Pochopení vztahu mezi senzomotorickými proměnnými a exerkiny, které ovlivňují funkci mozku a kognici, nám umožňuje hlouběji porozumět biologickému procesu stárnutí. Hlavním cílem této studie bylo zjistit, jak silně jsou mozkový neurotrofický faktor (brain-derived neurotrophic factor, BDNF), irisin, svalová hmota a svalová síla asociovány s výsledky testů vybraných kognitivních funkcí u starších žen a jak dobře je predikují. Padesát sedm starších žen (průměrný věk 70,4 ± 4,1 roku) absolvovalo baterii neuropsychologických testů, měření izometrické dynamometrie a bioelektrické impedance. Hladiny v krevním séru sledovaných exerkinů byly stanoveny enzymatickým imunosorbentním testem (ELISA). Pro testování predikcí byly využity hierarchické vícenásobné regresní modely. Odhadli jsme, že rozptyl 46,1 % v krátkodobé paměti byl zapříčiněn hladinami BDNF v séru, přičemž druhým statisticky významným prediktorem byl věk (beta = –0,22; p = 0,030). Síla dolních končetin (lower limb strength, LLS) prokázala významnou prediktivní sílu jak u paměti – bezprostřední vybavení (beta = 0,39; p = 0,004), tak u paměti – oddálené vybavení (beta = 0,45; p = 0,001). Hladiny BDNF v séru byly významným prediktorem u oddáleného vybavení (beta = 0,29; p = 0,048). Přidání hladin BDNF do modelu prokázalo významné zvýšení jeho prediktivní síly o přibližně 5,6 % (p = 0,048) u paměti – oddálené vybavení. Index kosterní svalové hmoty (skeletal muscle index, SMI) a úroveň vzdělání byly významnými prediktory mentální flexibility. Byla zjištěna silná pozitivní asociace mezi hladinami BDNF, irisinem, svalovou silou a kognitivní funkcí, přičemž irisin a svalová síla jsou silnými prediktory hladin BDNF u starších žen. Studie byla realizována s podporou grantu Univerzity Karlovy – PRIMUS/19/HUM/012, Specifického vysokoškolského výzkumu SVV 260599, projektu COOPERATIO a Grantové agentury UK číslo grantu 268321. Korespondenční adresa: PhDr. Veronika Holá Katedra gymnastiky a úpolových sportů FTVS UK José Martího 269/31 162 52 Praha 6-Veleslavín e-mail: veronika.hola@ftvs.cuni.cz
Understanding the relationship between sensorimotor variables and exerkines related to brain function and cognition may help better understand biological ageing. The main aim of this study was to determine how strongly brain-derived neurotrophic factor (BDNF), irisin, muscle mass and muscle strength are associated and predict scores on selected cognitive domain tests in older women. Fifty seven older women (mean age 70.4 ± 4.1 years) underwent a battery of cognitive and psychological tests and measurements of isometric dynamometry and bioelectrical impedance. Serum exerkines levels were measured by enzyme-linked immunosorbent assay (ELISA). Hierarchical multiple regression models were used to test the predictions. We estimated that 46.1% of the variance in short-term memory was accounted for by serum BDNF levels, with age being the second statistically significant predictor (Beta = -0.22; p = 0.030). Lower limb strength (LLS) showed significant predictive power in both immediate (Beta = 0.39; p = 0.004) and delayed memory (Beta = 0.45; p = 0.001), serum BDNF levels were a significant predictor in delayed memory (Beta = 0.29; p = 0.048). Adding serum BDNF levels to the model showed a significant increase in predictive power of approximately 5.6% (p = 0.048) in delayed memory. Skeletal muscle index (SMI) and education level were significant predictors of mental flexibility. A strong positive association between BDNF levels, irisin, muscle strength, and cognitive function was found, with irisin and muscle strength being strong predictors of BDNF levels in older women.
