New findings on the structure of the FOXO4: p53 complex - a key factor in senescence regulation (22.4. 2022)
Transcription factor p53 protects cells against tumorigenesis when subjected to various cellular stresses. Under stress conditions, p53 interacts with another transcription factor, FOXO4 (Forkhead box O 4), and together they increase the production of p21 protein, which triggers the process of cell aging (senescence). However, the molecular mechanism of upregulation of p21 transcription is still unclear. In the study published in the Protein Science journal, scientific teams of Dr. Obsilova (IPHYS CAS), prof. Obsil (Faculty of Science, Charles University and IPHYS CAS) and their colleagues from IOCB CAS characterized interactions between p53 and FOXO4 at the molecular level. New knowledge about the structure of the complex may enable the development of specific inhibitors of the interaction between these two proteins, and subsequently in the development of new drugs aimed at the selective elimination of senescent cells.
In this structural study, the researchers performed a detailed characterization of the interactions in the FOXO4: p53 complex using an integrated approach involving analytical ultracentrifugation, nuclear magnetic resonance, and chemical cross-linking coupled to mass spectrometry. Because both FOXO4 and p53 have multiple domains (see Figure), they studied the role of individual domains and disordered segments of both proteins and mapped their interaction interfaces. They found out that the interaction between p53 transactivation domain TAD and the FOXO4 Forkhead domain is crucial for the overall stability of the p53:FOXO4 complex. Furthermore, contacts involving the N-terminal disordered FOXO4 segment, the C-terminal negative regulatory domain of p53, and the DNA-binding domains of both proteins stabilize the complex formation. By measuring DNA binding, they further found that the p53: FOXO4 complex formation blocks p53 binding to DNA without affecting the DNA-binding properties of FOXO4.
Left, sedimentation velocity analytical ultracentrifugation analysis of interaction between FOXO4 and p53. Middle, chemical shift perturbations obtained from 1H-15N HSQC spectra of 15N-labeled FOXO4 in the presence of p53 mapped onto the crystal structure of the FOXO4 DBD:DNA complex. Right, fluorescence anisotropy measurements showing that the complex formation reduces the DNA-binding affinity of p53.
Mandal R, Kohoutova K, Petrvalska O, Horvath M, Srb P, Veverka V, Obsilova V and Obsil T. FOXO4 interacts with p53 TAD and CRD and inhibits its binding to DNA. Protein Sci. roč. 31, č. 5 (2022), č. článku e4287. IF = 6.725. DOI
An oral glucose tolerance test (OGTT) is the most commonly used method to diagnose diabetes mellitus from a drop of blood. It measures the ability of an organism to clear circulating glucose after ingestion of glucose bolus after an overnight fast. Although the dynamics of the blood glucose levels during the OGTT are well known, much less information about the metabolic changes in the target organs and the inter-organ communication are available. In our study, we investigated what is the fate of the sugar molecules in each organ and how it affects metabolic pathways in the body. Therefore, we performed the OGTT in mice using glucose with stable isotopic tracers (13C), analyzed 13C6-glucose tissue distribution and time profiles of metabolites and lipids across 12 organs and plasma. We found, that during the OGTT, the glucose use is turned on with specific kinetics at the organ level, but fasting substrates like β-hydroxybutyrate are switched off in all organs simultaneously. Timeline profiling of 13C-labeled fatty acids and triacylglycerols across tissues suggests that brown adipose tissue may contribute to the circulating fatty acid pool at maximal plasma glucose levels. We have created a virtual interactive atlas of metabolites (sugars, amino acids, lipids, etc.), which describes the interactions between organs after ingesting grape sugar. Metabolic fate of ingested glucose carbons was followed in 12 organs and plasma.
Visit the web application to explore virtual mouse metabolome yourself.
Lopes M, Brejchova K, Riecan M, Novakova M, Rossmeisl M, Cajka T, Kuda O. Metabolomics atlas of oral 13C-glucose tolerance test in mice. Cell Rep. 2021 Oct 12;37(2):109833. DOI. PMID: 34644567 IF = 9,423
The effectivity of opioid analgesics is reduced by CCL2 chemokine (8.12. 2021)
Opioid analgesics are the standard of care in the treatment of serious painful states. Treatment of neuropathic pain states, induced by damage to the nervous system, is especially difficult and opioid analgesics often do not have a beneficial effect. It was shown before that neuropathic states are accompanied with neuroinflammatory changes in the spinal cord and the level of different signaling molecules such as chemokine CCL2 is increased. This work shows that chemokine CCL2 is one of the important factors significantly reducing the effectivity of analgesics acting through opioid receptors. It acts probably directly on neurons and also through activation of microglial cells. Analgesic treatment with opioids has also number of serious unwanted side effects. One of them is a paradoxical increase of sensitivity, hyperalgesia/pain after opioids administration. This work shows that this hyperalgesia may be related to TRPV1 receptors activation. These published results suggest that to improve pain treatment with opioid analgesics, modulation of CCL2 and TRPV1 receptors may be needed, especially in cases of neuropathic pain.
