The main function of the bacterial phosphotransferase system is to transport and to phosphorylate mono- and disaccharides as well as sugar alcohols. However, the phosphotransferase system is also involved in regulation of carbon metabolism. In Gram-positive bacteria, it is implicated in carbon catabolite repression and regulation of expression of catabolic genes by controlling either catabolic enzyme activities, transcriptional activators or antiterminators. All these different regulations follow a protein phosphorylation mechanism.

• MeSH
• bakteriální proteiny metabolismus   MeSH
• fosfoenolpyruvát-cukr-fosfotransferasový systém metabolismus   MeSH
• fosforylace MeSH
• grampozitivní bakterie genetika   metabolismus   MeSH
• metabolismus sacharidů * MeSH
• modely genetické MeSH
• regulace genové exprese u bakterií * MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• bakteriální proteiny MeSH
• fosfoenolpyruvát-cukr-fosfotransferasový systém MeSH

Exocyst is a heterooctameric protein complex crucial for the tethering of secretory vesicles to the plasma membrane during exocytosis. Compared to other eukaryotes, exocyst subunit EXO70 is represented by many isoforms in land plants whose cell biological and biological roles, as well as modes of regulation remain largely unknown. Here, we present data on the phospho-regulation of exocyst isoform EXO70C2, which we previously identified as a putative negative regulator of exocyst function in pollen tube growth. A comprehensive phosphoproteomic analysis revealed phosphorylation of EXO70C2 at multiple sites. We have now performed localization and functional studies of phospho-dead and phospho-mimetic variants of Arabidopsis EXO70C2 in transiently transformed tobacco pollen tubes and stably transformed Arabidopsis wild type and exo70C2 mutant plants. Our data reveal a dose-dependent effect of AtEXO70C2 overexpression on pollen tube growth rate and cellular architecture. We show that changes of the AtEXO70C2 phosphorylation status lead to distinct outcomes in wild type and exo70c2 mutant cells, suggesting a complex regulatory pattern. On the other side, phosphorylation does not affect the cytoplasmic localization of AtEXO70C2 or its interaction with putative secretion inhibitor ROH1 in the yeast two-hybrid system.

• Klíčová slova
• Exo70, exocyst, membrane trafficking, phosphorylation, pollen tube, secretion inhibitor, tip-growth,
• Publikační typ
• časopisecké články MeSH

We posit the following hypothesis: Independently of whether malignant tumors are initiated by a fundamental reprogramming of gene expression or seeded by stem cells, "waves" of gene expression that promote metabolic changes occur during carcinogenesis, beginning with oncogene-mediated changes, followed by hypoxia-induced factor (HIF)-mediated gene expression, both resulting in the highly glycolytic "Warburg" phenotype and suppression of mitochondrial biogenesis. Because high proliferation rates in malignancies cause aglycemia and nutrient shortage, the third (second oncogene) "wave" of adaptation stimulates glutaminolysis, which in certain cases partially re-establishes oxidative phosphorylation; this involves the LKB1-AMPK-p53, PI3K-Akt-mTOR axes and MYC dysregulation. Oxidative glutaminolysis serves as an alternative pathway compensating for cellular ATP. Together with anoxic glutaminolysis it provides pyruvate, lactate, and the NADPH pool (alternatively to pentose phosphate pathway). Retrograde signaling from revitalized mitochondria might constitute the fourth "wave" of gene reprogramming. In turn, upon reversal of the two Krebs cycle enzymes, glutaminolysis may partially (transiently) function even during anoxia, thereby further promoting malignancy. The history of the carcinogenic process within each malignant tumor determines the final metabolic phenotype of the selected surviving cells, resulting in distinct cancer bioenergetic phenotypes ranging from the highly glycolytic "classic Warburg" to partial or enhanced oxidative phosphorylation. We discuss the bioenergetically relevant functions of oncogenes, the involvement of mitochondrial biogenesis/degradation in carcinogenesis, the yet unexplained Crabtree effect of instant glucose blockade of respiration, and metabolic signaling stemming from the accumulation of succinate, fumarate, pyruvate, lactate, and oxoglutarate by interfering with prolyl hydroxylase domain enzyme-mediated hydroxylation of HIFα prolines.

