Spermine and spermidine SI-PPCs: Molecular dynamics reveals enhanced biomolecular interactions.

In this paper the effects on the interaction of highly positively charged substitution-inert platinum polynuclear complexes (SI-PPCs) with negatively charged DNA and heparin are examined and compared by theoretical chemistry methods. Electrostatic and hydrogen bonding interactions contribute to the overall effects on the biomolecule. Root Mean Square (RMS) deviation, Solvent Accessible Surface, RMS fluctuation, and interaction analysis all confirm similar effects on both biomolecules, dictated predominantly by the total positive charge and total number of hydrogen bonds formed. Especially, changes in structural parameters suggesting condensation and reduction of available surface area will reduce or prevent normal protein recognition and may thus potentially inhibit biological mechanisms related to apoptosis (DNA) or reduced vascularization viability (HEP). Thermodynamic analyses supported these findings with favourable interaction energies. The comparison of DNA and heparin confirms the general intersectionality between the two biomolecules and confirms the intrinsic dual-nature function of this chemotype. The distinction between the two-limiting mode of actions (HS or DNA-centred) could reflect an intriguing balance between extracellular (GAG) and intracellular (DNA) binding and affinities. The results underline the need to fully understand GAG-small molecule interactions and their contribution to drug pharmacology and related therapeutic modalities. This report contributes to that understanding.

Phenanthroline and phenyl carboxylate mixed ligand copper complexes in developing drugs to treat cancer.

The success of a classic inorganic coordination compound, Cisplatin, cis-[Pt(NH3)2Cl2], as the first anticancer metallodrug started a field of research dedicated to discovering coordination compounds with antitumor activity, encompassing various metals. Among these, copper complexes have emerged as interesting candidates to develop drugs to treat cancer. In this work, mixed ligand complexes of Cu(II) with diimines (phenanthroline or 4-methylphenanthroline) and 3-(4-hydroxyphenyl)propanoate, phenylcarboxylate or phenylacetate were synthesized. They were characterized in the solid state, including a new crystal structure of [Cu2(3-(4-hydroxyphenyl)propanoate)3(phenanthroline)2]Cl·H2O. The obtained complexes presented a variety of stoichiometries. In solution, complexes were partially dissociated in the corresponding Cu-diimine complex. The complexes bound to the DNA by partial intercalation and groove binding, as assessed by Circular Dichroism, relative viscosity change and UV-Vis titration. The cytotoxicity of the complexes was determined in vitro on MDA-MB-231, MCF-7 (human metastatic breast adenocarcinomas, the first triple negative), MCF-10A (breast nontumoral), A549 (human lung epithelial carcinoma), and MRC-5 (human nontumoral lung epithelial cells), finding an activity higher than that of Cisplatin, although with less selectivity.

Dissecting the structural and functional consequences of the evolutionary proline-glycine deletion in the wing 1 region of the forkhead domain of human FoxP1.

The human FoxP transcription factors dimerize via three-dimensional domain swapping, a unique feature among the human Fox family, as result of evolutionary sequence adaptations in the forkhead domain. This is the case for the conserved glycine and proline residues in the wing 1 region, which are absent in FoxP proteins but present in most of the Fox family. In this work, we engineered both glycine (G) and proline-glycine (PG) insertion mutants to evaluate the deletion events in FoxP proteins in their dimerization, stability, flexibility, and DNA-binding ability. We show that the PG insertion only increases protein stability, whereas the single glycine insertion decreases the association rate and protein stability and promotes affinity to the DNA ligand.

Exploring the BSA- and DNA-binding, cytotoxicity, and cell cycle evaluation of ternary copper(II)/diimine complexes with N,N-dibenzyl-N'-benzoylthiourea as promising metallodrug candidates.

