus levels of the Lon protease are controlled through proteolysis. Lon turnover requires active Lon and ClpAP proteases. We show that specific determinants dictate Lon stability with a key carboxy-terminal histidine residue driving recognition. Expression of stabilized Lon variants results in toxic levels of protease that deplete normal Lon substrates, such as the replication initiator DnaA, to lethally low levels. Taken together, results of this work demonstrate a feedback mechanism in which ClpAP and Lon collaborate to tune Lon proteolytic capacity for the cell. IMPORTANCE Proteases are essential, but unrestrained activity can also kill cells by degrading essential proteins. The quality-control protease Lon must degrade many misfolded and native substrates. We show that Lon is itself controlled through proteolysis and that bypassing this control results in toxic consequences for the cell.

scholarly journals Lon recognition of the replication initiator DnaA is not confined to a single degron

2018 ◽  
Author(s):  
Jing Liu ◽  
Laura Francis ◽  
Peter Chien
Keyword(s):  
Acute Stress ◽  
Caulobacter Crescentus ◽  
Initiation Site ◽  
Active State ◽  
Lon Protease ◽  
Species Specific ◽  
Replication Initiator

SummaryDnaA initiates chromosome replication in bacteria. In Caulobacter crescentus, the Lon protease degrades DnaA to coordinate replication with nutrient availability and to halt the cell cycle during acute stress. Here we characterize the mechanism of DnaA recognition by Lon. We find that the native folded state of DnaA is crucial for its degradation, in contrast to the well-known role of Lon in degrading misfolded proteins. We fail to identify a single degradation motif (degron) sufficient for DnaA degradation, rather we show that both the ATPase domain and a species-specific N-terminal motif are important for productive Lon degradation of DnaA. Mutations in either of these determinants disrupt DnaA degradation in vitro and in vivo. DnaA switches from an inactive to active state depending on its nucleotide state and we find that locking DnaA in an active state inhibits degradation. Our working model is that Lon engages DnaA through at least two elements, one of which anchors DnaA to Lon and the other acting as an initiation site for degradation.

scholarly journals Expression of a gene duplication encoding conserved sperm tail proteins is translationally regulated in Drosophila melanogaster.

1993 ◽  
Vol 13 (3) ◽  
pp. 1708-1718 ◽  
Author(s):  
M Schäfer ◽  
D Börsch ◽  
A Hülster ◽  
U Schäfer
Keyword(s):  
Drosophila Melanogaster ◽  
Gene Family ◽  
Translational Control ◽  
Germ Line ◽  
Gene Families ◽  
Homologous Gene ◽  
Male Sterile ◽  
Sperm Tail ◽  
Repetitive Motif ◽  
Carboxy Terminal

We have analyzed a locus of Drosophila melanogaster located at 98C on chromosome 3, which contains two tandemly arranged genes, named Mst98Ca and Mst98Cb. They are two additional members of the Mst(3)CGP gene family by three criteria. (i) Both genes are exclusively transcribed in the male germ line. (ii) Both transcripts encode a protein with a high proportion of the repetitive motif Cys-Gly-Pro. (iii) Their expression is translationally controlled; while transcripts can be detected in diploid stages of spermatogenesis, association with polysomes can be shown only in haploid stages of sperm development. The genes differ markedly from the other members of the gene family in structure; they do not contain introns, they are of much larger size, and they have the Cys-Gly-Pro motifs clustered at the carboxy-terminal end of the encoded proteins. An antibody generated against the Mst98Ca protein recognizes both Mst98C proteins in D. melanogaster. In a male-sterile mutation in which spermiogenesis is blocked before individualization of sperm, both of these proteins are no longer synthesized. This finding provides proof of late translation for the Mst98C proteins and thereby independent proof of translational control of expression. Northern (RNA) and Western immunoblot analyses indicate the presence of homologous gene families in many other Drosophila species. The Mst98C proteins share sequence homology with proteins of the outer dense fibers in mammalian spermatozoa and can be localized to the sperm tail by immunofluorescence with an anti-Mst98Ca antibody.

