The insulin receptor (IR, with its isoforms IR-A and IR-B) and the insulin-like growth factor 1 receptor (IGF-1R) are related tyrosine kinase receptors. Recently, the portfolio of solved hormone-receptor structures has grown extensively thanks to advancements in cryo-electron microscopy. However, the dynamics of how these receptors transition between their inactive and active state are yet to be fully understood. The C-terminal part of the alpha subunit (αCT) of the receptors is indispensable for the formation of the hormone-binding site. We mutated the αCT residues Arg717 and His710 of IR-A and Arg704 and His697 of IGF-1R. We then measured the saturation binding curves of ligands on the mutated receptors and their ability to become activated. Mutations of Arg704 and His697 to Ala in IGF-1R decreased the binding of IGF-1. Moreover, the number of binding sites for IGF-1 on the His697 IGF-1R mutant was reduced to one-half, demonstrating the presence of two binding sites. Both mutations of Arg717 and His710 to Ala in IR-A inactivated the receptor. We have proved that Arg717 is important for the binding of insulin to its receptor, which suggests that Arg717 is a key residue for the transition to the active conformation.

• Klíčová slova
• mutagenesis in vitro, peptide hormone, receptor modification, receptor tyrosine kinase, structure–function,
• MeSH
• elektronová kryomikroskopie MeSH
• insulinu podobný růstový faktor I genetika   chemie   metabolismus   MeSH
• inzulin metabolismus   MeSH
• ligandy MeSH
• receptor IGF typ 1 * genetika   chemie   metabolismus   MeSH
• receptor inzulinu * genetika   chemie   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• insulinu podobný růstový faktor I MeSH
• inzulin MeSH
• ligandy MeSH
• receptor IGF typ 1 * MeSH
• receptor inzulinu * MeSH

Evidence accumulates that the functional plasticity of insulin and insulin-like growth factor signaling in insects could spring, among others, from the multiplicity of insulin receptors (InRs). Their multiple variants may be implemented in the control of insect polyphenism, such as wing or caste polyphenism. Here, we present a comprehensive phylogenetic analysis of insect InR sequences in 118 species from 23 orders and investigate the role of three InRs identified in the linden bug, Pyrrhocoris apterus, in wing polymorphism control. We identified two gene clusters (Clusters I and II) resulting from an ancestral duplication in a late ancestor of winged insects, which remained conserved in most lineages, only in some of them being subject to further duplications or losses. One remarkable yet neglected feature of InR evolution is the loss of the tyrosine kinase catalytic domain, giving rise to decoys of InR in both clusters. Within the Cluster I, we confirmed the presence of the secreted decoy of insulin receptor in all studied Muscomorpha. More importantly, we described a new tyrosine kinase-less gene (DR2) in the Cluster II, conserved in apical Holometabola for ∼300 My. We differentially silenced the three P. apterus InRs and confirmed their participation in wing polymorphism control. We observed a pattern of Cluster I and Cluster II InRs impact on wing development, which differed from that postulated in planthoppers, suggesting an independent establishment of insulin/insulin-like growth factor signaling control over wing development, leading to idiosyncrasies in the co-option of multiple InRs in polyphenism control in different taxa.

• Klíčová slova
• decoy of insulin receptor, gene structure, insects, insulin receptor, insulin signaling, wing polyphenism,
• MeSH
• biologická evoluce * MeSH
• duplikace genu MeSH
• Heteroptera genetika   růst a vývoj   MeSH
• hmyz anatomie a histologie   genetika   MeSH
• křídla zvířecí anatomie a histologie   růst a vývoj   MeSH
• receptor inzulinu genetika   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• receptor inzulinu MeSH