Four new copper(II) complexes were synthesized and characterized with the general formula [Cu(N-N)(Th)(NO3)], where N-N corresponds to the N-heterocyclic ligands 1,10-phenanthroline (phen), 2,2'-bipyridine (bipy), 4,7-diphenyl-1,10-phenanthroline (dpp), and 4,4-dimethyl-2,2'-bipyridine (dmbp) and Th represents the N,N-dibenzyl-N'-benzoylthiourea. Cytotoxic activities of the complexes against HCT116 (human colon carcinoma), HepG2 (human hepatocellular carcinoma), and non-tumor MRC-5 (human lung fibroblast) cells were investigated. The copper(II) complexes 1-4 were characterized by spectroscopic techniques while complexes 1 and 2 were studied using single-crystal X-ray diffraction as well. The complexes possessed a five-coordinated structure with one nitrate ligand as a monodentate at the axial position and two bidentate ligands N-heterocyclic and N,N-dibenzyl-N'-benzoylthiourea. The complexes showed promising IC50 values, ranging from 0.3 to 9.0 µM. Furthermore, interaction studies with biomolecules such as calf thymus DNA (ct-DNA) and Bovine Serum Albumin (BSA), which can act as possible biological targets of the complexes, were carried out. The studies suggested that the compounds interact moderately with ct-DNA and BSA. Complexes 1, 2, and 4 did not lead to cell accumulation at any stage of the cell cycle but caused a significant increase in internucleosomal DNA fragmentation. Whereas, compound 3 caused cell cycle arrest in the S phase while doxorubicin caused cell cycle arrest in the G2/M phase. The effect of structural modifications on the metal compounds was correlated with their biological properties and it was concluded that an increase in biological activity occurred with increasing the extension of the diimine ligands. Thus, complex 3 was the most promising one.

The impact of biomolecule interactions on the cytotoxic effects of rhenium(I) tricarbonyl complexes.

Rhenium complexes show great promise as anticancer drug candidates. Specifically, compounds with a Re(CO)3(NN)(py)+ core in their architecture have shown cytotoxicity equal to or greater than that of well-established anticancer drugs based on platinum or organic molecules. This study aimed to evaluate how the strength of the interaction between rhenium(I) tricarbonyl complexes fac-[Re(CO)3(NN)(py)]+, NN = 1,10-phenanthroline (phen), dipyrido[3,2-f:2',3'-h]quinoxaline (dpq) or dipyrido[3,2-a:2'3'-c]phenazine (dppz) and biomolecules (protein, lipid and DNA) impacted the corresponding cytotoxic effect in cells. Results showed that fac-[Re(CO)3(dppz)(py)]+ has higher Log Po/w and binding constant (Kb) with biomolecules (protein, lipid and DNA) compared to complexes of fac-[Re(CO)3(phen)(py)]+ and fac-[Re(CO)3(dpq)(py)]+. As consequence, fac-[Re(CO)3(dppz)(py)]+ exhibited the highest cytotoxicity (IC50 = 8.5 µM for HeLa cells) for fac-[Re(CO)3(dppz)(py)]+ among the studied compounds (IC50 > 15 µM). This highest cytotoxicity of fac-[Re(CO)3(dppz)(py)]+ are probably related to its lipophilicity, higher permeation of the lipid bilayers of cells, and a more potent interaction of the dppz ligand with biomolecules (protein and DNA). Our findings open novel avenues for rational drug design and highlight the importance of considering the chemical structures of rhenium complexes that strongly interact with biomolecules (proteins, lipids, and DNA).

Influence of diimine bidentate ligand in the nitrosyl and nitro terpyridine ruthenium complex on the HSA/DNA interaction and antiviral activity.

