RelA-SpoT Homolog (RSH) enzymes control bacterial physiology through synthesis and degradation of the nucleotide alarmone (p)ppGpp. We recently discovered multiple families of small alarmone synthetase (SAS) RSH acting as toxins of toxin-antitoxin (TA) modules, with the FaRel subfamily of toxSAS abrogating bacterial growth by producing an analog of (p)ppGpp, (pp)pApp. Here we probe the mechanism of growth arrest used by four experimentally unexplored subfamilies of toxSAS: FaRel2, PhRel, PhRel2, and CapRel. Surprisingly, all these toxins specifically inhibit protein synthesis. To do so, they transfer a pyrophosphate moiety from ATP to the tRNA 3' CCA. The modification inhibits both tRNA aminoacylation and the sensing of cellular amino acid starvation by the ribosome-associated RSH RelA. Conversely, we show that some small alarmone hydrolase (SAH) RSH enzymes can reverse the pyrophosphorylation of tRNA to counter the growth inhibition by toxSAS. Collectively, we establish RSHs as RNA-modifying enzymes.
- MeSH
- bakteriální toxiny genetika metabolismus farmakologie MeSH
- fosforylace účinky léků MeSH
- grampozitivní nesporulující tyčinky chemie metabolismus MeSH
- guanosinpentafosfát chemie metabolismus MeSH
- inhibitory syntézy proteinů farmakologie MeSH
- ligasy chemie genetika metabolismus MeSH
- proteosyntéza účinky léků fyziologie MeSH
- pyrofosfatasy MeSH
- ribozomy metabolismus MeSH
- RNA transferová metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Mitoribosomes consist of ribosomal RNA and protein components, coordinated assembly of which is critical for function. We used mitoribosomes from Trypanosoma brucei with reduced RNA and increased protein mass to provide insights into the biogenesis of the mitoribosomal large subunit. Structural characterization of a stable assembly intermediate revealed 22 assembly factors, some of which have orthologues/counterparts/homologues in mammalian genomes. These assembly factors form a protein network that spans a distance of 180 Å, shielding the ribosomal RNA surface. The central protuberance and L7/L12 stalk are not assembled entirely and require removal of assembly factors and remodeling of the mitoribosomal proteins to become functional. The conserved proteins GTPBP7 and mt-EngA are bound together at the subunit interface in proximity to the peptidyl transferase center. A mitochondrial acyl-carrier protein plays a role in docking the L1 stalk, which needs to be repositioned during maturation. Additional enzymatically deactivated factors scaffold the assembly while the exit tunnel is blocked. Together, this extensive network of accessory factors stabilizes the immature sites and connects the functionally important regions of the mitoribosomal large subunit.
- MeSH
- elektronová kryomikroskopie MeSH
- konformace proteinů MeSH
- mitochondriální ribozomy metabolismus MeSH
- proteiny vázající GTP metabolismus MeSH
- proteosyntéza fyziologie MeSH
- RNA ribozomální genetika MeSH
- Trypanosoma brucei brucei metabolismus MeSH
- vazba proteinů fyziologie MeSH
- velké ribozomální podjednotky metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Regeneration capacity is reduced as CNS axons mature. Using laser-mediated axotomy, proteomics and puromycin-based tagging of newly-synthesized proteins in a human embryonic stem cell-derived neuron culture system that allows isolation of axons from cell bodies, we show here that efficient regeneration in younger axons (d45 in culture) is associated with local axonal protein synthesis (local translation). Enhanced regeneration, promoted by co-culture with human glial precursor cells, is associated with increased axonal synthesis of proteins, including those constituting the translation machinery itself. Reduced regeneration, as occurs with the maturation of these axons by d65 in culture, correlates with reduced levels of axonal proteins involved in translation and an inability to respond by increased translation of regeneration promoting axonal mRNAs released from stress granules. Together, our results provide evidence that, as in development and in the PNS, local translation contributes to CNS axon regeneration.