• MeSH
• 1-fosfatidylinositol-3-kinasa metabolismus   MeSH
• biologická adaptace fyziologie   MeSH
• energetický metabolismus fyziologie   MeSH
• geny myc fyziologie   MeSH
• glukosa metabolismus   MeSH
• glutamin metabolismus   MeSH
• hypoxie buňky * MeSH
• kyselina mléčná metabolismus   MeSH
• kyselina pyrohroznová metabolismus   MeSH
• lidé MeSH
• metabolické sítě a dráhy fyziologie   MeSH
• mitochondrie metabolismus   MeSH
• nádory metabolismus   MeSH
• oxidativní fosforylace * MeSH
• proliferace buněk MeSH
• protein-serin-threoninkinasy metabolismus   MeSH
• regulace genové exprese fyziologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• 1-fosfatidylinositol-3-kinasa MeSH
• glukosa MeSH
• glutamin MeSH
• kyselina mléčná MeSH
• kyselina pyrohroznová MeSH
• protein-serin-threoninkinasy MeSH

p53 is a tetrameric protein with a thermodynamically unstable deoxyribonucleic acid (DNA)-binding domain flanked by intrinsically disordered regulatory domains that control its activity. The unstable and disordered segments of p53 allow high flexibility as it interacts with binding partners and permits a rapid on/off switch to control its function. The p53 tetramer can exist in multiple conformational states, any of which can be stabilized by a particular modification. Here, we apply the allostery model to p53 to ask whether evidence can be found that the "activating" C-terminal phosphorylation of p53 stabilizes a specific conformation of the protein in the absence of DNA. We take advantage of monoclonal antibodies for p53 that measure indirectly the following conformations: unfolded, folded, and tetrameric. A double antibody capture enzyme linked-immunosorbent assay was used to observe evidence of conformational changes of human p53 upon phosphorylation by casein kinase 2 in vitro. It was demonstrated that oligomerization and stabilization of p53 wild-type conformation results in differential exposure of conformational epitopes PAb1620, PAb240, and DO12 that indicates a reduction in the "unfolded" conformation and increases in the folded conformation coincide with increases in its oligomerization state. These data highlight that the oligomeric conformation of p53 can be stabilized by an activating enzyme and further highlight the utility of the allostery model when applied to understanding the regulation of unstable and intrinsically disordered proteins.

• Klíčová slova
• CK2, allosteric regulation, conformational change, oligomerization, p53, phosphorylation, protein conformation, protein folding,
• MeSH
• adenosintrifosfát metabolismus   MeSH
• alosterická regulace MeSH
• fosforylace MeSH
• kaseinkinasa II metabolismus   MeSH
• lidé MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• mutace MeSH
• nádorový supresorový protein p53 chemie   genetika   metabolismus   MeSH
• proteinové domény MeSH
• stabilita proteinů MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• adenosintrifosfát MeSH
• kaseinkinasa II MeSH
• nádorový supresorový protein p53 MeSH
• TP53 protein, human MeSH Prohlížeč

Aquaporin (AQP) proteins are involved in water homeostasis in cells at all taxonomic levels of life. Phosphorylation of some AQPs has been proposed to regulate water permeability via gating of the channel itself. We analyzed plasma membrane intrinsic proteins (PIP) from Camelina and characterized their biological functions under both stressful and favorable conditions. A three-dimensional theoretical model of the Camelina AQP proteins was built by homology modeling which could prove useful in further functional characterization of AQPs. CsPIP2;1 was strongly and constitutively expressed in roots and leaves of Camelina, suggesting that this gene is related to maintenance of homeostasis during salt and drought stresses. CsPIP2s exhibited water channel activity in Xenopus oocytes. We then examined the roles of CsPIP2;1 phosphorylation at Ser273 and Ser277 in the regulation of water permeability using phosphorylation mutants. A single deletion strain of CsPIP2;1 was generated to serve as the primary host for testing AQP expression constructs. A Ser277 to alanine mutation (to prevent phosphorylation) did not change CsPIP2;1 water permeability while a Ser273 mutation to alanine did affect water permeability. Furthermore, a CsPIP2;1 point mutation when ectopically expressed in yeast resulted in lower growth in salt and drought conditions compared with controls, and confirmation of Ser273 as the phosphorylation site. Our results support the idea that post-translational modifications in the Ser273 regulatory domains of the C-terminus fine tune water flux through CsPIP2;1.

