The close relationship between the Golgi trafficking machinery and protein glycosylation

La glicosilazione è la più comune modifica post-traduzionale delle proteine; media la loro corretta piegatura e stabilità, nonché il loro trasporto attraverso il trasporto secretorio. I cambiamenti nei glicani legati all'N e all'O sono stati associati a molteplici condizioni patologiche tra cui disturbi congeniti della glicosilazione, malattie infiammatorie e cancro. La glicosilazione della glicoproteina al Golgi coinvolge l'azione coordinata di centinaia di glicosiltransferasi e glicosidasi, che vengono mantenute nella posizione corretta attraverso il traffico di vescicole retrograde tra le cisterne di Golgi. In questa recensione, descriviamo il macchinario molecolare coinvolto nel traffico di vescicole e nel tethering presso l'apparato di Golgi e gli effetti delle mutazioni nel contesto della biosintesi dei glicani e delle malattie umane.Glycosylation is the most common post-translational modification of proteins; it mediates their correct folding and stability, as well as their transport through the secretory transport. Changes in N- and O-linked glycans have been associated with multiple pathological conditions including congenital disorders of glycosylation, inflammatory diseases and cancer. Glycoprotein glycosylation at the Golgi involves the coordinated action of hundreds of glycosyltransferases and glycosidases, which are maintained at the correct location through retrograde vesicle trafficking between Golgi cisternae. In this review, we describe the molecular machinery involved in vesicle trafficking and tethering at the Golgi apparatus and the effects of mutations in the context of glycan biosynthesis and human diseases

Oncogenic roles of GOLPH3 in the physiopathology of cancer

Golgi phosphoprotein 3 (GOLPH3), un effettore del fosfatidilinositolo 4-fosfato [PI (4) P] al Golgi, è necessaria per il mantenimento della struttura del nastro del Golgi, il traffico di vescicole e la glicosilazione del Golgi. GOLPH3 è stato convalidato come oncoproteina combinando la genomica integrativa con l'analisi clinopatologiche e funzionali. È spesso amplificato in diversi tipi di tumori solidi tra cui melanoma, cancro ai polmoni, cancro al seno, glioma e cancro del colon-retto. La sovraespressione di GOLPH3 è correlata a una prognosi infausta in più tipi di tumore, compreso il 52% dei tumori al seno e dal 41% al 53% del glioblastoma. I ruoli di GOLPH3 nella tumorigenesi possono essere correlati a diverse attività cellulari, tra cui: (i) regolazione del traffico dal Golgi alla membrana plasmatica e contributo a fenotipi secretori maligni; (ii) controllare l'internalizzazione e il riciclaggio di molecole di segnalazione chiave o aumentare la glicosilazione delle glicoproteine ​​rilevanti per il cancro; e (iii) influenzare la risposta al danno al DNA e il mantenimento della stabilità genomica. Qui riassumiamo le attuali conoscenze sui percorsi oncogeni che coinvolgono GOLPH3 nel cancro umano, l'influenza di GOLPH3 sul metabolismo del tumore e sullo stroma circostante e il suo possibile ruolo nella formazione di metastasi tumorali.Golgi phosphoprotein 3 (GOLPH3), a Phosphatidylinositol 4-Phosphate [PI(4)P] effector at the Golgi, is required for Golgi ribbon structure maintenance, vesicle trafficking and Golgi glycosylation. GOLPH3 has been validated as an oncoprotein through combining integrative genomics with clinopathological and functional analyses. It is frequently amplified in several solid tumor types including melanoma, lung cancer, breast cancer, glioma, and colorectal cancer. Overexpression of GOLPH3 correlates with poor prognosis in multiple tumor types including 52% of breast cancers and 41% to 53% of glioblastoma. Roles of GOLPH3 in tumorigenesis may correlate with several cellular activities including: (i) regulating Golgi-to-plasma membrane trafficking and contributing to malignant secretory phenotypes; (ii) controlling the internalization and recycling of key signaling molecules or increasing the glycosylation of cancer relevant glycoproteins; and (iii) influencing the DNA damage response and maintenance of genomic stability. Here we summarize current knowledge on the oncogenic pathways involving GOLPH3 in human cancer, GOLPH3 influence on tumor metabolism and surrounding stroma, and its possible role in tumor metastasis formation