- MeSH
- axony fyziologie MeSH
- embryonální kmenové buňky fyziologie MeSH
- kokultivační techniky MeSH
- lidé MeSH
- proteosyntéza fyziologie MeSH
- regenerace nervu fyziologie MeSH
- stárnutí buněk fyziologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Axon regeneration in the CNS is inhibited by many extrinsic and intrinsic factors. Because these act in parallel, no single intervention has been sufficient to enable full regeneration of damaged axons in the adult mammalian CNS. In the external environment, NogoA and CSPGs are strongly inhibitory to the regeneration of adult axons. CNS neurons lose intrinsic regenerative ability as they mature: embryonic but not mature neurons can grow axons for long distances when transplanted into the adult CNS, and regeneration fails with maturity in in vitro axotomy models. The causes of this loss of regeneration include partitioning of neurons into axonal and dendritic fields with many growth-related molecules directed specifically to dendrites and excluded from axons, changes in axonal signalling due to changes in expression and localization of receptors and their ligands, changes in local translation of proteins in axons, and changes in cytoskeletal dynamics after injury. Also with neuronal maturation come epigenetic changes in neurons, with many of the transcription factor binding sites that drive axon growth-related genes becoming inaccessible. The overall aim for successful regeneration is to ensure that the right molecules are expressed after axotomy and to arrange for them to be transported to the right place in the neuron, including the damaged axon tip.
- MeSH
- axonální transport fyziologie MeSH
- axony fyziologie MeSH
- centrální nervový systém cytologie fyziologie MeSH
- lidé MeSH
- nervový útlum fyziologie MeSH
- neurogeneze fyziologie MeSH
- proteosyntéza fyziologie MeSH
- regenerace nervu fyziologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Interferon-induced transmembrane proteins IFITM1 and IFITM3 (IFITM1/3) play a role in both RNA viral restriction and in human cancer progression. Using immunohistochemical staining of FFPE tissue, we identified subgroups of cervical cancer patients where IFITM1/3 protein expression is inversely related to metastasis. Guide RNA-CAS9 methods were used to develop an isogenic IFITM1/IFITM3 double null cervical cancer model in order to define dominant pathways triggered by presence or absence of IFITM1/3 signalling. A pulse SILAC methodology identified IRF1, HLA-B, and ISG15 as the most dominating IFNγ inducible proteins whose synthesis was attenuated in the IFITM1/IFITM3 double-null cells. Conversely, SWATH-IP mass spectrometry of ectopically expressed SBP-tagged IFITM1 identified ISG15 and HLA-B as dominant co-associated proteins. ISG15ylation was attenuated in IFNγ treated IFITM1/IFITM3 double-null cells. Proximity ligation assays indicated that HLA-B can interact with IFITM1/3 proteins in parental SiHa cells. Cell surface expression of HLA-B was attenuated in IFNγ treated IFITM1/IFITM3 double-null cells. SWATH-MS proteomic screens in cells treated with IFITM1-targeted siRNA cells resulted in the attenuation of an interferon regulated protein subpopulation including MHC Class I molecules as well as IFITM3, STAT1, B2M, and ISG15. These data have implications for the function of IFITM1/3 in mediating IFNγ stimulated protein synthesis including ISG15ylation and MHC Class I production in cancer cells. The data together suggest that pro-metastatic growth associated with IFITM1/3 negative cervical cancers relates to attenuated expression of MHC Class I molecules that would support tumor immune escape.