• Klíčová slova
• CsPIP2 ;1, Drought, Phosphorylation, Salt, Ser273,
• MeSH
• akvaporiny genetika   metabolismus   MeSH
• Brassicaceae účinky léků   genetika   fyziologie   MeSH
• chlorid sodný farmakologie   MeSH
• fosforylace MeSH
• fylogeneze MeSH
• fyziologický stres * MeSH
• listy rostlin účinky léků   genetika   fyziologie   MeSH
• molekulární sekvence - údaje MeSH
• molekulární struktura MeSH
• mutace MeSH
• období sucha MeSH
• oocyty MeSH
• regulace genové exprese u rostlin * MeSH
• rostlinné proteiny genetika   metabolismus   MeSH
• sekvence aminokyselin MeSH
• sekvenční seřazení MeSH
• serin MeSH
• voda metabolismus   MeSH
• Xenopus MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• akvaporiny MeSH
• chlorid sodný MeSH
• rostlinné proteiny MeSH
• serin MeSH
• voda MeSH

Changes of protein tyrosine phosphorylation in ejaculated boar sperm incubated in vitro were examined with the use of antiphosphotyrosine antibodies and immunoblotting. The intracellular levels of cAMP were modulated by treatment with various combinations of caffeine, 3-isobutyl-1-methylxanthine (IBMX), and dibutyryl cyclic AMP (dbcAMP), and acrosome reactions (ARs) were induced via treatment with divalent cation ionophore A23187. Proteins of Mr 34, 38, 40, and 44 (p34 ... p44) were strongly phosphorylated on tyrosine residues in freshly prepared sperm samples and at the same level during all subsequent treatments. Incubation of sperm in vitro for various periods of time induced an increase of tyrosine phosphorylation of p20, p93, and p175. The tyrosine phosphorylation of p93, p175, and several other sperm proteins was up-regulated in a concentration-dependent manner following treatment of the sperm with dbcAMP, caffeine, or IBMX alone, or with combinations of caffeine and IBMX, respectively, with dbcAMP; the tyrosine phosphorylation of p20 was not correlated with treatment of sperm with cAMP-elevating reagents. The percentage of sperm cells undergoing spontaneous ARs was not affected by the manipulation of cAMP levels and was not correlated with protein tyrosine phosphorylation. In contrast, the addition of calcium to the incubation media decreased protein tyrosine phosphorylation and elevated percentage of spontaneous ARs. The induction of ARs with A23187 caused a significant decrease of tyrosine phosphorylation of p93, p175, and p220/230, indicating that dephosphorylation on protein tyrosine residues might be associated with calcium influx during physiological ARs as well. Proteins p93 and p175 were effectively solubilized in greater than 9M urea/1% triton and in SDS sample buffer, but to only a small extent in triton, while p20 was virtually completely extractable with triton. In conjunction with the previously reported isolation of active tyrosine kinase sp42 from triton extracts of noncapacitated boar sperm cells (Berruti and Porzio, 1992: Biochim Biophys Acta 1118: 149-154), our results suggest that a cAMP-dependent event is required for tyrosine phosphorylation of triton-insoluble proteins such as p93 and p175. On the other hand, the tyrosine phosphorylation of p20 (and potentially other triton-soluble substrates) might not strictly require such cAMP up-regulation. We discuss the differences in the regulation of cAMP-dependent tyrosine phosphorylation in mouse, human, and boar sperm, and suggest that sensitivity to calcium and distinct basal levels of cyclic nucleotide PDE might correspond to species-specific reproduction strategies in mammals.