Expression of a gene duplication encoding conserved sperm tail proteins is translationally regulated in Drosophila melanogaster

1993 ◽  
Vol 13 (3) ◽  
pp. 1708-1718
Author(s):  
M Schäfer ◽  
D Börsch ◽  
A Hülster ◽  
U Schäfer
Keyword(s):  
Drosophila Melanogaster ◽  
Gene Family ◽  
Translational Control ◽  
Germ Line ◽  
Gene Families ◽  
Homologous Gene ◽  
Male Sterile ◽  
Sperm Tail ◽  
Repetitive Motif ◽  
Carboxy Terminal

We have analyzed a locus of Drosophila melanogaster located at 98C on chromosome 3, which contains two tandemly arranged genes, named Mst98Ca and Mst98Cb. They are two additional members of the Mst(3)CGP gene family by three criteria. (i) Both genes are exclusively transcribed in the male germ line. (ii) Both transcripts encode a protein with a high proportion of the repetitive motif Cys-Gly-Pro. (iii) Their expression is translationally controlled; while transcripts can be detected in diploid stages of spermatogenesis, association with polysomes can be shown only in haploid stages of sperm development. The genes differ markedly from the other members of the gene family in structure; they do not contain introns, they are of much larger size, and they have the Cys-Gly-Pro motifs clustered at the carboxy-terminal end of the encoded proteins. An antibody generated against the Mst98Ca protein recognizes both Mst98C proteins in D. melanogaster. In a male-sterile mutation in which spermiogenesis is blocked before individualization of sperm, both of these proteins are no longer synthesized. This finding provides proof of late translation for the Mst98C proteins and thereby independent proof of translational control of expression. Northern (RNA) and Western immunoblot analyses indicate the presence of homologous gene families in many other Drosophila species. The Mst98C proteins share sequence homology with proteins of the outer dense fibers in mammalian spermatozoa and can be localized to the sperm tail by immunofluorescence with an anti-Mst98Ca antibody.

STUDIES ON A HUMAN CHORIONIC GONADOTROPHIN-LIKE MATERIAL PRESENT IN NON-PREGNANT SUBJECTS

1978 ◽  
Vol 89 (3) ◽  
pp. 492-505 ◽  
Author(s):  
D. M. Robertson ◽  
H. Suginami ◽  
H. Hernandez Montes ◽  
C. P. Puri ◽  
S. K. Choi ◽  
...  
Keyword(s):  
Gel Filtration ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Specific Sequence ◽  
Gonadotrophin Secretion ◽  
Carboxy Terminal Region ◽  
Assay Methods ◽  
Low Levels ◽  
Carboxy Terminal

ABSTRACT The presence of an hCG-like material in urinary and pituitary extracts and plasma obtained from non-pregnant subjects was investigated. Two assay methods were used to detect this material following fractionation of pituitary and urinary extracts by gel filtration (Ultrogel AcA 54) and/or isoelectrofocusing: a) a radioimmunoassay employing an antiserum raised against a specific sequence of the carboxy terminal region (residues 115– 145) of the β-subunit of hCG, and b) an in vitro bioassay method which measures both hLH and hCG activities. The fractionation procedures employed provide a satisfactory separation of highly purified hCG and hLH preparations. In the pituitary and urinary extracts hCGβ-peptide-like immunoactive (PIA) material was found consistently, which co-eluted with iodinated hCG following gel filtration and possessed pI values similar to those of hCG when subjected to isoelectrofocusing. The PIA material also exhibited in vitro biological activity similar to that shown by hLH and hCG. Detectable levels of immunoactive material were also found in plasma; however, the plasma levels of this PIA material were not influenced by classical endocrine measures such as the stimulation or inhibition of gonadotrophin secretion. The low levels of this material in plasma precluded its further characterization by gel filtration or electrofocusing. Whereas the present data and those reported by other investigators seem to suggest the presence of some hCG-like material in urinary and pituitary extracts and possibly in plasma of non-pregnant subjects, it is emphasized that the available evidence is not sufficiently conclusive to exclude other interpretations as to the nature of this material.

scholarly journals SAT-435 A Direct Comparison of Thyroid Hormone Receptor (THR) Protein Levels in Mice Provides Unexpected Insights into TH Action

