Pancreatic and duodenal homeobox-1 nuclear localization is regulated by glucose in dispersed rat islets but not in insulin-secreting cell lines

Semache, Meriem; Ghislain, Julien; Zarrouki, Bader; Tremblay, Caroline; Poitout, Vincent

doi:10.4161/19382014.2014.982376

Cited by 5 publications

(3 citation statements)

References 61 publications

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“…MIN6 cells were cultured as previously described [ 20 ] and transfected with either 50 μmol Pax4 small interfering (si)RNA (Sigma) or scramble siRNA using Oligofectamine (Life Technologies, Madrid, Spain). Cells were either processed for RNA or treated with thapsigargin (Sigma-Aldrich, Madrid, Spain) to assess apoptosis 48 h after transfection.…”

Section: Methodsmentioning

confidence: 99%

PAX4 preserves endoplasmic reticulum integrity preventing beta cell degeneration in a mouse model of type 1 diabetes mellitus

et al. 2016

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Aims/hypothesisA strategy to enhance pancreatic islet functional beta cell mass (BCM) while restraining inflammation, through the manipulation of molecular and cellular targets, would provide a means to counteract the deteriorating glycaemic control associated with diabetes mellitus. The aims of the current study were to investigate the therapeutic potential of such a target, the islet-enriched and diabetes-linked transcription factor paired box 4 (PAX4), to restrain experimental autoimmune diabetes (EAD) in the RIP-B7.1 mouse model background and to characterise putative cellular mechanisms associated with preserved BCM.MethodsTwo groups of RIP-B7.1 mice were genetically engineered to: (1) conditionally express either PAX4 (BPTL) or its diabetes-linked mutant variant R129W (mutBPTL) using doxycycline (DOX); and (2) constitutively express luciferase in beta cells through the use of RIP. Mice were treated or not with DOX, and EAD was induced by immunisation with a murine preproinsulin II cDNA expression plasmid. The development of hyperglycaemia was monitored for up to 4 weeks following immunisation and alterations in the BCM were assessed weekly by non-invasive in vivo bioluminescence intensity (BLI). In parallel, BCM, islet cell proliferation and apoptosis were evaluated by immunocytochemistry. Alterations in PAX4- and PAX4R129W-mediated islet gene expression were investigated by microarray profiling. PAX4 preservation of endoplasmic reticulum (ER) homeostasis was assessed using thapsigargin, electron microscopy and intracellular calcium measurements.ResultsPAX4 overexpression blunted EAD, whereas the diabetes-linked mutant variant PAX4R129W did not convey protection. PAX4-expressing islets exhibited reduced insulitis and decreased beta cell apoptosis, correlating with diminished DNA damage and increased islet cell proliferation. Microarray profiling revealed that PAX4 but not PAX4R129W targeted expression of genes implicated in cell cycle and ER homeostasis. Consistent with the latter, islets overexpressing PAX4 were protected against thapsigargin-mediated ER-stress-related apoptosis. Luminal swelling associated with ER stress induced by thapsigargin was rescued in PAX4-overexpressing beta cells, correlating with preserved cytosolic calcium oscillations in response to glucose. In contrast, RNA interference mediated repression of PAX4-sensitised MIN6 cells to thapsigargin cell death.Conclusions/interpretationThe coordinated regulation of distinct cellular pathways particularly related to ER homeostasis by PAX4 not achieved by the mutant variant PAX4R129W alleviates beta cell degeneration and protects against diabetes mellitus. The raw data for the RNA microarray described herein are accessible in the Gene Expression Omnibus database under accession number GSE62846.Electronic supplementary materialThe online version of this article (doi:10.1007/s00125-016-3864-0) contains peer-reviewed but unedited supplementary material, which is available to authorised users.

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Section: Methodsmentioning

confidence: 99%

PAX4 preserves endoplasmic reticulum integrity preventing beta cell degeneration in a mouse model of type 1 diabetes mellitus

et al. 2016

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“…This antibody also recognized the same subcellular structures ( Supplementary Figure S6B ). This is not unexpected, since it is known that glucose levels induce a shift in the localization of a wide range of proteins [ 41 , 42 , 43 ]. In T. vaginalis , the availability of glucose affects the localization of other parasite proteins, such as TvCP2, TvAtg8a, TvTIM, and TvCat-D [ 19 , 30 , 44 , 45 ].…”

Section: Resultsmentioning

confidence: 85%

Trichomonas vaginalis Legumain-2, TvLEGU-2, Is an Immunogenic Cysteine Peptidase Expressed during Trichomonal Infection

