@article{MTMT:33063331, title = {STABILON, a Novel Sequence Motif That Enhances the Expression and Accumulation of Intracellular and Secreted Proteins}, url = {https://m2.mtmt.hu/api/publication/33063331}, author = {Réthi-Nagy, Zsuzsánna and Ábrahám, Edit and Udvardy, Katalin and Klement, Éva and Darula, Zsuzsanna and Pál, Margit and Katona, Robert L. and Tubak, Vilmos and Páli, Tibor and Kóta, Zoltán and Sinka, Rita and Udvardy, Andor and Lipinszki, Zoltán}, doi = {10.3390/ijms23158168}, journal-iso = {INT J MOL SCI}, journal = {INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES}, volume = {23}, unique-id = {33063331}, issn = {1661-6596}, abstract = {The dynamic balance of transcriptional and translational regulation together with degron-controlled proteolysis shapes the ever-changing cellular proteome. While a large variety of degradation signals has been characterized, our knowledge of cis-acting protein motifs that can in vivo stabilize otherwise short-lived proteins is very limited. We have identified and characterized a conserved 13-mer protein segment derived from the p54/Rpn10 ubiquitin receptor subunit of the Drosophila 26S proteasome, which fulfills all the characteristics of a protein stabilization motif (STABILON). Attachment of STABILON to various intracellular as well as medically relevant secreted model proteins resulted in a significant increase in their cellular or extracellular concentration in mammalian cells. We demonstrate that STABILON acts as a universal and dual function motif that, on the one hand, increases the concentration of the corresponding mRNAs and, on the other hand, prevents the degradation of short-lived fusion proteins. Therefore, STABILON may lead to a breakthrough in biomedical recombinant protein production.}, keywords = {SYSTEM; SUBUNIT; DROSOPHILA; Protein Stability; ENHANCER; proteasome; protein degradation; SECRETED PROTEINS; Biochemistry & Molecular Biology; Protein production; mRNA stability; stabilization motif}, year = {2022}, eissn = {1422-0067}, orcid-numbers = {Páli, Tibor/0000-0003-1649-1097; Kóta, Zoltán/0000-0003-2420-8773; Sinka, Rita/0000-0003-4040-4184; Lipinszki, Zoltán/0000-0002-2067-0832} } @article{MTMT:3376423, title = {Ubiquitylation Dynamics of the Clock Cell Proteome and TIMELESS during a Circadian Cycle}, url = {https://m2.mtmt.hu/api/publication/3376423}, author = {Szabó, Áron and Papin, C and Cornu, D and Chélot, E and Lipinszki, Zoltán and Udvardy, Andor and Redeker, V and Mayor, U and Rouyer, F}, doi = {10.1016/j.celrep.2018.04.064}, journal-iso = {CELL REP}, journal = {CELL REPORTS}, volume = {23}, unique-id = {3376423}, issn = {2211-1247}, year = {2018}, eissn = {2211-1247}, pages = {2273-2282}, orcid-numbers = {Lipinszki, Zoltán/0000-0002-2067-0832} } @article{MTMT:3339588, title = {A 26S PROTEASZÓMA POLIUBIKVITIN RECEPTOR ALEGYSÉGÉNEK KALANDOS AZONOSÍTÁSA}, url = {https://m2.mtmt.hu/api/publication/3339588}, author = {Haracska, Lajos and Lipinszki, Zoltán and Udvardy, Andor}, journal-iso = {BIOKÉMIA}, journal = {BIOKÉMIA: A MAGYAR BIOKÉMIAI EGYESÜLET FOLYÓIRATA}, volume = {41}, unique-id = {3339588}, issn = {0133-8455}, year = {2017}, eissn = {2060-8152}, pages = {64-77}, orcid-numbers = {Lipinszki, Zoltán/0000-0002-2067-0832} } @article{MTMT:2990307, title = {Involvement of small heat shock proteins, trehalose, and lipids in the thermal stress management in Schizosaccharomyces pombe.}, url = {https://m2.mtmt.hu/api/publication/2990307}, author = {Glatz, Attila and Pilbat, Ana Maria and Nemeth, GL and Kontár, Katalin and Jósvay, Katalin and Hunya, Ákos and Udvardy, Andor and Gombos, Imre and Péter, Mária and Balogh, Gábor and Horváth, Ibolya and Vigh, László and Török, Zsolt}, doi = {10.1007/s12192-015-0662-4}, journal-iso = {CELL STRESS CHAPERON}, journal = {CELL STRESS & CHAPERONES}, volume = {21}, unique-id = {2990307}, issn = {1355-8145}, abstract = {Changes in the levels of three structurally and functionally different important thermoprotectant molecules, namely small heat shock proteins (sHsps), trehalose, and lipids, have been investigated upon heat shock in Schizosaccharomyces pombe. Both alpha-crystallin-type sHsps (Hsp15.8 and Hsp16) were induced after prolonged high-temperature treatment but with different kinetic profiles. The shsp null mutants display a weak, but significant, heat sensitivity indicating their importance in the thermal stress management. The heat induction of sHsps is different in wild type and in highly heat-sensitive trehalose-deficient (tps1Delta) cells; however, trehalose level did not show significant alteration in shsp mutants. The altered timing of trehalose accumulation and induction of sHsps suggest that the disaccharide might provide protection at the early stage of the heat stress while elevated amount of sHsps are required at the later phase. The cellular lipid compositions of two different temperature-adapted wild-type S. pombe cells are also altered according to the rule of homeoviscous adaptation, indicating their crucial role in adapting to the environmental temperature changes. Both Hsp15.8 and Hsp16 are able to bind to different lipids isolated from S. pombe, whose interaction might provide a powerful protection against heat-induced damages of the membranes. Our data suggest that all the three investigated thermoprotectant macromolecules play a pivotal role during the thermal stress management in the fission yeast.}, year = {2016}, eissn = {1466-1268}, pages = {327-338}, orcid-numbers = {Hunya, Ákos/0000-0002-4547-9284} } @article{MTMT:2871663, title = {Role of the Deubiquitylating Enzyme DmUsp5 in Coupling Ubiquitin Equilibrium to Development and Apoptosis in Drosophila melanogaster.}, url = {https://m2.mtmt.hu/api/publication/2871663}, author = {Kovács, Levente and Nagy, Olga Mária and Pál, Margit and Udvardy, Andor and Popescu, O and Deák, Péter}, doi = {10.1371/journal.pone.0120875}, journal-iso = {PLOS ONE}, journal = {PLOS ONE}, volume = {10}, unique-id = {2871663}, issn = {1932-6203}, year = {2015}, eissn = {1932-6203}, orcid-numbers = {Kovács, Levente/0000-0002-3226-3740; Nagy, Olga Mária/0000-0003-1471-5165} } @article{MTMT:2731564, title = {Generation of induced pluripotent stem cells by using a mammalian artificial chromosome expression system.}, url = {https://m2.mtmt.hu/api/publication/2731564}, author = {Tóth, Anna and Fodor, Katalin and Blazsó, Péter and Cserpán, Imre and Praznovszky, Tünde and Tubak, Vilmos and Udvardy, Andor and Hadlaczky, Gyula and Katona, Róbert László}, doi = {10.1556/ABiol.65.2014.3.9}, journal-iso = {ACTA BIOL HUNG}, journal = {ACTA BIOLOGICA HUNGARICA (1983-2018)}, volume = {65}, unique-id = {2731564}, issn = {0236-5383}, abstract = {Direct reprogramming of mouse fibroblasts into induced pluripotent stem cells (iPS) was achieved recently by overexpression of four transcription factors encoded by retroviral vectors. Most of the virus vectors, however, may cause insertional mutagenesis in the host genome and may also induce tumor formation. Therefore, it is very important to discover novel and safer, non-viral reprogramming methods. Here we describe the reprogramming of somatic cells into iPS cells by a novel protein-based technique. Engineered Oct4, Sox2 and Klf4 transcription factors carrying an N-terminal Flag-tag and a C-terminal polyarginine tail were synthesized by a recently described mammalian artificial chromosome expression system (ACEs). This system is suitable for the high-level production of recombinant proteins in mammalian tissue culture cells. Recombinant proteins produced in this system contain all the post-translational modifications essential for the stability and the authentic function of the proteins. The engineered Oct4, Sox2 and Klf4 proteins efficiently induced the reprogramming of mouse embryonic fibroblasts by means of protein transduction. This novel method allows for the generation of iPS cells, which may be suitable for therapeutic applications in the future.}, year = {2014}, eissn = {1588-256X}, pages = {331-345}, orcid-numbers = {Blazsó, Péter/0000-0003-4404-8068; Tubak, Vilmos/0000-0002-6141-3920} } @article{MTMT:2508201, title = {Novel Method to Load Multiple Genes onto a Mammalian Artificial Chromosome.}, url = {https://m2.mtmt.hu/api/publication/2508201}, author = {Tóth, Anna and Fodor, Katalin and Praznovszky, Tünde and Tubak, Vilmos and Udvardy, Andor and Hadlaczky, Gyula and Katona, Róbert László}, doi = {10.1371/journal.pone.0085565}, journal-iso = {PLOS ONE}, journal = {PLOS ONE}, volume = {9}, unique-id = {2508201}, issn = {1932-6203}, abstract = {Mammalian artificial chromosomes are natural chromosome-based vectors that may carry a vast amount of genetic material in terms of both size and number. They are reasonably stable and segregate well in both mitosis and meiosis. A platform artificial chromosome expression system (ACEs) was earlier described with multiple loading sites for a modified lambda-integrase enzyme. It has been shown that this ACEs is suitable for high-level