EGF regulates tyrosine phosphorylation and membrane-translocation of the scaffold protein Tks5

Background: Tks5/FISH is a scaffold protein comprising of five SH3 domains and one PX domain. Tks5 is a substrate of the tyrosine kinase Src and is required for the organization of podosomes/invadopodia implicated in invasion of tumor cells. Recent data have suggested that a close homologue of Tks5, Tks4, is implicated in the EGF signaling.Results: Here, we report that Tks5 is a component of the EGF signaling pathway. In EGF-treated cells, Tks5 is tyrosine phosphorylated within minutes and the level of phosphorylation is sustained for at least 2 hours. Using specific kinase inhibitors, we demonstrate that tyrosine phosphorylation of Tks5 is catalyzed by Src tyrosine kinase. We show that treatment of cells with EGF results in plasma membrane translocation of Tks5. In addition, treatment of cells with LY294002, an inhibitor of PI 3-kinase, or mutation of the PX domain reduces tyrosine phosphorylation and membrane translocation of Tks5.Conclusions: Our results identify Tks5 as a novel component of the EGF signaling pathway. © 2013 Fekete et al.; licensee BioMed Central Ltd

Complex formation of EphB1/Nck/Caskin1 leads to tyrosine phosphorylation and structural changes of the Caskin1 SH3 domain.

Scaffold proteins have an important role in the regulation of signal propagation. These proteins do not possess any enzymatic activity but can contribute to the formation of multiprotein complexes. Although scaffold proteins are present in all cell types, the nervous system contains them in the largest amount. Caskin proteins are typically present in neuronal cells, particularly, in the synapses. However, the signaling mechanisms by which Caskin proteins are regulated are largely unknown. Here we demonstrate that EphB1 receptor tyrosine kinase can recruit Caskin1 through the adaptor protein Nck. Upon activation of the receptor kinase, the SH2 domain of Nck binds to one of its tyrosine residues, while Nck SH3 domains interact with the proline-rich domain of Caskin1. Complex formation of the receptor, adaptor and scaffold proteins results in the tyrosine phosphorylation of Caskin1 on its SH3 domain. The phosphorylation sites were identified by mass-spectrometry as tyrosines 296 and 336. To reveal the structural consequence of this phosphorylation, CD spectroscopy was performed. This measurement suggests that upon tyrosine phosphorylation the structure of the Caskin1 SH3 domain changes significantly. Taken together, we propose that the scaffold protein Caskin1 can form a complex with the EphB1 tyrosine kinase via the Nck protein as a linker. Complex formation results in tyrosine phosphorylation of the Caskin1 SH3 domain. Although we were not able to identify any physiological partner of the SH3 domain so far, we could demonstrate that phosphorylation on conserved tyrosine residues results in marked changes in the structure of the SH3 domain

Determination of chlorine species by capillary electrophoresis: mass spectrometry

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Asztmás betegek életminőségének változása komplex rehabilitációs kezelés után = Quality of life of asthmatic patients after complex rehabilitation treatmen