Euceda-Padilla,

Mateo-Cruz,

Ávila-González

et al. 2024

Pathogens

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Trichomonas vaginalis is the causative agent of trichomoniasis, the most prevalent nonviral, neglected sexually transmitted disease worldwide. T. vaginalis has one of the largest degradomes among unicellular parasites. Cysteine peptidases (CPs) are the most abundant peptidases, constituting 50% of the degradome. Some CPs are virulence factors recognized by antibodies in trichomoniasis patient sera, and a few are found in vaginal secretions that show fluctuations in glucose concentrations during infection. The CPs of clan CD in T. vaginalis include 10 genes encoding legumain-like peptidases of the C13 family. TvLEGU-2 is one of them and has been identified in multiple proteomes, including the immunoproteome obtained with Tv (+) patient sera. Thus, our goals were to assess the effect of glucose on TvLEGU-2 expression, localization, and in vitro secretion and determine whether TvLEGU-2 is expressed during trichomonal infection. We performed qRT-PCR assays using parasites grown under different glucose conditions. We also generated a specific anti-TvLEGU-2 antibody against a synthetic peptide of the most divergent region of this CP and used it in Western blot (WB) and immunolocalization assays. Additionally, we cloned and expressed the tvlegu-2 gene (TVAG_385340), purified the recombinant TvLEGU-2 protein, and used it as an antigen for immunogenicity assays to test human sera from patients with vaginitis. Our results show that glucose does not affect tvlegu-2 expression but does affect localization in different parasite organelles, such as the plasma membrane, Golgi complex, hydrogenosomes, lysosomes, and secretion vesicles. TvLEGU-2 is secreted in vitro, is present in vaginal secretions, and is immunogenic in sera from Tv (+) patients, suggesting its relevance during trichomonal infection.

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“…Several authors have described that the TF Pdx1 plays a central role in the function of mature β‐cells, as it has been described as a key regulator of insulin gene expression. Mutations of the Pdx1 gene in humans result in defective insulin secretion, and the development of one form of maturity onset diabetes of the young 4 (MODY4) in humans (Fujimoto & Polonsky, 2009 ; Semache et al, 2014 ). Additionally, conditional deletion of Pdx1 in 80% of mature β‐cells in an inducible conditional knockout model induces diabetes by switching the identity of β‐cells to an α‐cell‐like phenotype (Gao et al, 2014 ).…”

Section: Discussionmentioning

confidence: 99%

IMT504 protects beta cells against apoptosis and maintains beta cell identity, without modifying proliferation

Converti

Bianchi

Martinez

et al. 2023

Physiological Reports

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We have demonstrated that oligodeoxynucleotide IMT504 promotes significant improvement in the diabetic condition in diverse animal models. Based on these results, here we evaluated whether these effects observed in vivo could be due to direct effects on β‐cells. We demonstrate by immunofluorescence that IMT504 enters the cell and locates in cytoplasm where it induces GSK‐3β phosphorylation that inactivates this kinase. As GSK‐3β tags Pdx1 for proteasomal degradation, by inactivating GSK‐3β, IMT504 induces an increase in Pdx1 protein levels, demonstrated by Western blotting. Concomitantly, an increase in Ins2 and Pdx1 gene transcription was observed, with no significant increase in insulin content or secretion. Enhanced Pdx1 is promising since it is a key transcription factor for insulin synthesis and is also described as an essential factor for the maintenance β‐cell phenotype and function. Dose‐dependent inhibition of H2O2‐induced apoptosis determined by ELISA as well as decreased expression of Bax was also observed. These results were confirmed in another β‐cell line, beta‐TC‐6 cells, in which a cytokine mix induced apoptosis that was reversed by IMT504. In addition, an inhibitor of IMT504 entrance into cells abrogated the effect IMT504. Based on these results we conclude that the β‐cell recovery observed in vivo may include direct effects of IMT504 on β‐cells, by maintaining their identity/phenotype and protecting them from oxidative stress and cytokine‐induced apoptosis. Thus, this work positions IMT504 as a promising option in the framework of the search of new therapies for type I diabetes treatment.

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Pancreatic and duodenal homeobox-1 nuclear localization is regulated by glucose in dispersed rat islets but not in insulin-secreting cell lines

Cited by 5 publications

References 61 publications

PAX4 preserves endoplasmic reticulum integrity preventing beta cell degeneration in a mouse model of type 1 diabetes mellitus

PAX4 preserves endoplasmic reticulum integrity preventing beta cell degeneration in a mouse model of type 1 diabetes mellitus

Trichomonas vaginalis Legumain-2, TvLEGU-2, Is an Immunogenic Cysteine Peptidase Expressed during Trichomonal Infection

IMT504 protects beta cells against apoptosis and maintains beta cell identity, without modifying proliferation

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