• MeSH
• AMP cyklický metabolismus   MeSH
• fosforylace MeSH
• kultivační média MeSH
• kultivované buňky MeSH
• lidé MeSH
• myši MeSH
• prasata MeSH
• signální transdukce * MeSH
• spermie metabolismus   MeSH
• tyrosin metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• AMP cyklický MeSH
• kultivační média MeSH
• tyrosin MeSH

Trypanosomatids regulate gene expression mainly at the post-transcriptional level through processing, exporting and stabilising mRNA and control of translation. In most eukaryotes, protein synthesis is regulated by phosphorylation of eukaryotic initiation factor 2 (eIF2) at serine 51. Phosphorylation halts overall translation by decreasing availability of initiator tRNAmet to form translating ribosomes. In trypanosomatids, the N-terminus of eIF2α is extended with threonine 169 the homologous phosphorylated residue. Here, we evaluated whether eIF2α phosphorylation varies during the Trypanosoma cruzi life cycle, the etiological agent of Chagas' disease. Total levels of eIF2α are diminished in infective and non-replicative trypomastigotes compared with proliferative forms from the intestine of the insect vector or amastigotes from mammalian cells, consistent with decreased protein synthesis reported in infective forms. eIF2α phosphorylation increases in proliferative intracellular forms prior to differentiation into trypomastigotes. Parasites overexpressing eIF2αT169A or with an endogenous CRISPR/Cas9-generated eIF2αT169A mutation were created and analysis revealed alterations to the proteome, largely unrelated to the presence of μORF in epimastigotes. eIF2αT169A mutant parasites produced fewer trypomastigotes with lower infectivity than wild type, with increased levels of sialylated mucins and oligomannose glycoproteins, and decreased galactofuranose epitopes and the surface protease GP63 on the cell surface. We conclude that eIF2α expression and phosphorylation levels affect proteins relevant for intracellular progression of T. cruzi.

• Klíčová slova
• Trypanosoma cruzi, differentiation, eIF2, phosphorylation, translation, virulence,
• MeSH
• buněčné linie MeSH
• Chagasova nemoc parazitologie   MeSH
• CRISPR-Cas systémy MeSH
• eukaryotický iniciační faktor 2 genetika   metabolismus   MeSH
• fosforylace MeSH
• lidé MeSH
• mutace MeSH
• nádorové buněčné linie MeSH
• parazitemie MeSH
• proteom metabolismus   MeSH
• proteosyntéza MeSH
• protozoální proteiny analýza   biosyntéza   metabolismus   MeSH
• regulace genové exprese MeSH
• stadia vývoje MeSH
• Trypanosoma cruzi růst a vývoj   metabolismus   patogenita   MeSH
• virulence MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• eukaryotický iniciační faktor 2 MeSH
• proteom MeSH
• protozoální proteiny MeSH

Although it is known that the Src family tyrosine kinase Lyn initiates Fc epsilon receptor I (Fc epsilon RI) signaling by phosphorylation of the receptor subunits, regulation of Lyn kinase activity and its consequences for receptor signaling are incompletely understood. Using a phospho-Lyn-specific antiserum, we show an increased phosphorylation of the Lyn C-terminal regulatory tyrosine and decreased Lyn kinase activity during Fc epsilon RI-mediated mast cell activation. Mutant Lyn, defective in the C-terminal tyrosine, constitutively phosphorylated several substrates in resting cells, but did not cause Fc epsilon RI internalization or spontaneous degranulation. Fc epsilon RI-induced signaling in the presence of constitutively active Lyn exhibited enhanced phosphorylation of the receptor subunits, Syk, LAT, Gab2, phospholipase C (PLC)gamma 1 and PLC gamma 2, and production of phosphatidylinositol 3,4,5-trisphosphate. Although enzymatic activities of PLC gamma 1 and PLC gamma 2 were also up-regulated, amounts of inositol 1,4,5-trisphosphate, mobilization of intracellular calcium and degranulation were suppressed. Additionally, constitutively active Lyn was strikingly less efficient than wild-type Lyn in restoring the receptor-mediated calcium responses in bone marrow mast cells derived from Lyn(-/-) mice. These findings pinpoint the tight regulation of Lyn kinase activity as a critical event in mast cell degranulation.

• MeSH
• fosforylace MeSH
• kultivované buňky MeSH
• mastocyty imunologie   metabolismus   MeSH
• receptory IgE imunologie   MeSH
• signální transdukce imunologie   MeSH
• skupina kinas odvozených od src-genu imunologie   metabolismus   MeSH
• test degranulace bazofilů MeSH
• tyrosin metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• lyn protein-tyrosine kinase MeSH Prohlížeč
• receptory IgE MeSH
• skupina kinas odvozených od src-genu MeSH
• tyrosin MeSH