Absztrakt: Az asztma valamennyi korosztály számára világszerte súlyos népegészségügyi kihívást jelent. A természetes gyógytényezők használata a betegségek gyógyításában fokozott figyelmet kap hazai és nemzetközi viszonylatban is, ami a gyógyszeres kezeléseket költséghatékonyabbá teheti. A kutatásunk újszerűsége, hogy igazoljuk: már a 700–1000 méter tengerszint feletti magasságú gyógyhelyen is érvényesül a klíma gyógyhatása. Kutatásunk során arra kerestük a választ, hogyan alakul az asztmások életminősége a klímaterápiával kiegészített orvosi kezelés után, s igazolható-e a 700–1000 méteren végzett klímaterápia hatása idős, felnőtt betegeken. A légzőszervi megbetegedésben szenvedők a Mátrai Gyógyintézetben 3 hetes komplex terápián vettek részt. A jelen cikkben 514 asztmás beteg adatait elemeztük. Az orvosi vizsgálatokon és kezeléseken túl a betegek napi két alkalommal kúrateraszokon vagy a szabadban végzett légzőtornán, valamint napi egy alkalommal gyógytornászok által vezetett gyógytorna-mozgásprogramban, dietetikai és életmód-tanácsadáson vettek részt, továbbá a betegségükhöz igazodó diétás étrend lehetőségét is biztosítottuk számukra. A légzésfunkció vizsgálatát az intézetbe jövetelkor a kúra előtt és az intézetből történő távozáskor, a kúra után végeztük el. A betegek terhelhetőségét a 6 perces járásteszttel mértük fel a kúra előtt, majd 3 héttel később. A betegek részére kérdőívet készítettünk, melyben az intézetből való távozás utáni állapotukra (1–6 hónap) kérdeztünk rá. Eredményeink azt mutatják, hogy az asztmás betegek légzésfunkciós értékei, a 6 perces járásteszt eredményei szignifikánsan javultak a kezelés hatására. Az intézetből való távozás után az Asthma Control Test eredményei a kezelés utáni 1. hónapban a betegek szignifikánsan jobb életminőségét és jobb kontrollszintjét mutatták, mint a 3. hónap elteltével. Orv Hetil. 2018; 159(27): 1103–1112. | Abstract: Asthma is a serious public health challenge for all age groups around the world. Heightened attention surrounds the use of the natural therapeutic factors that potentially make medication more cost-efficient in the treatment of diseases. Our research has tried to reveal how the medical conditions, quality of life of asthmatic patients change after medical treatments that are accompanied by climate therapy. Asthma patients (514 patients) participated in a 3-week complex therapy at Mátra Health Resort. Beyond medical examinations and treatments, patients were involved in breathing exercises on the curing terraces or outdoors twice a day, they were to attend the rehabilitative physical exercises led by a physiotherapist once a day, were given dietetic and lifestyle advice and provided with diets that were fitted to their individual medical conditions. Their respiratory function was examined before the commencement of the cure as well as after the therapy just before departing from the institution. The patients’ exercise capacities were assessed with the 6-minute walk test before the treatment and then 3 weeks later. For the patients, a questionnaire was compiled to reveal information in relation to their conditions in the first 6 months after leaving the institution. Our results show that values of respiratory function in the sample and 6-minute walk test significantly improved with the treatment. After leaving the institute, the results of the Asthma Control Test in the 1st month after treatment revealed a significantly better quality of life and better control-level of patients than after the 3rd month. Orv Hetil. 2018; 159(27): 1103–1112

Bipolar membrane electrodialysis integration into the biotechnological production of itaconic acid: a proof-of-concept study

Itaconic acid is a promising biobased organic acid that can be industrially produced in an eco-friendly way by aerobic fungal fermentation. It has many applications, e.g. in biopolymers, and has the potential to help replace or facilitate the green production of other similar but fossil-based chemicals. Nowadays, its production costs are still relatively high, partly due to the multistep product recovery process from the fermentation broth. In this study, a reduced number of downstream processes were evaluated to recover itaconic acid from model and real fermentation effluents. To the best of our knowledge, this is the first time the whole procedure is presented from the substrates to the solid and pure product, including fermentation, electrodialysis with bipolar membrane, evaporation and optimized crystallization. Based on the fermentation temperature and elevated pH, a synergistic effect was observed that intensified certain mass transfer mechanisms. Consequently, a significant amount of water (400 cm3 ) and itaconic acid (15 g) was transported into the alkaline concentrate. Moreover, novel outcomes about the condition, decolorization and process enhancements of the membrane (product recovery: 65–80 %, current efficiency: 42–76 %, product purity: 99 %) were recorded. Based on the results, our group is one step closer to the final concept of a continuously operated fermentation-electrodialysis integrated system

Accumulation of the PX domain mutant Frank-ter Haar syndrome protein Tks4 in aggresomes