- Klíčová slova
- irisin,
- MeSH
- fibronektinová doména typu III fyziologie MeSH
- kognice fyziologie MeSH
- kognitivní stárnutí * fyziologie MeSH
- lidé MeSH
- mozkový neurotrofický faktor krev MeSH
- neuropsychologické testy * statistika a číselné údaje MeSH
- paměť fyziologie MeSH
- prognóza MeSH
- průřezové studie MeSH
- regresní analýza MeSH
- senioři MeSH
- svalová atrofie etiologie MeSH
- svalová síla fyziologie MeSH
- Check Tag
- lidé MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- práce podpořená grantem MeSH
The overexpression of MYC genes is frequently found in many human cancers, including adult and pediatric malignant brain tumors. Targeting MYC genes continues to be challenging due to their undruggable nature. Using our prediction algorithm, the nine-amino-acid activation domain (9aaTAD) has been identified in all four Yamanaka factors, including c-Myc. The predicted activation function was experimentally demonstrated for all these short peptides in transactivation assay. We generated a set of c-Myc constructs (1-108, 69-108 and 98-108) in the N-terminal regions and tested their ability to initiate transcription in one hybrid assay. The presence and absence of 9aaTAD (region 100-108) in the constructs strongly correlated with their activation functions (5-, 3- and 67-times respectively). Surprisingly, we observed co-activation function of the myc region 69-103, called here acetyl-TAD, previously described by Faiola et al. (Mol Cell Biol 25:10220-10234, 2005) and characterized in this study as a new domain collaborating with the 9aaTAD. We discovered strong interactions on a nanomolar scale between the Myc-9aaTAD activation domains and the KIX domain of CBP coactivator. We showed conservation of the 9aaTADs in the MYC family. In summary for the c-Myc oncogene, the acetyl-TAD and the 9aaTAD domains jointly mediated activation function. The c-Myc protein is largely intrinsically disordered and therefore difficult to target with small-molecule inhibitors. For the c-Myc driven tumors, the strong c-Myc interaction with the KIX domain represents a promising druggable target.
- MeSH
- aktivace transkripce MeSH
- lidé MeSH
- proteinové domény MeSH
- protoonkogenní proteiny c-myc * metabolismus genetika MeSH
- sekvence aminokyselin MeSH
- vazba proteinů * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
MICAL proteins play a crucial role in cellular dynamics by binding and disassembling actin filaments, impacting processes like axon guidance, cytokinesis, and cell morphology. Their cellular activity is tightly controlled, as dysregulation can lead to detrimental effects on cellular morphology. Although previous studies have suggested that MICALs are autoinhibited, and require Rab proteins to become active, the detailed molecular mechanisms remained unclear. Here, we report the cryo-EM structure of human MICAL1 at a nominal resolution of 3.1 Å. Structural analyses, alongside biochemical and functional studies, show that MICAL1 autoinhibition is mediated by an intramolecular interaction between its N-terminal catalytic and C-terminal coiled-coil domains, blocking F-actin interaction. Moreover, we demonstrate that allosteric changes in the coiled-coil domain and the binding of the tripartite assembly of CH-L2α1-LIM domains to the coiled-coil domain are crucial for MICAL activation and autoinhibition. These mechanisms appear to be evolutionarily conserved, suggesting a potential universality across the MICAL family.
- MeSH
- aktiny metabolismus chemie MeSH
- alosterická regulace MeSH
- calponiny MeSH
- elektronová kryomikroskopie * MeSH
- lidé MeSH
- mikrofilamenta metabolismus ultrastruktura MeSH
- mikrofilamentové proteiny metabolismus chemie ultrastruktura MeSH
- molekulární modely MeSH
- oxygenasy se smíšenou funkcí MeSH
- proteinové domény MeSH
- proteiny s doménou LIM metabolismus chemie genetika MeSH
- vazba proteinů * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Lens epithelium-derived growth factor p75 (LEDGF/p75), member of the hepatoma-derived growth-factor-related protein (HRP) family, is a transcriptional co-activator and involved in several pathologies including HIV infection and malignancies such as MLL-rearranged leukemia. LEDGF/p75 acts by tethering proteins to the chromatin through its integrase binding domain. This chromatin interaction occurs between the PWWP domain of LEDGF/p75 and nucleosomes carrying a di- or trimethylation mark on histone H3 Lys36 (H3K36me2/3). Our aim is to rationally devise small molecule drugs capable of inhibiting such interaction. To bootstrap this development, we resorted to X-ray crystallography-based fragment screening (FBS-X). Given that the LEDGF PWWP domain crystals were not suitable for FBS-X, we employed crystals of the closely related PWWP domain of paralog HRP-2. As a result, as many as 68 diverse fragment hits were identified, providing a detailed sampling of the H3K36me2/3 pocket pharmacophore. Subsequent structure-guided fragment