RNA splicing, the process of intron removal from pre-mRNA, is essential for the regulation of gene expression. It is controlled by the spliceosome, a megadalton RNA-protein complex that assembles de novo on each pre-mRNA intron through an ordered assembly of intermediate complexes1,2. Spliceosome activation is a major control step that requires substantial protein and RNA rearrangements leading to a catalytically active complex1-5. Splicing factor 3B subunit 1 (SF3B1) protein-a subunit of the U2 small nuclear ribonucleoprotein6-is phosphorylated during spliceosome activation7-10, but the kinase that is responsible has not been identified. Here we show that cyclin-dependent kinase 11 (CDK11) associates with SF3B1 and phosphorylates threonine residues at its N terminus during spliceosome activation. The phosphorylation is important for the association between SF3B1 and U5 and U6 snRNAs in the activated spliceosome, termed the Bact complex, and the phosphorylation can be blocked by OTS964, a potent and selective inhibitor of CDK11. Inhibition of CDK11 prevents spliceosomal transition from the precatalytic complex B to the activated complex Bact and leads to widespread intron retention and accumulation of non-functional spliceosomes on pre-mRNAs and chromatin. We demonstrate a central role of CDK11 in spliceosome assembly and splicing regulation and characterize OTS964 as a highly selective CDK11 inhibitor that suppresses spliceosome activation and splicing.

• MeSH
• aktivace enzymů účinky léků   MeSH
• chinolony farmakologie   MeSH
• chromatin metabolismus   MeSH
• cyklin-dependentní kinasy * antagonisté a inhibitory   metabolismus   MeSH
• fosfoproteiny * chemie   metabolismus   MeSH
• fosforylace MeSH
• malý jaderný ribonukleoprotein U2 * chemie   metabolismus   MeSH
• prekurzory RNA * genetika   metabolismus   MeSH
• sestřih RNA * účinky léků   MeSH
• spliceozomy * účinky léků   metabolismus   MeSH
• threonin metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• chinolony MeSH
• chromatin MeSH
• cyklin-dependentní kinasy * MeSH
• fosfoproteiny * MeSH
• malý jaderný ribonukleoprotein U2 * MeSH
• OTS964 MeSH Prohlížeč
• prekurzory RNA * MeSH
• threonin MeSH

EF-Tu from E. coli, one of the superfamily of GTPase switch proteins, plays a central role in the fast and accurate delivery of aminoacyl-tRNAs to the translating ribosome. An overview is given about the regulatory effects of methylation, phosphorylation and phage-induced cleavage of EF-Tu on its function. During exponential growth, EF-Tu becomes monomethylated at Lys56 which is converted to Me2Lys upon entering the stationary phase. Lys56 is in the GTPase switch-1 region (residues 49-62), a strongly conserved site involved in interactions with the nucleotide and the 5' end of tRNA. Methylation was found to attenuate GTP hydrolysis and may thus enhance translational accuracy. In vivo 5-10% of EF-Tu is phosphorylated at Thr382 by a ribosome-associated kinase. In EF-Tu-GTP, Thr382 in domain 3 has a strategic position in the interface with domain 1; it is hydrogen-bonded to Glu117 that takes part in the switch-2 mechanism, and is close to the T-stem binding site of the tRNA, in a region known for many kirromycin-resistance mutations. Phosphorylation is enhanced by EF-Ts, but inhibited by kirromycin. In reverse, phosphorylated EF-Tu has an increased affinity for EF-Ts, does not bind kirromycin and can no longer bind aminoacyi tRNA. The in vivo role of this reversible modification is still a matter of speculation. T4 infection of E. coli may trigger a phase-exclusion mechanism by activation of Lit, a host-encoded proteinase. As a result, EF-Tu is cleaved site-specifically between Gly59-Ile60 in the switch-1 region. Translation was found to drop beyond a minimum level. Interestingly, the identical sequence in the related EF-G appeared to remain fully intact. Although the Lit cleavage-mechanism may eventually lead to programmed cell death, the very efficient prevention of phage multiplication may be caused by a novel mechanism of in cis inhibition of late T4 mRNA translation.

• MeSH
• elongační faktor Tu metabolismus   MeSH
• Escherichia coli genetika   metabolismus   MeSH
• fosforylace MeSH
• metylace MeSH
• proteosyntéza * MeSH
• regulace genové exprese * MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• elongační faktor Tu MeSH