Nitric oxide (NO) acts in different physiological processes, such as blood pressure control, antiparasitic activities, neurotransmission, and antitumor action. Among the exogenous NO donors, ruthenium nitrosyl/nitro complexes are potential candidates for prodrugs, due to their physicochemical properties, such as thermal and physiological pH stability. In this work, we proposed the synthesis and physical characterization of the new nitro terpyridine ruthenium (II) complexes of the type [RuII(L)(NO2)(tpy)]PF6 where tpy = 2,2':6',2″-terpyridine; L = 3,4-diaminobenzoic acid (bdq) or o-phenylenediamine (bd) and evaluation of influence of diimine bidentate ligand NH.NHq-R (R = H or COOH) in the HSA/DNA interaction as well as antiviral activity. The interactions between HSA and new nitro complexes [RuII(L)(NO2)(tpy)]+ were evaluated. The Ka values for the HSA-[RuII(bdq)(NO2)(tpy)]+ is 10 times bigger than HSA-[RuII(bd)(NO2)(tpy)]+. The sites of interaction between HSA and the complexes via synchronous fluorescence suppression indicate that the [RuII(bdq)(NO2)(tpy)]+ is found close to the Trp-241 residue, while the [RuII(bd)(NO2)(tpy)]+ complex is close to Tyr residues. The interaction with fish sperm fs-DNA using direct spectrophotometric titration (Kb) and ethidium bromide replacement (KSV and Kapp) showed weak interaction in the system fs-DNA-[RuII(bdq)(NO)(tpy)]+. Furthermore, fs-DNA-[RuII(bd)(NO2)(tpy)]+ and fs-DNA-[RuII(bd)(NO)(tpy)]3+ system showed higher intercalation constant. Circular dichroism spectra for fs-DNA-[RuII(bd)(NO2)(tpy)]+ and fs-DNA-[RuII(bd)(NO)(tpy)]3+, suggest semi-intercalative accompanied by major groove binding interaction modes. The [RuII(bd)(NO2)(tpy)]+ and [RuII(bd)(NO)(tpy)]3+ inhibit replication of Zika and Chikungunya viruses based in the nitric oxide release under S-nitrosylation reaction with cysteine viral.

Vanadium(V) complexes derived from triphenylphosphonium and hydrazides: cytotoxicity evaluation and interaction with biomolecules.

The development of coordination compounds with antineoplastic therapeutic properties is currently focused on non-covalent interactions with deoxyribonucleic acid (DNA). Additionally, the interaction profiles of these compounds with globular plasma proteins, particularly serum albumin, warrant thorough evaluation. In this study, we report on the interactions between biomolecules and complexes featuring hydrazone-type imine ligands coordinated with vanadium. The potential to enhance the therapeutic efficiency of these compounds through mitochondrial targeting is explored. This targeting is facilitated by the derivatization of ligands with triphenylphosphonium groups. Thus, this work presents the synthesis, characterization, interactions, and cytotoxicity of dioxidovanadium(V) complexes (C1-C5) with a triphenylphosphonium moiety. These VV-species are coordinated to hydrazone-type iminic ligands derived from (3-formyl-4-hydroxybenzyl)triphenylphosphonium chloride ([AH]Cl) and aromatic hydrazides ([H2L1]Cl-[H2L5]Cl). The structures of the five complexes were elucidated through single-crystal X-ray diffraction and vibrational spectroscopies, confirming the presence of dioxidovanadium(V) species in various geometries with degrees of distortion (τ = 0.03-0.50) and highlighting their zwitterionic characteristics. The molecular structural stability of C1-C5 in solution was ascertained using 1H, 19F, 31P, and 51V-nuclear magnetic resonance. Moreover, their interactions with biomolecules were evaluated using diverse spectroscopic methodologies and molecular docking, indicating moderate interactions (Kb ≈ 104 M-1) with calf thymus DNA in the minor groove and with human serum albumin, predominantly in the superficial IB subdomain. Lastly, the cytotoxic potentials of these complexes were assessed in keratinocytes of the HaCaT lineage, revealing that C1-C5 induce a reduction in metabolic activity and cell viability through apoptotic pathways.

The transcription of the main gene associated with Treacher-Collins syndrome (TCOF1) is regulated by G-quadruplexes and cellular nucleic acid binding protein (CNBP).