COG7 deficiency in Drosophila generates multifaceted developmental, behavioral and protein glycosylation phenotypes

Congenital disorders of glycosylation (CDG) comprise a family of human multisystemic diseases caused by recessive mutations in genes required for protein N-glycosylation. More than 100 distinct forms of CDGs have been identified and most of them cause severe neurological impairment. The Conserved Oligomeric Golgi (COG) complexmediates tethering of vesicles carrying glycosylation enzymes across the Golgi cisternae. Mutations affecting human COG1, COG2 and COG4-COG8 cause monogenic forms of inherited, autosomal recessive CDGs.We have generated a Drosophila COG7-CDG model that closely parallels the pathological characteristics of COG7-CDG patients, including pronounced neuromotor defects associated with altered N-glycome profiles. Consistent with these alterations, larval neuromuscular junctions of Cog7 mutants exhibit a significant reduction in bouton numbers. We demonstrate that the COG complex cooperates with Rab1 and Golgi phosphoprotein 3 to regulate Golgi trafficking and that overexpression of Rab1 can rescue the cytokinesis and locomotor defects associated with loss of Cog7. Our results suggest that the Drosophila COG7-CDG model can be used to test novel potential therapeutic strategies by modulating trafficking pathways

A novel coordinated function of myosin II with GOLPH3 controls centralspindlin localization during cytokinesis in drosophila

Nella citochinesi delle cellule animali, l'interazione della miosina II non muscolare (NMII) con l'actina F fornisce la forza dominante per dividere la cellula madre in due figlie. Qui dimostriamo che celibe (cbe) è un allele missenso di zipper, che codifica per la catena pesante della miosina in Drosophila. La mutazione di cbe altera il legame della proteina Zipper con la catena leggera di spaghetti squash (Sqh). Negli spermatociti in divisione dai maschi cbe, Sqh non riesce a concentrarsi nella corteccia equatoriale, determinando anelli sottili di actomiosina che non sono in grado di restringersi. Mostriamo che la mutazione cbe altera la localizzazione di Golgi phosphoprotein 3 (GOLPH3, anche conosciuta come Sauron) che lega il fosfatidilinositolo 4-fosfato [PI (4) P] e il mantenimento della centralspindlina al cell equator durante la telofase. I nostri risultati dimostrano ulteriormente che la proteina GOLPH3 si associa a Sqh e si lega direttamente alla subunità centralspindlin Pavarotti. Proponiamo che durante la citochinesi, la dipendenza reciproca tra miosina e PI (4) P-GOLPH3 regola la stabilizzazione centralspindlin presso la membrana plasmatica invaginante e l'assemblaggio dell'anello contrattile.In animal cell cytokinesis, interaction of non-muscle myosin II (NMII) with F-actin provides the dominant force for pinching the mother cell into two daughters. Here we demonstrate that celibe (cbe) is a missense allele of zipper, which encodes the Drosophila Myosin heavy chain. Mutation of cbe impairs binding of Zipper protein to the regulatory light chain Spaghetti squash (Sqh). In dividing spermatocytes from cbe males, Sqh fails to concentrate at the equatorial cortex, resulting in thin actomyosin rings that are unable to constrict. We show that cbe mutation impairs localization of the phosphatidylinositol 4-phosphate [PI(4)P]-binding protein Golgi phosphoprotein 3 (GOLPH3, also known as Sauron) and maintenance of centralspindlin at the cell equator of telophase cells. Our results further demonstrate that GOLPH3 protein associates with Sqh and directly binds the centralspindlin subunit Pavarotti. We propose that during cytokinesis, the reciprocal dependence between Myosin and PI(4)P-GOLPH3 regulates centralspindlin stabilization at the invaginating plasma membrane and contractile ring assembly

Modeling congenital disorders of N-linked glycoprotein glycosylation in Drosophila melanogaster