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Sanya Bansal ◽  
Svetlana Minakhina ◽  
Alice Zhang ◽  
Michael Brotherton ◽  
Rucha Janodia ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Translational Control ◽  
Thyroid Hormone Receptor ◽  
Mrna Levels ◽  
Peripheral Tissues ◽  
Protein Levels ◽  
Peripheral Organs ◽  
Low Levels ◽  
First Time ◽  
Predominant Expression

Abstract Thyroid Hormone (TH) action is mediated by three major THR isoforms α1, β1 and β2 (THRA1, THRB1 and THRB2), encoded by two genes Thra and Thrb. A fourth non-T3 binding isoform is termed THRA2. Previous characterization and comparison of THR isoform expression using QPCR of mRNA levels does not assess translational or post-translational control of THR protein levels. Moreover, reliable antibodies against all THR isoforms are not currently available. To address these concerns, we generated knock-in mouse models expressing endogenously and identically 2X HA tagged THRs (THRA1/2, THRB1 and THRB2), which could then be detected by commercially available anti-HA antibodies. We characterized THR expression in 16 mouse organs. We found that in all peripheral tissues tested except the liver, the dominant THR isoform is THRA1, and THRB1 and THRA2 are expressed at significantly lower levels or are undetectable. Surprising THRB1 levels were very low in metabolically active organs such fat and muscle. In some organs, mRNA transcript levels predicted THR protein amounts, while in others the prediction was inaccurate. For instance, adipose depots have similar levels of Thrb1 and Thra1 transcripts, however THRA1 protein levels are up to 10-fold higher than THRB1. In contrast to peripheral organs, brain tissues express low levels of both THRB1 and THRA1, but have very high levels of THRA2. As expected and confirmed in this study, THRB2 has limited expression and was only detected in pituitary of the organs tested. Interestingly, THRB2 expression in female was much higher than male mice (the only sex-difference in THR expression found); and expression of the THRB2 target gene, Tshb, was lower in female mice. For the first time, these HA-THR mouse lines can be used to accurately compare isoform-specific actions of THRs in organs. The predominant expression of THRA1 in most peripheral organs, for example, suggests that many peripheral actions of THRB1 could be indirectly mediated.

Translational control of oskar generates short OSK, the isoform that induces pole plasma assembly

Development ◽  
1995 ◽  
Vol 121 (11) ◽  
pp. 3723-3732 ◽  
Author(s):  
F.H. Markussen ◽  
A.M. Michon ◽  
W. Breitwieser ◽  
A. Ephrussi
Keyword(s):  
Positive Feedback ◽  
Translational Control ◽  
Feedback Mechanism ◽  
Posterior Pole ◽  
Start Codon ◽  
Anterior Pole ◽  
Wild Type ◽  
Alternative Start Codon ◽  
Positive Feedback Mechanism ◽  
Pole Plasm

At the posterior pole of the Drosophila oocyte, oskar induces a tightly localized assembly of pole plasm. This spatial restriction of oskar activity has been thought to be achieved by the localization of oskar mRNA, since mislocalization of the RNA to the anterior induces anterior pole plasm. However, ectopic pole plasm does not form in mutant ovaries where oskar mRNA is not localized, suggesting that the unlocalized mRNA is inactive. As a first step towards understanding how oskar activity is restricted to the posterior pole, we analyzed oskar translation in wild type and mutants. We show that the targeting of oskar activity to the posterior pole involves two steps of spatial restriction, cytoskeleton-dependent localization of the mRNA and localization-dependent translation. Furthermore, our experiments demonstrate that two isoforms of Oskar protein are produced by alternative start codon usage. The short isoform, which is translated from the second in-frame AUG of the mRNA, has full oskar activity. Finally, we show that when oskar RNA is localized, accumulation of Oskar protein requires the functions of vasa and tudor, as well as oskar itself, suggesting a positive feedback mechanism in the induction of pole plasm by oskar.

scholarly journals Involvement of bone-specific alkaline phosphatase and procollagen I carboxy-terminal propeptide as predictors of early fracture risk in Chinese Children with juvenile osteoporosis: An interventional clinical trial

2018 ◽  
Vol 13 (2) ◽  
pp. 164
Author(s):  
Yu Wang ◽  
Jingxin Zhao ◽  
Lingwei Kong ◽  
Yu Jin
Keyword(s):  
Clinical Trial ◽  
Vitamin D ◽  
Alkaline Phosphatase ◽  
Bone Fracture ◽  
Specific Alkaline Phosphatase ◽  
Bone Specific Alkaline Phosphatase ◽  
Low Levels ◽  
Carboxy Terminal ◽  
To Receive

