Anterior gradient 2 (AGR2) is an endoplasmic reticulum (ER)-resident protein disulfide isomerase (PDI) known to be overexpressed in many human epithelial cancers and is involved in cell migration, cellular transformation, angiogenesis, and metastasis. This protein inhibits the activity of the tumor suppressor p53, and its expression levels can be used to predict cancer patient outcome. However, the precise network of AGR2-interacting partners and clients remains to be fully characterized. Herein, we used label-free quantification and also stable isotope labeling with amino acids in cell culture-based LC-MS/MS analyses to identify proteins interacting with AGR2. Functional annotation confirmed that AGR2 and its interaction partners are associated with processes in the ER that maintain intracellular metabolic homeostasis and participate in the unfolded protein response, including those associated with changes in cellular metabolism, energy, and redox states in response to ER stress. As a proof of concept, the interaction between AGR2 and PDIA3, another ER-resident PDI, was studied in more detail. Pathway analysis revealed that AGR2 and PDIA3 play roles in protein folding in ER, including post-translational modification and in cellular response to stress. We confirmed the AGR2-PDIA3 complex formation in cancer cells, which was enhanced in response to ER stress. Accordingly, molecular docking characterized potential quaternary structure of this complex; however, it remains to be elucidated whether AGR2 rather contributes to PDIA3 maturation in ER, the complex directly acts in cellular signaling, or mediates AGR2 secretion. Our study provides a comprehensive insight into the protein-protein interaction network of AGR2 by identifying functionally relevant proteins and related cellular and biochemical pathways associated with the role of AGR2 in cancer cells.
- MeSH
- chromatografie kapalinová MeSH
- lidé MeSH
- mapy interakcí proteinů MeSH
- mukoproteiny * metabolismus MeSH
- nádory * MeSH
- onkogenní proteiny * metabolismus MeSH
- proteindisulfidisomerasy * MeSH
- simulace molekulového dockingu MeSH
- tandemová hmotnostní spektrometrie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Anterior gradient 2 (AGR2) is a dimeric protein disulfide isomerase family member involved in the regulation of protein quality control in the endoplasmic reticulum (ER). Mouse AGR2 deletion increases intestinal inflammation and promotes the development of inflammatory bowel disease (IBD). Although these biological effects are well established, the underlying molecular mechanisms of AGR2 function toward inflammation remain poorly defined. Here, using a protein-protein interaction screen to identify cellular regulators of AGR2 dimerization, we unveiled specific enhancers, including TMED2, and inhibitors of AGR2 dimerization, that control AGR2 functions. We demonstrate that modulation of AGR2 dimer formation, whether enhancing or inhibiting the process, yields pro-inflammatory phenotypes, through either autophagy-dependent processes or secretion of AGR2, respectively. We also demonstrate that in IBD and specifically in Crohn's disease, the levels of AGR2 dimerization modulators are selectively deregulated, and this correlates with severity of disease. Our study demonstrates that AGR2 dimers act as sensors of ER homeostasis which are disrupted upon ER stress and promote the secretion of AGR2 monomers. The latter might represent systemic alarm signals for pro-inflammatory responses.
- MeSH
- endoplazmatické retikulum genetika metabolismus MeSH
- HEK293 buňky MeSH
- homeostáze proteinů * MeSH
- lidé MeSH
- mukoproteiny genetika metabolismus MeSH
- multimerizace proteinu * MeSH
- myši MeSH
- onkogenní proteiny genetika metabolismus MeSH
- stres endoplazmatického retikula * MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Banana Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc) is soil-borne disease of banana (Musa spp.) causing significant economic losses. Extensins and arabinogalactan proteins (AGPs) are cell wall components important for pathogen defence. Their significance for Foc resistance in banana was not reported so far. In this study, two banana cultivars differing in Foc sensitivity were used to monitor the changes in transcript levels, abundance and distribution of extensins and AGPs after wounding and Foc inoculation. Extensins mainly appeared in the root cap and meristematic cells. AGPs recognized by JIM13, JIM8, PN16.4B4 and CCRC-M134 antibodies located in root hairs, xylem and root cap. Individual AGPs and extensins showed specific radial distribution in banana roots. At the transcript level, seven extensins and 23 AGPs were differentially expressed between two banana cultivars before and after treatments. Two extensins and five AGPs responded to the treatments at the protein level. Most extensins and AGPs were up-regulated by wounding and pathogen inoculation of intact plants but down-regulated by pathogen attack of wounded plants. Main components responsible for the resistance of banana were MaELP-2 and MaPELP-2. Our data revealed that AGPs and extensins represent dynamic cell wall components involved in wounding and Foc resistance.