expansion in three directions yielded multiple compound series binding to the pocket, as verified through X-ray crystallography, nuclear magnetic resonance and differential scanning fluorimetry. Our best compounds have double-digit micromolar affinity and optimally sample the interactions available in the pocket, judging by the Kd-based ligand efficiency exceeding 0.5 kcal/mol per non-hydrogen atom. Beyond π-stacking within the aromatic cage of the pocket and hydrogen bonding, the best compounds engage in a σ-hole interaction between a halogen atom and a conserved water buried deep in the pocket. Notably, the binding pocket in LEDGF PWWP is considerably smaller compared to the related PWWP1 domains of NSD2 and NSD3 which feature an additional subpocket and for which nanomolar affinity compounds have been developed recently. The absence of this subpocket in LEDGF PWWP limits the attainable affinity. Additionally, these structural differences in the H3K36me2/3 pocket across the PWWP domain family translate into a distinct selectivity of the compounds we developed. Our top-ranked compounds are interacting with both homologous LEDGF and HRP-2 PWWP domains, yet they showed no affinity for the NSD2 PWWP1 and BRPF2 PWWP domains which belong to other PWWP domain subfamilies. Nevertheless, our developed compound series provide a strong foundation for future drug discovery targeting the LEDGF PWWP domain as they can further be explored through combinatorial chemistry. Given that the affinity of H3K36me2/3 nucleosomes to LEDGF/p75 is driven by interactions within the pocket as well as with the DNA-binding residues, we suggest that future compound development should target the latter region as well. Beyond drug discovery, our compounds can be employed to devise tool compounds to investigate the mechanism of LEDGF/p75 in epigenetic regulation.
- MeSH
- knihovny malých molekul chemie farmakologie chemická syntéza MeSH
- krystalografie rentgenová MeSH
- lidé MeSH
- mezibuněčné signální peptidy a proteiny metabolismus chemie MeSH
- molekulární modely MeSH
- molekulární struktura MeSH
- proteinové domény MeSH
- racionální návrh léčiv * MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Adar null mutant mouse embryos die with aberrant double-stranded RNA (dsRNA)-driven interferon induction, and Adar Mavs double mutants, in which interferon induction is prevented, die soon after birth. Protein kinase R (Pkr) is aberrantly activated in Adar Mavs mouse pup intestines before death, intestinal crypt cells die, and intestinal villi are lost. Adar Mavs Eifak2 (Pkr) triple mutant mice rescue all defects and have long-term survival. Adenosine deaminase acting on RNA 1 (ADAR1) and PKR co-immunoprecipitate from cells, suggesting PKR inhibition by direct interaction. AlphaFold studies on an inhibitory PKR dsRNA binding domain (dsRBD)-kinase domain interaction before dsRNA binding and on an inhibitory ADAR1 dsRBD3-PKR kinase domain interaction on dsRNA provide a testable model of the inhibition. Wild-type or editing-inactive human ADAR1 expressed in A549 cells inhibits activation of endogenous PKR. ADAR1 dsRNA binding is required for, but is not sufficient for, PKR inhibition. Mutating the ADAR1 dsRBD3-PKR contact prevents co-immunoprecipitation, ADAR1 inhibition of PKR activity, and co-localization of ADAR1 and PKR in cells.
- MeSH
- adenosindeaminasa * metabolismus genetika MeSH
- aktivace enzymů MeSH
- buňky A549 MeSH
- dvouvláknová RNA * metabolismus MeSH
- kinasa eIF-2 * metabolismus MeSH
- lidé MeSH
- myši MeSH
- proteinové domény MeSH
- proteiny vázající RNA * metabolismus genetika MeSH
- vazba proteinů MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
AMPA glutamate receptors (AMPARs) are ion channel tetramers that mediate the majority of fast excitatory synaptic transmission. They are composed of four subunits (GluA1-GluA4); the GluA2 subunit dominates AMPAR function throughout the forebrain. Its extracellular N-terminal domain (NTD) determines receptor localization at the synapse, ensuring reliable synaptic transmission and plasticity. This synaptic anchoring function requires a compact NTD tier, stabilized by a GluA2-specific NTD interface. Here we show that low pH conditions, which accompany synaptic activity, rupture this interface. All-atom molecular dynamics simulations reveal that protonation of an interfacial histidine residue (H208) centrally contributes to NTD rearrangement. Moreover, in stark contrast to their canonical compact arrangement at neutral pH, GluA2 cryo-electron microscopy structures exhibit a wide spectrum of NTD conformations under acidic conditions. We show that the consequences of this pH-dependent conformational control are twofold: rupture of the NTD tier slows recovery from desensitized states and increases receptor mobility at mouse hippocampal synapses. Therefore, a proton-triggered NTD switch will shape both AMPAR location and kinetics, thereby impacting synaptic signal transmission.