<p class="Abstract">This clinical trial was designed to understand whether the children with juvenile osteoporosis receiving tablet containing vitamin D and calcium had lower incidence of bone fracture compared to the children receiving a diet rich in calcium, vitamin D, and protein. We assessed whether plasma levels of bone-specific alkaline phosphatase (BSAP) and procollagen I carboxy-terminal propeptide levels (PIPC) could be used as predictors of early bone fracture in children. A total of 120 children of either gender with a juvenile osteoporosis were enrolled and randomized (1:1 ratio) to receive tablet containing vitamin D and calcium (n=60) or diet rich in calcium, vitamin D, and protein (n=60), and undergone follow-up for up to 3 years.  Blood sample was collected and BSAP and PIPC levels were measured. The results suggested that therapeutic intervention (vitamin D and calcium) does not predict bone fracture in children. However, correlations analysis revealed that the decreased level of BSAP and PIPC were associated with higher incidence of fracture. The results suggest that the low levels of BSAP and PIPC cause increase susceptibility of fracture among children with juvenile osteoporosis.</p>

RNA thermometers are common in α- and γ-proteobacteria

2005 ◽  
Vol 386 (12) ◽  
pp. 1279-1286 ◽  
Author(s):  
Torsten Waldminghaus ◽  
Anja Fippinger ◽  
Juliane Alfsmann ◽  
Franz Narberhaus
Keyword(s):  
Heat Shock ◽  
Translational Control ◽  
Caulobacter Crescentus ◽  
Site Directed Mutagenesis ◽  
Reporter System ◽  
Heat Shock Genes ◽  
Regulatory Modules ◽  
Secondary Structure Predictions ◽  
Shine Dalgarno Sequence ◽  
The Rose

AbstractExpression of many rhizobial small heat-shock genes is controlled by the ROSE element, a thermoresponsive structure in the 5′-untranslated region of the corresponding mRNAs. Using a bioinformatics approach, we found more than 20 new potential ROSE-like RNA thermometers upstream of small heat-shock genes in a wide variety of α- and γ-proteobacteria. Northern blot analyses revealed heat-inducible transcripts of the representative candidateCaulobacter crescentus CC2258,Escherichia coli ibpAandSalmonella typhimurium ibpAgenes. Typical σ32-type promoters were mapped upstream of the potential RNA thermometers by primer extension. Additional translational control was demonstrated in alacZreporter system and by site-directed mutagenesis. RNA secondary structure predictions strongly suggest that the Shine-Dalgarno sequence in the RNA thermometers is masked at low temperatures. Combining two regulatory modules, a σ32promoter and a ROSE-type RNA thermometer, provides a novel stringent mechanism to control expression of small heat-shock genes.

Photocontrol of enzyme activation and inactivation

1983 ◽  
Vol 303 (1116) ◽  
pp. 433-441 ◽  
Keyword(s):  
Translational Control ◽  
Messenger Rna ◽  
Enzyme Synthesis ◽  
Good Evidence ◽  
Enzyme Activation ◽  
Transcriptional Level ◽  
Plant Enzymes ◽  
Activation Mechanisms ◽  
Theoretical Considerations ◽  
Plant Enzyme

During recent years the question of whether phytochrome regulates certain plant enzyme activities by influencing the rates of enzyme synthesis or by post-translational activation mechanisms has been vigorously debated. There is now good evidence that phytochrome can regulate concentrations of specific messenger RNA populations in some situations. However, the capacity to exert control at a transcriptional level in some systems does not necessarily preclude the possibility that regulation could occur at a post-translational level elsewhere, or even within the same cells. Theoretical considerations apart, the evidence for activation of plant enzymes by phytochrome is not generally strong. Some of the enzymes whose activity is known to be modulated by phytochrome have also been shown to possess post-translational control systems or exist in inactive forms so that the molecular possibilities for such modulation do seem to occur. The lack of direct evidence for the control of such processes by phytochrome may well reflect the technical difficulties involved in this sort of investigation.

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