industrial protein production and the treatment of a mouse model for a devastating human disorder, Krabbe's disease. ACEs-treated mutant mice carrying a therapeutic gene lived more than four times longer than untreated counterparts. This novel gene therapy method is called combined mammalian artificial chromosome-stem cell therapy. At present, this method suffers from the limitation that a new selection marker gene should be present for each therapeutic gene loaded onto the ACEs. Complex diseases require the cooperative action of several genes for treatment, but only a limited number of selection marker genes are available and there is also a risk of serious side-effects caused by the unwanted expression of these marker genes in mammalian cells, organs and organisms. We describe here a novel method to load multiple genes onto the ACEs by using only two selectable marker genes. These markers may be removed from the ACEs before therapeutic application. This novel technology could revolutionize gene therapeutic applications targeting the treatment of complex disorders and cancers. It could also speed up cell therapy by allowing researchers to engineer a chromosome with a predetermined set of genetic factors to differentiate adult stem cells, embryonic stem cells and induced pluripotent stem (iPS) cells into cell types of therapeutic value. It is also a suitable tool for the investigation of complex biochemical pathways in basic science by producing an ACEs with several genes from a signal transduction pathway of interest.}, year = {2014}, eissn = {1932-6203}, orcid-numbers = {Tubak, Vilmos/0000-0002-6141-3920} } @article{MTMT:2490590, title = {A novel interplay between the ubiquitin-proteasome system and serine proteases during Drosophila development.}, url = {https://m2.mtmt.hu/api/publication/2490590}, author = {Lipinszki, Zoltán and Klement, Éva and Hunyadi-Gulyás Éva, Csilla and Medzihradszky F., Katalin and Márkus, Róbert and Pál, Margit and Deák, Péter and Udvardy, Andor}, doi = {10.1042/BJ20130040}, journal-iso = {BIOCHEM J}, journal = {BIOCHEMICAL JOURNAL}, volume = {454}, unique-id = {2490590}, issn = {0264-6021}, abstract = {The concentrations of the Drosophila proteasomal and extraproteasomal polyubiquitin receptors fluctuate in a developmentally regulated fashion. This fluctuation is generated by a previously unidentified proteolytic activity. In the present paper, we describe the purification, identification and characterization of this protease (endoproteinase I). Its expression increases sharply at the L1-L2 larval stages, remains high until the second half of the L3 stage, then declines dramatically. This sharp decrease coincides precisely with the increase of polyubiquitin receptor concentrations in late L3 larvae, which suggests a tight developmental co-regulation. RNAi-induced down-regulation of endoproteinase I results in pupal lethality. Interestingly, we found a cross-talk between the 26S proteasome and this larval protease: transgenic overexpression of the in vivo target of endoproteinase I, the C-terminal half of the proteasomal polyubiquitin receptor subunit p54/Rpn10 results in transcriptional down-regulation of endoproteinase I and consequently a lower level of proteolytic elimination of the polyubiquitin receptors. Another larval protease, Jonah65A-IV, which degrades only unfolded proteins and exhibits similar cross-talk with the proteasome has also been purified and characterized. It may prevent the accumulation of polyubiquitylated proteins in larvae contrary to the low polyubiquitin receptor concentration.}, keywords = {Animals; PROTEOLYSIS; Amino Acid Motifs; RNA, Small Interfering/genetics; Gene Expression Regulation, Developmental; Conserved Sequence; Enzyme Induction; Drosophila melanogaster/genetics/*growth & development/metabolism; Proteasome Endopeptidase Complex/*metabolism; Unfolded protein response; Gene Knockdown Techniques; *Ubiquitination; Serine Endopeptidases/*chemistry/genetics/*metabolism; Larva/genetics/growth & development/metabolism; Drosophila Proteins/*chemistry/genetics/*metabolism}, year = {2013}, eissn = {1470-8728}, pages = {571-583}, orcid-numbers = {Lipinszki, Zoltán/0000-0002-2067-0832} } @article{MTMT:2015354, title = {lemmingA encodes the Apc11 subunit of the APC/C in Drosophila melanogaster that forms a ternary complex with the E2-C type ubiquitin conjugating enzyme, Vihar and Morula/Apc2}, url = {https://m2.mtmt.hu/api/publication/2015354}, author = {Nagy, Olga Mária and Pál, Margit and