Treacle ribosome biogenesis factor 1 (TCOF1) is responsible for about 80% of mandibular dysostosis (MD) cases. We have formerly identified a correlation between TCOF1 and CNBP (CCHC-type zinc finger nucleic acid binding protein) expression in human mesenchymal cells. Given the established role of CNBP in gene regulation during rostral development, we explored the potential for CNBP to modulate TCOF1 transcription. Computational analysis for CNBP binding sites (CNBP-BSs) in the TCOF1 promoter revealed several putative binding sites, two of which (Hs791 and Hs2160) overlap with putative G-quadruplex (G4) sequences (PQSs). We validated the folding of these PQSs measuring circular dichroism and fluorescence of appropriate synthetic oligonucleotides. In vitro studies confirmed binding of purified CNBP to the target PQSs (both folded as G4 and unfolded) with Kd values in the nM range. ChIP assays conducted in HeLa cells chromatin detected the CNBP binding to TCOF1 promoter. Transient transfections of HEK293 cells revealed that Hs2160 cloned upstream SV40 promoter increased transcription of downstream firefly luciferase reporter gene. We also detected a CNBP-BS and PQS (Dr2393) in the zebrafish TCOF1 orthologue promoter (nolc1). Disrupting this G4 in zebrafish embryos by microinjecting DNA antisense oligonucleotides complementary to Dr2393 reduced the transcription of nolc1 and recapitulated the craniofacial anomalies characteristic of Treacher Collins Syndrome. Both cnbp overexpression and Morpholino-mediated knockdown in zebrafish induced nolc1 transcription. These results suggest that CNBP modulates the transcriptional expression of TCOF1 through a mechanism involving G-quadruplex folding/unfolding, and that this regulation is active in vertebrates as distantly related as bony fish and humans. These findings may have implications for understanding and treating MD.

The interaction of InvF-RNAP is mediated by the chaperone SicA in Salmonella sp: an in silico prediction.

In this work we carried out an in silico analysis to understand the interaction between InvF-SicA and RNAP in the bacterium Salmonella Typhimurium strain LT2. Structural analysis of InvF allowed the identification of three possible potential cavities for interaction with SicA. This interaction could occur with the structural motif known as tetratricopeptide repeat (TPR) 1 and 2 in the two cavities located in the interface of the InvF and α-CTD of RNAP. Indeed, molecular dynamics simulations showed that SicA stabilizes the Helix-turn-Helix DNA-binding motifs, i.e., maintaining their proper conformation, mainly in the DNA Binding Domain (DBD). Finally, to evaluate the role of amino acids that contribute to protein-protein affinity, an alanine scanning mutagenesis approach, indicated that R177 and R181, located in the DBD motif, caused the greatest changes in binding affinity with α-CTD, suggesting a central role in the stabilization of the complex. However, it seems that the N-terminal region also plays a key role in the protein-protein interaction, especially the amino acid R40, since we observed conformational flexibility in this region allowing it to interact with interface residues. We consider that this analysis opens the possibility to validate experimentally the amino acids involved in protein-protein interactions and explore other regulatory complexes where chaperones are involved.

Importancia biológica de los telómeros / Biological importance of telomeres

Se exponen los hallazgos históricos y la importancia biológica de los telómeros en la vida celular y en los aspectos genéticos del ADN humano. (AU)

The discovery and the biological importance of the telomeres are exposed. (AU)

Non-biofilm-forming Staphylococcus epidermidis planktonic cell supernatant induces alterations in osteoblast biological function.