La glicosilazione delle proteine, l'aggiunta enzimatica di glicani N-legati o O-legati alle proteine, svolge funzioni cruciali nelle cellule animali e richiede l'azione di glicosiltransferasi, glicosidasi e trasportatori nucleotidi-zucchero, localizzati nel reticolo endoplasmatico e nell'apparato di Golgi. I disturbi congeniti della glicosilazione (CDG) comprendono una famiglia di malattie multisistemiche causate da mutazioni nei geni che codificano per proteine ​​coinvolte nelle vie di glicosilazione. Le CDG sono classificati in due grandi gruppi. le CDG di tipo I influenzano la sintesi del precursore Glc3Man9GlcNac2 legato al dolicolo della glicosilazione legata all'N o il suo trasferimento alle proteine ​​accettrici. Le malattie CDG di tipo II (CDG-II) compromettono o il taglio dell'oligosaccaride legato all'N, l'aggiunta di glicani terminali o la biosintesi degli oligosaccaridi legati all'O, che si verificano nell'apparato di Golgi. Finora, sono note oltre 100 forme distinte di CDG, la maggior parte delle quali caratterizzata da difetti neurologici tra cui ritardo mentale, convulsioni e ipotonia. Tuttavia, non è chiaro in che modo la glicosilazione difettosa causi la patologia delle CDG. Questo problema può essere affrontato solo sviluppando modelli animali di CDG specifici. Drosophila melanogaster sta emergendo come un organismo altamente adatto per analizzare le funzioni dipendenti dai glicani nel sistema nervoso centrale (SNC) e il coinvolgimento della N-glicosilazione nelle neuropatologie. In questa recensione illustriamo un lavoro recente che evidenzia i vantaggi genetici e neurobiologici offerti da D. melanogaster per la dissezione delle vie di glicosilazione e la modellazione della fisiopatologia del CDG.Protein glycosylation, the enzymatic addition of N-linked or O-linked glycans to proteins, serves crucial functions in animal cells and requires the action of glycosyltransferases, glycosidases and nucleotide-sugar transporters, localized in the endoplasmic reticulum and Golgi apparatus. Congenital Disorders of Glycosylation (CDGs) comprise a family of multisystemic diseases caused by mutations in genes encoding proteins involved in glycosylation pathways. CDGs are classified into two large groups. Type I CDGs affect the synthesis of the dolichol-linked Glc3Man9GlcNac2 precursor of N-linked glycosylation or its transfer to acceptor proteins. Type II CDG (CDG-II) diseases impair either the trimming of the N-linked oligosaccharide, the addition of terminal glycans or the biosynthesis of O-linked oligosaccharides, which occur in the Golgi apparatus. So far, over 100 distinct forms of CDGs are known, with the majority of them characterized by neurological defects including mental retardation, seizures and hypotonia. Yet, it is unclear how defective glycosylation causes the pathology of CDGs. This issue can be only addressed by developing animal models of specific CDGs. Drosophila melanogaster is emerging as a highly suitable organism for analyzing glycan-dependent functions in the central nervous system (CNS) and the involvement of N-glycosylation in neuropathologies. In this review we illustrate recent work that highlights the genetic and neurobiologic advantages offered by D. melanogaster for dissecting glycosylation pathways and modeling CDG pathophysiology

Rab1 interacts with GOLPH3 and controls Golgi structure and contractile ring constriction during cytokinesis in Drosophila melanogaster

Cytokinesis requires a tight coordination between actomyosin ring constriction and new membrane addition along the ingressing cleavage furrow. However, the molecular mechanisms underlying vesicle trafficking to the equatorial site and how this process is coupled with the dynamics of the contractile apparatus are poorly defined. Here we provide evidence for the requirement of Rab1 during cleavage furrow ingression in cytokinesis. We demonstrate that the gene omelette (omt) encodes the Drosophila orthologue of human Rab1 and is required for successful cytokinesis in both mitotic and meiotic dividing cells of Drosophila melanogaster. We show that Rab1 protein colocalizes with the conserved oligomeric Golgi (COG) complex Cog7 subunit and the phosphatidylinositol 4-phosphate effector GOLPH3 at the Golgi stacks. Analysis by transmission electron microscopy and 3D-SIM super-resolution microscopy reveals loss of normal Golgi architecture in omt mutant spermatocytes indicating a role for Rab1 in Golgi formation. In dividing cells, Rab1 enables stabilization and contraction of actomyosin rings. We further demonstrate that GTP-bound Rab1 directly interacts with GOLPH3 and controls its localization at the Golgi and at the cleavage site. We propose that Rab1, by associating with GOLPH3, controls membrane trafficking and contractile ring constriction during cytokinesis