Udvardy, Andor and Shirras, CAM and Boros, Imre Miklós and Shirras, AD and Deák, Péter}, doi = {10.1186/1747-1028-7-9}, journal-iso = {CELL DIV}, journal = {CELL DIVISION}, volume = {7}, unique-id = {2015354}, abstract = {Background: Ubiquitin-dependent protein degradation is a critical step in key cell cycle events, such as metaphase-anaphase transition and mitotic exit. The anaphase promoting complex/cyclosome (APC/C) plays a pivotal role in these transitions by recognizing and marking regulatory proteins for proteasomal degradation. Its overall structure and function has been elucidated mostly in yeasts and mammalian cell lines. The APC/C is, however, a multisubunit assembly with at least 13 subunits and their function and interaction within the complex is still relatively uncharacterized, particularly in metazoan systems. Here, lemming (lmg) mutants were used to study the APC/C subunit, Apc11, and its interaction partners in Drosophila melanogaster. Results: The lmg gene was initially identified through a pharate adult lethal P element insertion mutation expressing developmental abnormalities and widespread apoptosis in larval imaginal discs and pupal abdominal histoblasts. Larval neuroblasts were observed to arrest mitosis in a metaphase-like state with highly condensed, scattered chromosomes and frequent polyploidy. These neuroblasts contain high levels of both cyclin A and cyclin B. The lmg gene was cloned by virtue of the lmg(03424) P element insertion which is located in the 5' untranslated region. The lemming locus is transcribed to give a 2.0 kb mRNA that contains two ORFs, lmgA and lmgB. The lmgA ORF codes for a putative protein with more than 80% sequence homology to the APC11 subunit of the human APC/C. The 85 amino acid protein also contains a RING-finger motif characteristic of known APC11 subunits. The lmgA ORF alone was sufficient to rescue the lethal and mitotic phenotypes of the lmg(138) null allele and to complement the temperature sensitive lethal phenotype of the APC11-myc9 budding yeast mutant. The LmgA protein interacts with Mr/Apc2, and they together form a binding site for Vihar, the E2-C type ubiquitin conjugating enzyme. Despite being conserved among Drosophila species, the LmgB protein is not required for viability or fertility. Conclusions: Our work provides insight into the subunit structure of the Drosophila APC/C with implications for its function. Based on the presented data, we suggest that the Lmg/Apc11 subunit recruits the E2-C type ubiquitin conjugating enzyme, Vihar, to the APC/C together with Mr/Apc2 by forming a ternary complex.}, year = {2012}, eissn = {1747-1028}, orcid-numbers = {Nagy, Olga Mária/0000-0003-1471-5165; Boros, Imre Miklós/0000-0001-8504-9687} } @article{MTMT:2015348, title = {Ubiquitylation of Drosophila p54/Rpn10/S5a Regulates Its Interaction with the UBA-UBL Polyubiquitin Receptors}, url = {https://m2.mtmt.hu/api/publication/2015348}, author = {Lipinszki, Zoltán and Kovács, Levente and Deák, Péter and Udvardy, Andor}, doi = {10.1021/bi3001006}, journal-iso = {BIOCHEMISTRY-US}, journal = {BIOCHEMISTRY}, volume = {51}, unique-id = {2015348}, issn = {0006-2960}, abstract = {Analysis of the in vivo ubiquitylation of the p54/Rpn10 polyubiquitin receptor subunit of the Drosophila 26S proteasome revealed that the site of ubiquitylation is the C-terminal cluster of lysines, which is conserved in higher eukaryotes. Extraproteasomal p54 was extensively multiubiquitylated, but only very modest modification was detected in the proteasome-assembled subunit. Ubiquitylation of p54 seriously jeopardizes one of its most important functions, i.e., the interaction of its ubiquitin-interacting motifs with the ubiquitin-like domain of Dsk2 and Rad23 extraproteasomal polyubiquitin receptors. This modification of p54 supports the previous notion that p54 is a shuttling subunit of the 26S proteasome with a specific extraproteasomal function. This assumption is supported by the observation that, while transgenic p54 can fully rescue the lethal phenotype of the Delta p54 null mutation, its derivative from which the cluster of conserved lysines is deleted shifts the lethality from the early pupa to pharate adult stage but cannot rescue the Delta p54 mutation, suggesting that ubiquitylated extraproteasomal p54 has an essential role in the pupa adult transition.}, year = {2012}, eissn = {1520-4995}, pages = {2461-2470}, orcid-numbers = {Lipinszki, Zoltán/0000-0002-2067-0832; Kovács, Levente/0000-0002-3226-3740} }