- MeSH
- buněčné linie MeSH
- diferenciační antigeny fyziologie MeSH
- histokompatibilita - antigeny třídy I metabolismus MeSH
- lidé MeSH
- membránové proteiny fyziologie MeSH
- nádory děložního čípku metabolismus MeSH
- proteiny vázající RNA fyziologie MeSH
- proteosyntéza fyziologie MeSH
- Check Tag
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Protein production must be strictly controlled at its beginning and end to synthesize a polypeptide that faithfully copies genetic information carried in the encoding mRNA. In contrast to viruses and prokaryotes, the majority of mRNAs in eukaryotes contain only one coding sequence, resulting in production of a single protein. There are, however, many exceptional mRNAs that either carry short open reading frames upstream of the main coding sequence (uORFs) or even contain multiple long ORFs. A wide variety of mechanisms have evolved in microbes and higher eukaryotes to prevent recycling of some or all translational components upon termination of the first translated ORF in such mRNAs and thereby enable subsequent translation of the next uORF or downstream coding sequence. These specialized reinitiation mechanisms are often regulated to couple translation of the downstream ORF to various stimuli. Here we review all known instances of both short uORF-mediated and long ORF-mediated reinitiation and present our current understanding of the underlying molecular mechanisms of these intriguing modes of translational control.
- MeSH
- Bacteria genetika metabolismus MeSH
- Eukaryota genetika MeSH
- lidé MeSH
- otevřené čtecí rámce genetika MeSH
- proteosyntéza genetika fyziologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
- Research Support, N.I.H., Intramural MeSH
Sarkopenie patří mezi nejzávažnější poruchy spojené s nutriční dysbalancí. Přestože je sarkopenie s postupujícím věkem neodlučitelnou složkou v procesu stárnutí, hraje samostatnou vitální roli v patologických situacích, jako je podvýživa proteino-kalorického i kwashiorkorového typu, provází řadu onemocnění revmatologického charakteru, je součástí prakticky vždy klinického obrazu u onkologických pacientů. Sarkopenie ve stresové zátěži se fatálně projevuje zejména ve stáří u tzv. křehkých pacientů. Vzhledem k tomu, že sarkopenie rozhoduje o průběhu chronických i akutních onemocnění, je nezbytné ji včas diagnostikovat, monitorovat a správně léčit. I když výživa není jediným faktorem, který zejména ve stáří se na vzniku a rozvoji sarkopenie podílí, je typ nutriční podpory a v nové době, zejména nutriční farmakologie, principiálním přístupem v komplexní léčbě pacienta.
Sarcopenia is one of the severe disorders associated with nutritional imbalance. While sarcopenia is an inseparable part of the process of ageing as people get older, it plays a vital role in pathological situations such as protein-calorie and kwashiorkor malnutrition, it accompanies various diseases of rheumatic character and is an almost ever-present feature of the clinical picture of oncological patients. Sarcopenia in people under strain is fatally manifested mainly in the elderly, medically fragile patients. In view of the fact that sarcopenia is the decisive factor regarding the course of chronic as well as acute illnesses, it must be diagnosed, monitored and appropriately treated in time. Although nutrition is not the only factor involved especially in older age in the development of sarcopenia, the type of nutritional support and, in the new era, particularly nutritional pharmacology, form the underlying principles of the approach to the comprehensive care of the patient.
- MeSH
- aminokyseliny metabolismus MeSH
- dietní proteiny metabolismus MeSH
- elektrická impedance MeSH
- kachexie komplikace MeSH
- kosterní svaly metabolismus MeSH
- kreatinin metabolismus MeSH
- leucin metabolismus MeSH
- lidé MeSH
- nedostatek vitaminu D MeSH
- oxidační stres fyziologie MeSH
- podvýživa * komplikace metabolismus MeSH
- proteosyntéza fyziologie MeSH
- sarkopenie * diagnóza etiologie prevence a kontrola MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- stárnutí * MeSH
- svalová síla - dynamometr MeSH
- tloušťka kožní řasy MeSH
- Check Tag
- lidé MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- Publikační typ
- práce podpořená grantem MeSH
- přehledy MeSH
Nucleic acid sequence complementarity underlies many fundamental biological processes. Although first noticed a long time ago, sequence complementarity between mRNAs and ribosomal RNAs still lacks a meaningful biological interpretation. Here we used statistical analysis of large-scale sequence data sets and high-throughput computing to explore complementarity between 18S and 28S rRNAs and mRNA 3' UTR sequences. By the analysis of 27,646 full-length 3' UTR sequences from 14 species covering both protozoans and metazoans, we show that the computed 18S rRNA complementarity creates an evolutionarily conserved localization pattern centered around the ribosomal mRNA entry channel, suggesting its biological relevance and functionality. Based on this specific pattern and earlier data showing that post-termination 80S ribosomes are not stably anchored at the stop codon and can migrate in both directions to codons that are cognate to the P-site deacylated tRNA, we propose that the 18S rRNA-mRNA complementarity selectively stabilizes post-termination ribosomal complexes to facilitate ribosome recycling. We thus demonstrate that the complementarity between 18S rRNA and 3' UTRs has a non-random nature and very likely carries information with a regulatory potential for translational control.