Annona and ginger have prominent uses in traditional medicine; their therapeutic properties have not been sufficiently explored. The ameliorative effect of Annona or ginger extracts on hyperglycaemia associated with oxidative stress, inflammation, and apoptosis in experimentally induced diabetes was addressed. Type 1 diabetes in male rats was induced by a single injection of streptozotocin (STZ; 40 mg/kg, i.p.), then Annona (100 mg/kg) or ginger (200 mg/kg) extracts were orally administered daily for 30 days. The Annona and ginger extracts ameliorated hyperglycaemia, insulin level, glycosylated haemoglobin (HbA1c) and insulin resistance (HOMA-IR) levels in the diabetic rats. The treatments significantly ameliorated liver function enzymes and total proteins; this was confirmed by histopathological examination of liver sections. Annona and ginger extracts significantly reduced elevated malondialdehyde (MDA) and restored activity of antioxidant enzymes in the liver such as glutathione peroxidase (GPx), glutathione reductase (GR), superoxide dismutase (SOD), and catalase (CAT) and the hepatic content of reduced glutathione (GSH). The oxidative stressdependent inflammation was regulated by both Annona and ginger extracts, which was indicated by down-regulation of TNF-α, NF-κB, pro-apoptotic proteins Bax, p53, and anti-apoptotic protein Bcl-2. Moreover, the expression of insulin receptor (INSR) and glucose transporter 2 (GLUT2) genes was markedly regulated by both these extracts. The results suggest that Annona and ginger extracts ameliorate the hepatic damage resulting from diabetes by advocating antioxidants and modulating apoptotic mediator proteins in the liver of diabetic rats. In conclusion, Annona and ginger extracts have a potential therapeutic effect in the treatment of diabetes and its complications.

• MeSH
• Annona chemie   MeSH
• antioxidancia metabolismus   MeSH
• apoptóza účinky léků   MeSH
• biologické markery krev   MeSH
• experimentální diabetes mellitus krev   farmakoterapie   enzymologie   MeSH
• glykovaný hemoglobin metabolismus   MeSH
• inzulin krev   MeSH
• inzulinová rezistence MeSH
• jaterní testy MeSH
• játra účinky léků   patofyziologie   MeSH
• krevní glukóza metabolismus   MeSH
• malondialdehyd metabolismus   MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• NF-kappa B metabolismus   MeSH
• oxidační stres účinky léků   MeSH
• potkani Wistar MeSH
• přenašeč glukosy typ 2 genetika   metabolismus   MeSH
• protein X asociovaný s bcl-2 metabolismus   MeSH
• receptor inzulinu genetika   metabolismus   MeSH
• regulace genové exprese MeSH
• rostlinné extrakty farmakologie   terapeutické užití   MeSH
• TNF-alfa metabolismus   MeSH
• zázvor lékařský chemie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antioxidancia MeSH
• biologické markery MeSH
• glykovaný hemoglobin MeSH
• inzulin MeSH
• krevní glukóza MeSH
• malondialdehyd MeSH
• nádorový supresorový protein p53 MeSH
• NF-kappa B MeSH
• přenašeč glukosy typ 2 MeSH
• protein X asociovaný s bcl-2 MeSH
• receptor inzulinu MeSH
• rostlinné extrakty MeSH
• TNF-alfa MeSH

Parasitic nematodes transition between dramatically different free-living and parasitic stages, with correctly timed development and migration crucial to successful completion of their lifecycle. However little is known of the mechanisms controlling these transitions. microRNAs (miRNAs) negatively regulate gene expression post-transcriptionally and regulate development of diverse organisms. Here we used microarrays to determine the expression profile of miRNAs through development and in gut tissue of the pathogenic nematode Haemonchus contortus. Two miRNAs, mir-228 and mir-235, were enriched in infective L3 larvae, an arrested stage analogous to Caenorhabditis elegans dauer larvae. We hypothesized that these miRNAs may suppress development and maintain arrest. Consistent with this, inhibitors of these miRNAs promoted H. contortus development from L3 to L4 stage, while genetic deletion of C. elegans homologous miRNAs reduced dauer arrest. Epistasis studies with C. elegans daf-2 mutants showed that mir-228 and mir-235 synergise with FOXO transcription factor DAF-16 in the insulin signaling pathway. Target prediction suggests that these miRNAs suppress metabolic and transcription factor activity required for development. Our results provide novel insight into the expression and functions of specific miRNAs in regulating nematode development and identify miRNAs and their target genes as potential therapeutic targets to limit parasite survival within the host.