The inhibition of the opioid agonists induced reduction of painful/nociceptive signaling in the spinal cord dorsal horn by the CCL2 chemokine. The control blue line demonstrates the inhibition of the nociceptive synaptic signaling after the DAMGO application and later (in about 13minutes) its potentiation. DAMGO is µ opioid receptors agonist and simulates thus application of opioid analgesics. The analgesic effect of DAMGO application is completely reversed in the presence of CCL2 chemokine (red line). The purple line demonstrates that the effect of CCL2 is dependent on microglia cells activation as it is prevented by microglia inhibitor minocycline.
Chemokine CCL2 preventsopioid‑inducedinhibitionofnociceptivesynaptictransmission in spinalcorddorsalhorn. Mario Heles, Petra Mrozkova, Dominika Sulcova, Pavel Adamek, Diana Spicarova and Jiri Palecek, JournalofNeuroinflammation (2021) 18:279, DOI, IF=8.23
Laboratory of Pain Research, Institute of Physiology, The Czech Academy of Sciences, Videnska 1083, 142 20 Praha 4, Czech Republic
Gliflozins - more than just antidiabetics (8.12. 2021)
Gliflozins are commonly prescribed for the treatment of diabetes (patients may know e.g. empagliflozin under the brand name Jardiance). Gliflozins inhibit the activity of sodium-glucose transporter in kidneys, which leads to higher glucose excretion in urine and normalization of blood glucose levels. Besides alleviation of hyperglycemia, other beneficial effects were observed in patients taking gliflozins, including decreased body weight, reduced blood pressure or improved kidney function. How this is possible? Here comes the right time to take a step backwards and to look once again and in greater detail on gliflozins effects in rodent models. And that is exactly what dr. Vaneckova and her colleagues from IKEM and departments of IPHYS are doing. They characterized the effects of empagliflozin in various non-diabetic rodent models prone to hypertension (Ren-2 transgenic rats; TGR) and lipid imbalance (hereditary hypertensive rats). Although the details of treatment effects vary among the individual models, empagliflozin generally attenuated inflammation, and normalized plasma and tissue lipid levels. In addition, it inhibited the activity of sympathetic nervous system, which resulted in a decrease of blood pressure in TGR. Hopefully, this and similar findings will allow to extend the use of gliflozins in the future to more diagnoses than just diabetes.
Hojná S, Rauchová H, Malínská H, Marková I, Hüttl M, Papoušek F, Behuliak M, Miklánková D, Vaňourková Z, Neckář J, Kadlecová M, Kujal P, Zicha J, Vaněčková I. Antihypertensive and metabolic effects of empagliflozin in Ren-2 transgenic rats, an experimental non-diabetic model of hypertension. Biomed Pharmacother. 2021 Dec;144:112246. Epub 2021 Oct 1. IF-5,98, DOI
Targeted modulation of NMDA receptors is a key for the effective treatment of neurological and neurodevelopmental diseases (14.10. 2021)
N-methyl-D-aspartate (NMDA) receptors are glutamate-gated ion channels critically involved in excitatory synaptic transmission that play a key role in learning and memory. Impaired NMDA receptor function leads to major neurological, neurodevelopmental and psychiatric disorders such as schizophrenia, autistic spectrum disorder, epilepsy or Alzheimer's disease. For an effective design of novel drugs capable of specifically modulating NMDA receptors, it is essential not only to understand the NMDA receptor atomic structure, but also to uncover the specific sequence of conformational changes that are involved in receptor activation and allosteric modulation.
We used single-molecule FRET to identify and quantify the sequence of conformational changes in the amino-terminal domain of the NMDA receptor during its activation. Next, we uncovered distinct roles of receptor subunits in receptor activation, and last but not least, we have identified the molecular mechanism of receptor modulation by pH during pathophysiological conditions such as stroke.
Vyklický, Vojtěch - Stanley, Ch. - Habrian, Ch. - Isacoff, E. Y. Conformational rearrangement of the NMDA receptor amino-terminal domain during activation and allosteric modulation. Nature Communications. Roč. 12, č. 1 (2021), č. článku 2694. ISSN 2041-1723. E-ISSN 2041-1723, IF: 14.919, rok: 2020, DOI