Exocyst-Dependent Membrane Addition Is Required for Anaphase Cell Elongation and Cytokinesis in <i>Drosophila</i>

<div><p>Mitotic and cytokinetic processes harness cell machinery to drive chromosomal segregation and the physical separation of dividing cells. Here, we investigate the functional requirements for exocyst complex function during cell division <i>in vivo</i>, and demonstrate a common mechanism that directs anaphase cell elongation and cleavage furrow progression during cell division. We show that <i>onion rings (onr)</i> and <i>funnel cakes (fun)</i> encode the <i>Drosophila</i> homologs of the Exo84 and Sec8 exocyst subunits, respectively. In <i>onr</i> and <i>fun</i> mutant cells, contractile ring proteins are recruited to the equatorial region of dividing spermatocytes. However, cytokinesis is disrupted early in furrow ingression, leading to cytokinesis failure. We use high temporal and spatial resolution confocal imaging with automated computational analysis to quantitatively compare wild-type versus <i>onr</i> and <i>fun</i> mutant cells. These results demonstrate that anaphase cell elongation is grossly disrupted in cells that are compromised in exocyst complex function. Additionally, we observe that the increase in cell surface area in wild type peaks a few minutes into cytokinesis, and that <i>onr</i> and <i>fun</i> mutant cells have a greatly reduced rate of surface area growth specifically during cell division. Analysis by transmission electron microscopy reveals a massive build-up of cytoplasmic astral membrane and loss of normal Golgi architecture in <i>onr</i> and <i>fun</i> spermatocytes, suggesting that exocyst complex is required for proper vesicular trafficking through these compartments. Moreover, recruitment of the small GTPase Rab11 and the PITP Giotto to the cleavage site depends on wild-type function of the exocyst subunits Exo84 and Sec8. Finally, we show that the exocyst subunit Sec5 coimmunoprecipitates with Rab11. Our results are consistent with the exocyst complex mediating an essential, coordinated increase in cell surface area that potentiates anaphase cell elongation and cleavage furrow ingression.</p></div

Vertical knowledge transfer in Czech organizations

For organizations the losing key employees is the greatest threat; if the organizations lose the key workers, they cannot use their premises, data, information and other resources. The article aims at evaluation of the level of vertical knowledge transfer in organizations in the Czech Republic. The partial goals of this article are to determine dependencies between the examined qualitative features. The article has been drawn up using scientific methods, in particular logical methods, such as analysis, synthesis, induction, deduction and comparison. The results were obtained from longitudinal quantitative research in organizations in the Czech Republic from 2010 to 2014. The article identifies the factors affecting vertical knowledge transfer and presents a method of eliminating the risk of losing key knowledge workers. Vertical knowledge transfer is a new supporting factor of organizations’ productivity and business continuity

Įmonių pasirengimo įgyvendinti inovacijų strategiją vertinimo sistema

Development and implementation of innovation strategy requires increased attention of the businesses. Rather than the business acceding to its implementation needs to know the current status of work in innovation and the key elements that will be crucial in this process. The aim of this paper is based on the analysis of literary sources and carried out research on the proposal of the evaluation system of preparedness of businesses for implementation of an innovation strategy. The proposal describes different levels of preparedness, the basic evaluation methodology and evaluation procedure. The paper brings the main results of the authors who conducted research on a sample of 462 respondents to show the current situation in the Slovak businesses in the use of innovation strategy. A survey used the following methods: comparative method, qualitative evaluation, and method of structured and semi-structured interviews, observation methods, method of document analysis (a method of content analysis) and the questionnaire method