• MeSH
• Caenorhabditis elegans genetika   MeSH
• cholesteny farmakologie   MeSH
• delece genu MeSH
• druhová specificita MeSH
• genová ontologie MeSH
• Haemonchus účinky léků   genetika   růst a vývoj   MeSH
• larva MeSH
• messenger RNA genetika   metabolismus   MeSH
• mikro RNA biosyntéza   genetika   MeSH
• proteiny Caenorhabditis elegans genetika   MeSH
• receptor inzulinu genetika   MeSH
• RNA helmintů biosyntéza   genetika   MeSH
• vývojová regulace genové exprese účinky léků   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• cholesteny MeSH
• DAF-2 protein, C elegans MeSH Prohlížeč
• dafachronic acid MeSH Prohlížeč
• messenger RNA MeSH
• mikro RNA MeSH
• proteiny Caenorhabditis elegans MeSH
• receptor inzulinu MeSH
• RNA helmintů MeSH

Information on how insulin and insulin-like growth factors 1 and 2 (IGF-1 and -2) activate insulin receptors (IR-A and -B) and the IGF-1 receptor (IGF-1R) is crucial for understanding the difference in the biological activities of these peptide hormones. Cryo-EM studies have revealed that insulin uses its binding sites 1 and 2 to interact with IR-A and have identified several critical residues in binding site 2. However, mutagenesis studies suggest that Ile-A10, Ser-A12, Leu-A13, and Glu-A17 also belong to insulin's site 2. Here, to resolve this discrepancy, we mutated these insulin residues and the equivalent residues in IGFs. Our findings revealed that equivalent mutations in the hormones can result in differential biological effects and that these effects can be receptor-specific. We noted that the insulin positions A10 and A17 are important for its binding to IR-A and IR-B and IGF-1R and that A13 is important only for IR-A and IR-B binding. The IGF-1/IGF-2 positions 51/50 and 54/53 did not appear to play critical roles in receptor binding, but mutations at IGF-1 position 58 and IGF-2 position 57 affected the binding. We propose that IGF-1 Glu-58 interacts with IGF-1R Arg-704 and belongs to IGF-1 site 1, a finding supported by the NMR structure of the less active Asp-58-IGF-1 variant. Computational analyses indicated that the aforementioned mutations can affect internal insulin dynamics and inhibit adoption of a receptor-bound conformation, important for binding to receptor site 1. We provide a molecular model and alternative hypotheses for how the mutated insulin residues affect activity.

• Klíčová slova
• NMR structure, complex, hormone analog, insulin, insulin-like growth factor (IGF), molecular dynamics, mutagenesis, peptide hormone, receptor autophosphorylation, receptor binding, receptor tyrosine kinase, structural biology, structure-function,
• MeSH
• insulinu podobný růstový faktor I chemie   genetika   MeSH
• insulinu podobný růstový faktor II chemie   genetika   MeSH
• inzulin analogy a deriváty   chemická syntéza   chemie   genetika   MeSH
• lidé MeSH
• mnohočetné abnormality genetika   MeSH
• mutace genetika   MeSH
• nukleární magnetická rezonance biomolekulární MeSH
• poruchy růstu genetika   MeSH
• proteinové domény genetika   MeSH
• receptor IGF typ 1 chemie   genetika   MeSH
• receptor inzulinu chemie   genetika   MeSH
• sekvence aminokyselin genetika   MeSH
• vazba proteinů genetika   MeSH
• vazebná místa genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• IGF1R protein, human MeSH Prohlížeč
• IGF2 protein, human MeSH Prohlížeč
• insulinu podobný růstový faktor I MeSH
• insulinu podobný růstový faktor II MeSH
• inzulin MeSH
• receptor IGF typ 1 MeSH
• receptor inzulinu MeSH

Insulin and insulin-like growth factor 1 (IGF-1) are closely related hormones involved in the regulation of metabolism and growth. They elicit their functions through activation of tyrosine kinase-type receptors: insulin receptors (IR-A and IR-B) and IGF-1 receptor (IGF-1R). Despite similarity in primary and three-dimensional structures, insulin and IGF-1 bind the noncognate receptor with substantially reduced affinity. We prepared [d-HisB24, GlyB31, TyrB32]-insulin, which binds all three receptors with high affinity (251 or 338% binding affinity to IR-A respectively to IR-B relative to insulin and 12.4% binding affinity to IGF-1R relative to IGF-1). We prepared other modified insulins with the aim of explaining the versatility of [d-HisB24, GlyB31, TyrB32]-insulin. Through structural, activity, and kinetic studies of these insulin analogs, we concluded that the ability of [d-HisB24, GlyB31, TyrB32]-insulin to stimulate all three receptors is provided by structural changes caused by a reversed chirality at the B24 combined with the extension of the C terminus of the B chain by two extra residues. We assume that the structural changes allow the directing of the B chain C terminus to some extra interactions with the receptors. These unusual interactions lead to a decrease of dissociation rate from the IR and conversely enable easier association with IGF-1R. All of the structural changes were made at the hormones' Site 1, which is thought to interact with the Site 1 of the receptors. The results of the study suggest that merely modifications of Site 1 of the hormone are sufficient to change the receptor specificity of insulin.