BACKGROUND: Cells deploy quality control mechanisms to remove damaged or misfolded proteins. Recently, we have reported that a mutation (R43W) in the Frank-ter Haar syndrome protein Tks4 resulted in aberrant intracellular localization. RESULTS: Here we demonstrate that the accumulation of Tks4(R43W) depends on the intact microtubule network. Detergent-insoluble Tks4 mutant colocalizes with the centrosome and its aggregate is encaged by the intermediate filament protein vimentin. Both the microtubule inhibitor nocodazole and the histone deacetylase inhibitor Trichostatin A inhibit markedly the aggresome formation in cells expressing Tks4(R43W). Finally, pretreatment of cells with the proteasome inhibitor MG132 markedly increases the level of aggresomes formed by Tks4(R43W). Furthermore, two additional mutant Tks4 proteins (Tks4(1-48) or Tks4(1-341)) have been investigated. Whereas the shorter Tks4 mutant, Tks4(1-48), shows no expression at all, the longer Tks4 truncation mutant accumulates in the nuclei of the cells. CONCLUSIONS: Our results suggest that misfolded Frank-ter Haar syndrome protein Tks4(R43W) is transported via the microtubule system to the aggresomes. Lack of expression of Tks4(1-48) or aberrant intracellular expressions of Tks4(R43W) and Tks4(1-341) strongly suggest that these mutations result in dysfunctional proteins which are not capable of operating properly, leading to the development of FTHS

Klórvegyületek meghatározása kapilláris elektroforézissel kapcsolt tömegspektrométerrel

A kutatómunkánk során különböző klórvegyületek egymás melletti meghatározását végeztük nátriumsóik vizes oldatából. A meghatározás szempontjából jelentős ionok a klorid-, hipoklorit-, klorit-, klorát- és perklorátionok voltak. A kutató munkánk során a célunk egy egyszerű, megbízható és reprodukálható CE-MS módszer kifejlesztése volt a fent felsorolt klórvegyületek egymás melletti kimutatására. Az optimálási folyamat részeként elsőként CE készüléken vizsgáltuk az áramerősség alakulását a kapilláris végeinél elhelyezett különböző elektrolitok esetében. Miután kiválasztottuk az ígéretesnek adódó elektrolitpárokat a CE-MS készüléken folytattuk tovább a kísérleteinket, optimáltuk a háttérelektrolit és a pótlólagos folyadékáram összetételét, majd vizsgáltuk a kapilláris bemeneti végénél alkalmazott plusznyomás hatását az elválasztásra. Az optimális paraméterek meghatározása után az analitikai teljesítőképességet és reprodukálhatóságot vizsgáltuk, majd valós mintákon végeztünk méréseket, hogy az általunk kidolgozott módszer jóságát teszteljük és hatékonyságát bizonyítsuk.BSc/BAVegyészmérnökg

RESEARCH Open Access Complex formation of EphB1/Nck/Caskin1 leads to tyrosine phosphorylation and structural

Background: Scaffold proteins have an important role in the regulation of signal propagation. These proteins do not possess any enzymatic activity but can contribute to the formation of multiprotein complexes. Although scaffold proteins are present in all cell types, the nervous system contains them in the largest amount. Caskin proteins are typically present in neuronal cells, particularly, in the synapses. However, the signaling mechanisms by which Caskin proteins are regulated are largely unknown. Results: Here we demonstrate that EphB1 receptor tyrosine kinase can recruit Caskin1 through the adaptor protein Nck. Upon activation of the receptor kinase, the SH2 domain of Nck binds to one of its tyrosine residues, while Nck SH3 domains interact with the proline-rich domain of Caskin1. Complex formation of the receptor, adaptor and scaffold proteins results in the tyrosine phosphorylation of Caskin1 on its SH3 domain. The phosphorylation sites were identified by mass-spectrometry as tyrosines 296 and 336. To reveal the structural consequence of this phosphorylation, CD spectroscopy was performed. This measurement suggests that upon tyrosine phosphorylation the structure of the Caskin1 SH3 domain changes significantly. Conclusion: Taken together, we propose that the scaffold protein Caskin1 can form a complex with the EphB1 tyrosine kinase via the Nck protein as a linker. Complex formation results in tyrosine phosphorylation of the Caskin1 SH3 domain. Although we were not able to identify any physiological partner of the SH3 domain so far, we could demonstrate that phosphorylation on conserved tyrosine residues results in marked changes in the structure of the SH3 domain