- MeSH
- banánovník mikrobiologie fyziologie MeSH
- epitopy imunologie MeSH
- Fusarium * MeSH
- glykoproteiny genetika metabolismus MeSH
- interakce hostitele a patogenu genetika MeSH
- kořeny rostlin genetika metabolismus mikrobiologie MeSH
- mukoproteiny genetika metabolismus MeSH
- nemoci rostlin genetika mikrobiologie MeSH
- odolnost vůči nemocem genetika MeSH
- regulace genové exprese u rostlin * MeSH
- rostlinné proteiny genetika imunologie metabolismus MeSH
- transport proteinů MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
Uromodulin (UMOD) malfunction has been found in a range of autosomal dominant tubulointerstitial nephropathies associated with hyperuricaemia, gouty arthritis, medullary cysts and renal failure-labelled as familial juvenile hyperuricaemic nephropathy, medullary cystic disease type 2 and glomerulocystic kidney disease. To gain knowledge of the spectrum of UMOD changes in various genetic diseases with renal involvement we examined urinary UMOD excretion and found significant quantitative and qualitative changes in 15 male patients at various clinical stages of Fabry disease. In untreated patients, the changes ranged from normal to a marked decrease, or even absence of urinary UMOD. This was accompanied frequently by the presence of aberrantly processed UMOD lacking the C-terminal part following the K432 residue. The abnormal patterns normalized in all patients on enzyme replacement therapy and in some patients on substrate reduction therapy. Immunohistochemical analysis of the affected kidney revealed abnormal UMOD localization in the thick ascending limb of Henle's loop and the distal convoluted tubule, with UMOD expression inversely proportional to the degree of storage. Our observations warrant evaluation of tubular functions in Fabry disease and suggest UMOD as a potential biochemical marker of therapeutic response of the kidney to therapy. Extended comparative studies of UMOD expression in kidney specimens obtained during individual types of therapies are therefore of great interest.
- MeSH
- alfa-galaktosidasa terapeutické užití MeSH
- biologické markery metabolismus MeSH
- dospělí MeSH
- Fabryho nemoc farmakoterapie metabolismus patologie MeSH
- financování organizované MeSH
- ledvinové kanálky metabolismus patologie MeSH
- lidé středního věku MeSH
- lidé MeSH
- molekulární sekvence - údaje MeSH
- mucin 1 metabolismus MeSH
- mukoproteiny metabolismus moč MeSH
- nemoci ledvin etiologie metabolismus patologie MeSH
- posttranslační úpravy proteinů MeSH
- sekvence aminokyselin MeSH
- trihexosylceramidy metabolismus MeSH
- uromodulin MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
Autosomal dominant hyperuricemia, gout, renal cysts, and progressive renal insufficiency are hallmarks of a disease complex comprising familial juvenile hyperuricemic nephropathy and medullary cystic kidney diseases type 1 and type 2. In some families the disease is associated with mutations of the gene coding for uromodulin, but the link between the genetic heterogeneity and mechanism(s) leading to the common phenotype symptoms is not clear. In 19 families, we investigated relevant biochemical parameters, performed linkage analysis to known disease loci, sequenced uromodulin gene, expressed and characterized mutant uromodulin proteins, and performed immunohistochemical and electronoptical investigation in kidney tissues. We proved genetic heterogeneity of the disease. Uromodulin mutations were identified in six families. Expressed, mutant proteins showed distinct glycosylation patterns, impaired intracellular trafficking, and decreased ability to be exposed on the plasma membrane, which corresponded with the observations in the patient's kidney tissue. We found a reduction in urinary uromodulin excretion as a common feature shared by almost all of the families. This was associated with case-specific differences in the uromodulin immunohistochemical staining patterns in kidney. Our results suggest that various genetic defects interfere with uromodulin biology, which could lead to the development of the common disease phenotype. 'Uromodulin-associated kidney diseases' may be thus a more appropriate term for this syndrome.