• Klíčová slova
• Site 1, binding, insulin, insulin receptor, insulin-like growth factor (IGF), kinetics, protein design, structure-function,
• MeSH
• insulinu podobný růstový faktor I chemie   genetika   metabolismus   MeSH
• inzulin agonisté   metabolismus   MeSH
• kinetika MeSH
• krystalografie rentgenová MeSH
• lidé MeSH
• receptor IGF typ 1 MeSH
• receptor inzulinu chemie   genetika   metabolismus   MeSH
• receptory somatomedinů chemie   genetika   metabolismus   MeSH
• sekvence aminokyselin MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• IGF1R protein, human MeSH Prohlížeč
• insulinu podobný růstový faktor I MeSH
• inzulin MeSH
• receptor IGF typ 1 MeSH
• receptor inzulinu MeSH
• receptory somatomedinů MeSH

Insulin and insulin-like growth factors I and II are closely related protein hormones. Their distinct evolution has resulted in different yet overlapping biological functions with insulin becoming a key regulator of metabolism, whereas insulin-like growth factors (IGF)-I/II are major growth factors. Insulin and IGFs cross-bind with different affinities to closely related insulin receptor isoforms A and B (IR-A and IR-B) and insulin-like growth factor type I receptor (IGF-1R). Identification of structural determinants in IGFs and insulin that trigger their specific signaling pathways is of increasing importance in designing receptor-specific analogs with potential therapeutic applications. Here, we developed a straightforward protocol for production of recombinant IGF-II and prepared six IGF-II analogs with IGF-I-like mutations. All modified molecules exhibit significantly reduced affinity toward IR-A, particularly the analogs with a Pro-Gln insertion in the C-domain. Moreover, one of the analogs has enhanced binding affinity for IGF-1R due to a synergistic effect of the Pro-Gln insertion and S29N point mutation. Consequently, this analog has almost a 10-fold higher IGF-1R/IR-A binding specificity in comparison with native IGF-II. The established IGF-II purification protocol allowed for cost-effective isotope labeling required for a detailed NMR structural characterization of IGF-II analogs that revealed a link between the altered binding behavior of selected analogs and conformational rearrangement of their C-domains.

• Klíčová slova
• insulin, insulin receptor, insulin-like growth factor (IGF), nuclear magnetic resonance (NMR), structural biology, structure-function,
• MeSH
• CD antigeny chemie   genetika   metabolismus   MeSH
• insulinu podobný růstový faktor II chemie   genetika   metabolismus   MeSH
• lidé MeSH
• missense mutace MeSH
• protein - isoformy chemie   genetika   metabolismus   MeSH
• proteinové domény MeSH
• receptor IGF typ 1 chemie   genetika   metabolismus   MeSH
• receptor inzulinu chemie   genetika   metabolismus   MeSH
• rekombinantní proteiny chemie   genetika   metabolismus   MeSH
• substituce aminokyselin MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• CD antigeny MeSH
• IGF2 protein, human MeSH Prohlížeč
• INSR protein, human MeSH Prohlížeč
• insulinu podobný růstový faktor II MeSH
• protein - isoformy MeSH
• receptor IGF typ 1 MeSH
• receptor inzulinu MeSH
• rekombinantní proteiny MeSH

Despite the recent first structural insight into the insulin-insulin receptor complex, the role of the C terminus of the B-chain of insulin in this assembly remains unresolved. Previous studies have suggested that this part of insulin must rearrange to reveal amino acids crucial for interaction with the receptor. The role of the invariant Phe(B24), one of the key residues of the hormone, in this process remains unclear. For example, the B24 site functionally tolerates substitutions to D-amino acids but not to L-amino acids. Here, we prepared and characterized a series of B24-modified insulin analogues, also determining the structures of [D-HisB24]-insulin and [HisB24]-insulin. The inactive [HisB24]-insulin molecule is remarkably rigid due to a tight accommodation of the L-His side chain in the B24 binding pocket that results in the stronger tethering of B25-B28 residues to the protein core. In contrast, the highly active [D-HisB24]-insulin is more flexible, and the reverse chirality of the B24C(α) atom swayed the D-His(B24) side chain into the solvent. Furthermore, the pocket vacated by Phe(B24) is filled by Phe(B25), which mimics the Phe(B24) side and main chains. The B25→B24 downshift results in a subsequent downshift of Tyr(B26) into the B25 site and the departure of B26-B30 residues away from the insulin core. Our data indicate the importance of the aromatic L-amino acid at the B24 site and the structural invariance/integrity of this position for an effective binding of insulin to its receptor. Moreover, they also suggest limited, B25-B30 only, unfolding of the C terminus of the B-chain upon insulin activation.