- MeSH
- 3' nepřekládaná oblast * MeSH
- kodon MeSH
- proteosyntéza fyziologie MeSH
- RNA ribozomální chemie fyziologie MeSH
- terminátorové oblasti (genetika) * MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Příjem bílkovin a odporový trénink jsou faktory ovlivňující míru svalové proteosyntézy. Jedním z adaptačních projevů je svalová hypertrofie jako důsledek pozitivní bílkovinné bilance mezi mírou svalové proteosyntézy a svalové proteolýzy. Příjem bílkovin s obsahem esenciálních aminokyselin v období po zatížení je nutným předpokladem účinné adaptace. Postprandiální vzestup aminoacidémie je stěžejním faktorem efektivity proteosyntetických dějů. Mezi nejpoužívanější druhy bílkovin uplatňované ve sportovní výživě patří mléčné bílkoviny, vaječná a sójová bílkovina. Existují mezi nimi značné rozdíly v digesci a kinetice resorbovaných esenciálních aminokyselin. Syrovátková bílkovina je pro svůj obsah esenciálních aminokyselin a zastoupení leucinu nadřazená ostatním bílkovinným zdrojům a je považována za významný anabolický stimulus. Jednorázové množství ~20–25 g syrovátkové bílkoviny zkonzumované po skončení zatížení optimalizuje podmínky pro svalovou hypertrofii. Vhodné načasování příjmu bílkovin do těsné blízkosti zatížení pozitivně ovlivňuje snahu sportovce dosáhnout svalové hypertrofie.
Protein intake and resistance training are factors affecting the rate of muscle protein synthesis. Muscle hypertrophy manifests as a result of a positive protein balance between the rate of muscle protein synthesis and muscle protein breakdown. An intake of protein containing essential amino acids in the period after the resistance exercise is a key prerequisite of an effective adaptation. Postprandial hyperaminoacidemia is a key factor in the effectiveness of proteosynthetic processes. The most common types of protein used in sports nutrition include milk protein, egg and soy protein. There are significant differences among them in the context of digestion and essential amino acids resorptio kinetics. Whey protein represents due to the high content of essential amino acids and leucine a superior source and is considered an important anabolic stimuli. An amount of ~20–25 g of whey protein consumed after restistance exercise optimizes conditions for muscle hypertrophy. Appropriate timing of protein intake in close proximity to the load positively affects an athletes attempt to achieve muscle hypertrophy.
- MeSH
- časové faktory MeSH
- esenciální aminokyseliny MeSH
- fyziologie sportovní výživy MeSH
- hypertrofie MeSH
- kosterní svaly metabolismus MeSH
- leucin MeSH
- lidé MeSH
- mléčné bílkoviny MeSH
- odporový trénink * MeSH
- proteosyntéza * fyziologie MeSH
- regenerace fyziologie MeSH
- sportovní nutriční věda MeSH
- svalové proteiny * biosyntéza MeSH
- syrovátka MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- přehledy MeSH