- MeSH
- AMPA receptory * metabolismus chemie MeSH
- elektronová kryomikroskopie * MeSH
- hipokampus metabolismus MeSH
- kinetika MeSH
- koncentrace vodíkových iontů MeSH
- konformace proteinů MeSH
- lidé MeSH
- myši MeSH
- nervový přenos MeSH
- proteinové domény MeSH
- protony * MeSH
- simulace molekulární dynamiky * MeSH
- synapse * metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
This study investigates the factors modulating the reactivity of 5'-deoxyadenosyl (5'dAdo ̇) radical, a potent hydrogen atom abstractor that forms in the active sites of radical SAM enzymes and that otherwise undergoes a rapid self-decay in aqueous solution. Here, we compare hydrogen atom abstraction (HAA) reactions between native substrates of radical SAM enzymes and 5'dAdo ̇ in aqueous solution and in two enzymatic microenvironments. With that we reveal that HAA efficiency of 5'dAdo ̇ is due to (i) the in situ formation of 5'dAdo ̇ in a pre-ordered complex with a substrate, which attenuates the unfavorable effect of substrate:5'dAdo ̇ complex formation, and (ii) the prevention of the conformational changes associated with self-decay by a tight active-site cavity. The enzymatic cavity, however, does not have a strong effect on the HAA activity of 5'dAdo ̇. Thus, we performed an analysis of in-water HAA performed by 5'dAdo ̇ based on a three-component thermodynamic model incorporating the diagonal effect of the free energy of reaction, and the off-diagonal effect of asynchronicity and frustration. To this aim, we took advantage of the straightforward relationship between the off-diagonal thermodynamic effects and the electronic-structure descriptor - the redistribution of charge between the reactants during the reaction. It allows to access HAA-competent redox and acidobasic properties of 5'dAdo ̇ that are otherwise unavailable due to its instability upon one-electron reduction and protonation. The results show that all reactions feature a favourable thermodynamic driving force and tunneling, the latter of which lowers systematically barriers by ∼2 kcal mol-1. In addition, most of the reactions experience a favourable off-diagonal thermodynamic contribution. In HAA reactions, 5'dAdo ̇ acts as a weak oxidant as well as a base, also 5'dAdo ̇-promoted HAA reactions proceed with a quite low degree of asynchronicity of proton and electron transfer. Finally, the study elucidates the crucial and dual role of asynchronicity. It directly lowers the barrier as a part of the off-diagonal thermodynamic contribution, but also indirectly increases the non-thermodynamic part of the barrier by presumably controlling the adiabatic coupling between proton and electron transfer. The latter signals that the reaction proceeds as a hydrogen atom transfer rather than a proton-coupled electron transfer.
Isoforms of microtubule-associated protein 2 (MAP2) differ from their homolog Tau in the sequence and interactions of the N-terminal region. Binding of the N-terminal region of MAP2c (N-MAP2c) to the dimerization/docking domains of the regulatory subunit RIIα of cAMP-dependent protein kinase (RIIDD2) and to the Src-homology domain 2 (SH2) of growth factor receptor-bound protein 2 (Grb2) have been described long time ago. However, the structural features of the complexes remained unknown due to the disordered nature of MAP2. Here, we provide structural description of the complexes. We have solved solution structure of N-MAP2c in complex with RIIDD2, confirming formation of an amphiphilic α-helix of MAP2c upon binding, defining orientation of the α-helix in the complex and showing that its binding register differs from previous predictions. Using chemical shift mapping, we characterized the binding interface of SH2-Grb2 and rat MAP2c phosphorylated by the tyrosine kinase Fyn in their complex and proposed a model explaining differences between SH2-Grb2 complexes with rat MAP2c and phosphopeptides with a Grb2-specific sequence. The results provide the structural basis of a potential role of MAP2 in regulating cAMP-dependent phosphorylation cascade via interactions with RIIDD2 and Ras signaling pathway via interactions with SH2-Grb2.