- MeSH
- bazální membrána patologie ultrastruktura MeSH
- biopsie MeSH
- dítě MeSH
- dna (nemoc) MeSH
- dospělí MeSH
- financování organizované MeSH
- genetická heterogenita MeSH
- genetická vazba MeSH
- hyperurikemie genetika metabolismus MeSH
- hypofýza cytologie MeSH
- imunohistochemie MeSH
- kultivované buňky MeSH
- ledvinové kanálky patologie ultrastruktura MeSH
- ledviny chirurgie metabolismus patologie ultrastruktura MeSH
- lidé MeSH
- lidské chromozomy, pár 16 MeSH
- missense mutace MeSH
- mladiství MeSH
- mukoproteiny genetika metabolismus moč MeSH
- mutační analýza DNA MeSH
- polycystické ledviny autozomálně dominantní genetika MeSH
- polymorfismus délky restrikčních fragmentů MeSH
- rodokmen MeSH
- sekvence nukleotidů MeSH
- syndrom MeSH
- transfekce MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- srovnávací studie MeSH
BACKGROUND: Autosomal-dominant juvenile hyperuricemia, gouty arthritis, medullary cysts, and progressive renal insufficiency are features associated with familial juvenile hyperuricemic nephropathy (FJHN), medullary cystic kidney disease type 1 (MCKD1) and type 2 (MCKD2). MCKD1 has been mapped to chromosome 1q21. FJHN and MCKD2 have been mapped to chromosome 16p11.2. FJHN and MCKD2 are allelic, result from uromodulin (UMOD) mutations and the term uromodulin-associated kidney disease (UAKD) has been proposed for them. Linkage studies also reveal families that do not show linkage to any of the identified loci. To identify additional UAKD loci, we analyzed one of these families, with features suggestive of FJHN. METHODS: Clinical, biochemical, and immunohistochemical investigations were used for phenotype characterization. Genotyping, linkage and haplotype analyses were employed to identify the candidate disease region. Bioinformatics and sequencing were used for candidate gene selection and analyses. RESULTS: We identified a new candidate UAKD locus on chromosome 1q41, bounded by markers D1S3470 and D1S1644. We analyzed and found no linkage to this region in eight additional families, who did not map to the previously established loci. We noted that affected individuals showed, in addition to the characteristic urate hypoexcretion, significant reductions in urinary excretion of calcium and UMOD. Immunohistochemical analysis showed that low UMOD excretion resulted from its reduced expression, which is a different mechanism to intracellular UMOD accumulation observed in cases with UMOD mutations. CONCLUSION: We have mapped a new candidate UAKD locus and shown that UAKD may be a consequence of various defects affecting uromodulin biology.
- MeSH
- chronická renální insuficience genetika metabolismus MeSH
- dítě MeSH
- dospělí MeSH
- financování organizované MeSH
- genetická vazba MeSH
- genotyp MeSH
- hyperurikemie genetika metabolismus MeSH
- ledviny metabolismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- lidské chromozomy, pár 1 MeSH
- mapování chromozomů MeSH
- mladiství MeSH
- mukoproteiny genetika metabolismus MeSH
- rodokmen MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- Check Tag
- dítě MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- MeSH
- glykosaminoglykany metabolismus MeSH
- granulační tkáň metabolismus MeSH
- hojení ran MeSH
- izotopy stroncia metabolismus MeSH
- kosti a kostní tkáň metabolismus MeSH
- krysa rodu rattus MeSH
- monitorování radiace MeSH
- mukoproteiny metabolismus MeSH
- vazebná místa MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- MeSH
- alfa-1-antitrypsin izolace a purifikace krev metabolismus MeSH
- chemické modely MeSH
- elektroforéza sérových bílkovin metody využití MeSH
- glykoproteiny * izolace a purifikace krev metabolismus MeSH
- hexosaminy izolace a purifikace krev metabolismus MeSH
- hexosy izolace a purifikace krev metabolismus MeSH
- lidé MeSH
- membrány krevní zásobení metabolismus ultrastruktura MeSH
- mukoproteiny izolace a purifikace krev metabolismus MeSH
- polysacharidy krev metabolismus ultrastruktura MeSH
- sérum * enzymologie fyziologie metabolismus MeSH
- statistika jako téma MeSH
- ultrafiltrace * metody využití MeSH
- Check Tag
- lidé MeSH