• MeSH
• inzulin chemie   genetika   metabolismus   MeSH
• lidé MeSH
• receptor inzulinu chemie   genetika   metabolismus   MeSH
• sekundární struktura proteinů MeSH
• vazba proteinů fyziologie   MeSH
• vazebná místa MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• inzulin MeSH
• receptor inzulinu MeSH

The pathogenesis of myotonic dystrophy type 2 includes the sequestration of MBNL proteins by expanded CCUG transcripts, which leads to an abnormal splicing of their target pre-mRNAs. We have found CCUG(exp) RNA transcripts of the ZNF9 gene associated with the formation of ribonuclear foci in human skeletal muscle and some non-muscle tissues present in muscle biopsies and skin excisions from myotonic dystrophy type 2 patients. Using RNA-FISH and immunofluorescence-FISH methods in combination with a high-resolution confocal microscopy, we demonstrate a different frequency of nuclei containing the CCUG(exp) foci, a different expression pattern of MBNL1 protein and a different sequestration of MBNL1 by CCUG(exp) repeats in skeletal muscle, vascular smooth muscle and endothelia, Schwann cells, adipocytes, and ectodermal derivatives. The level of CCUG(exp) transcription in epidermal and hair sheath cells is lower compared with that in other tissues examined. We suppose that non-muscle tissues of myotonic dystrophy type 2 patients might be affected by a similar molecular mechanism as the skeletal muscle, as suggested by our observation of an aberrant insulin receptor splicing in myotonic dystrophy type 2 adipocytes.

• MeSH
• aktiny metabolismus   MeSH
• analýza rozptylu MeSH
• antigeny CD34 metabolismus   MeSH
• endotel metabolismus   patologie   MeSH
• konfokální mikroskopie MeSH
• kosterní svaly metabolismus   MeSH
• kůže metabolismus   patologie   MeSH
• lidé MeSH
• myotonická dystrofie MeSH
• myotonické poruchy * diagnóza   genetika   metabolismus   patologie   MeSH
• neurofilamentové proteiny metabolismus   MeSH
• proteiny S100 metabolismus   MeSH
• proteiny vázající RNA genetika   metabolismus   MeSH
• receptor inzulinu genetika   MeSH
• repetitivní sekvence nukleových kyselin genetika   MeSH
• RNA metabolismus   MeSH
• sestřih RNA genetika   MeSH
• transport proteinů fyziologie   MeSH
• tukové buňky metabolismus   patologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aktiny MeSH
• antigeny CD34 MeSH
• CNBP protein, human MeSH Prohlížeč
• MBNL1 protein, human MeSH Prohlížeč
• neurofilamentové proteiny MeSH
• proteiny S100 MeSH
• proteiny vázající RNA MeSH
• receptor inzulinu MeSH
• RNA MeSH

During 3 years of collaboration we have detected 13 patients with insulin resistance syndromes among the 800 patients referred to our genetic counseling center for diagnosis during that time. It represents about 1.5% of all diagnostic problems of our genetic counseling center. Thus it seems to us that insulin receptor disorders are not as rare as previously appreciated. We describe here the clinical and biochemical findings of four of them as well as characteristics of disorders of insulin receptors. On cellular level these four patients represents the full scale of insulin receptor defects and demonstrate genetic heterogeneity of these disorders.

• MeSH
• dítě MeSH
• inzulin metabolismus   MeSH
• inzulinová rezistence * MeSH
• lidé MeSH
• mladiství MeSH
• předškolní dítě MeSH
• receptor inzulinu genetika   MeSH
• RNA biosyntéza   MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• Názvy látek
• inzulin MeSH
• receptor inzulinu MeSH
• RNA MeSH