- MeSH
- adaptorový protein Grb2 * metabolismus chemie MeSH
- lidé MeSH
- proteinové domény MeSH
- proteiny asociované s mikrotubuly * metabolismus chemie genetika MeSH
- protoonkogenní proteiny c-fyn metabolismus chemie genetika MeSH
- signální transdukce MeSH
- src homologní domény MeSH
- vazba proteinů * MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
YKL-40, also known as human cartilage glycoprotein-39 (HC-gp39) or CHI3L1, shares structural similarities with chitotriosidase (CHIT1), an active chitinase, but lacks chitinase activity. Despite being a biomarker for inflammatory disorders and cancer, the reasons for YKL-40's inert chitinase function have remained elusive. This study reveals that the loss of chitinase activity in YKL-40 has risen from multiple sequence modifications influencing its chitin affinity. Contrary to the common belief associating the lack of chitinase activity with amino acid substitutions in the catalytic motif, attempts to activate YKL-40 by creating two amino acid mutations in the catalytic motif (MT-YKL-40) proved ineffective. Subsequent exploration that included creating chimeras of MT-YKL-40 and CHIT1 catalytic domains (CatDs) identified key exons responsible for YKL-40 inactivation. Introducing YKL-40 exons 3, 6, or 8 into CHIT1 CatD resulted in chitinase inactivation. Conversely, incorporating CHIT1 exons 3, 6, and 8 into MT-YKL-40 led to its activation. Our recombinant proteins exhibited properly formed disulfide bonds, affirming a defined structure in active molecules. Biochemical and evolutionary analysis indicated that the reduced chitinase activity of MT-YKL-40 correlates with specific amino acids in exon 3. M61I and T69W substitutions in CHIT1 CatD diminished chitinase activity and increased chitin binding. Conversely, substituting I61 with M and W69 with T in MT-YKL-40 triggered chitinase activity while reducing the chitin-binding activity. Thus, W69 plays a crucial role in a unique subsite within YKL-40. These findings emphasize that YKL-40, though retaining the structural framework of a mammalian chitinase, has evolved to recognize chitin while surrendering chitinase activity.
- MeSH
- chitin * metabolismus chemie MeSH
- chitinasy metabolismus genetika chemie MeSH
- exony MeSH
- hexosaminidasy metabolismus chemie genetika MeSH
- katalytická doména MeSH
- lidé MeSH
- molekulární evoluce MeSH
- protein CHI3L1 * metabolismus genetika chemie MeSH
- sekvence aminokyselin MeSH
- substituce aminokyselin MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Phosphorylation plays a crucial role in the regulation of many fundamental cellular processes. Phosphorylation levels are increased in many cancer cells where they may promote changes in mitochondrial homeostasis. Proteomic studies on various types of cancer identified 17 phosphorylation sites within the human ATP-dependent protease Lon, which degrades misfolded, unassembled and oxidatively damaged proteins in mitochondria. Most of these sites were found in Lon's N-terminal (NTD) and ATPase domains, though little is known about the effects on their function. By combining the biochemical and cryo-electron microscopy studies, we show the effect of Tyr186 and Tyr394 phosphorylations in Lon's NTD, which greatly reduce all Lon activities without affecting its ability to bind substrates or perturbing its tertiary structure. A substantial reduction in Lon's activities is also observed in the presence of polyphosphate, whose amount significantly increases in cancer cells. Our study thus provides an insight into the possible fine-tuning of Lon activities in human diseases, which highlights Lon's importance in maintaining proteostasis in mitochondria.
- MeSH
- elektronová kryomikroskopie MeSH
- fosforylace MeSH
- lidé MeSH
- mitochondrie * metabolismus MeSH
- polyfosfáty * metabolismus MeSH
- proteasa La * metabolismus MeSH
- proteinové domény MeSH
- tyrosin * metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