Staphylococcal biofilms significantly contribute to prosthetic joint infection (PJI). However, 40% of S. epidermidis PJI isolates do not produce biofilms, which does not explain the role of biofilms in these cases. We studied whether the supernatant from planktonic S. epidermidis alters osteoblast function. Non-biofilm-forming S. epidermidis supernatants (PJI- clinical isolate, healthy skin isolate (HS), and ATCC12228 reference strain) and biofilm-forming supernatants (PJI+ clinical isolate, ATCC35984 reference strain, and Staphylococcus aureus USA300 reference strain) were included. Osteoblasts stimulated with supernatants from non-biofilm-forming isolates for 3, 7, and 14 days showed significantly reduced cellular DNA content compared with unstimulated osteoblasts, and apoptosis was induced in these osteoblasts. Similar results were obtained for biofilm-forming isolates, but with a greater reduction in DNA content and higher apoptosis. Alkaline phosphatase activity and mineralization were significantly reduced in osteoblasts treated with supernatants from non-biofilm-forming isolates compared to the control at the same time points. However, the supernatants from biofilm-forming isolates had a greater effect than those from non-biofilm-forming isolates. A significant decrease in the expression of ATF4, RUNX2, ALP, SPARC, and BGLAP, and a significant increase in RANK-L expression were observed in osteoblasts treated with both supernatants. These results demonstrate that the supernatants of the S. epidermidis isolate from the PJI- and HS (commensal) with a non-biofilm-forming phenotype alter the function of osteoblasts (apoptosis induction, failure of cell differentiation, activation of osteoblasts, and induction of bone resorption), similar to biofilm-forming isolates (PJI+, ATCC35984, and S. aureus USA300), suggesting that biofilm status contributes to impaired osteoblast function and that the planktonic state can do so independently of biofilm production.

Reduced protein intake and aging affects the sustainment of hematopoiesis by impairing bone marrow mesenchymal stem cells.

Protein malnourishment (PM) is common among the elderly, but how aging and PM impact hematopoiesis is not fully understood. This study aimed to assess how aging and PM affect the hematopoietic regulatory function of bone marrow (BM) mesenchymal stem cells (MSCs). Young and aged male C57BL/6J mice were fed with normoproteic or hypoproteic diets and had their nutritional, biochemical, and hematological parameters evaluated. BM MSCs were characterized and had their secretome, gene expression, autophagy, reactive oxygen species production (ROS), and DNA double-stranded breaks evaluated. The modulation of hematopoiesis by MSCs was assayed using in vitro and in vivo models. Lastly, BM invasiveness and mice survival were evaluated after being challenged with leukemic cells of the C1498 cell line. Aging and PM alter biochemical parameters, changing the peripheral blood and BM immunophenotype. MSC autophagy was affected by aging and the frequencies for ROS and DNA double-stranded breaks. Regarding the MSCs' secretome, PM and aging affected CXCL12, IL-6, and IL-11 production. Aging and PM up-regulated Akt1 and PPAR-γ while down-regulating Cdh2 and Angpt-1 in MSCs. Aged MSCs increased C1498 cell proliferation while reducing their colony-forming potential. PM and aging lowered mice survival, and malnourishment accumulated C1498 cells at the BM. Finally, aged and/or PM MSCs up-regulated Sox2, Nanog, Pou5f1, and Akt1 expression while down-regulating Cdkn1a in C1498 cells. Together, aging and PM can induce cell-intrinsic shifts in BM MSCs, creating an environment that alters the regulation of hematopoietic populations and favoring the development of malignant cells.

APGW/AKH Precursor from Rotifer Brachionus plicatilis and the DNA Loss Model Explain Evolutionary Trends of the Neuropeptide LWamide, APGWamide, RPCH, AKH, ACP, CRZ, and GnRH Families.

In the year 2002, DNA loss model (DNA-LM) postulated that neuropeptide genes to emerged through codons loss via the repair of damaged DNA from ancestral gene namely Neuropeptide Precursor Predictive (NPP), which organization correspond two or more neuropeptides precursors evolutive related. The DNA-LM was elaborated according to amino acids homology among LWamide, APGWamide, red pigment-concentrating hormone (RPCH), adipokinetic hormones (AKHs) and in silico APGW/RPCH NPPAPGW/AKH NPP were proposed. With the above principle, it was proposed the evolution of corazonin (CRZ), gonadotropin-releasing hormone (GnRH), AKH, and AKH/CRZ (ACP), but any NPP never was considered. However, the evolutive relation via DNA-LM among these neuropeptides precursors not has been established yet. Therefore, the transcriptomes from crabs Callinectes toxotes and Callinectes arcuatus were used to characterized ACP and partial CRZ precursors, respectively. BLAST alignment with APGW/RPCH NPP and APGW/AKH NPP allow identified similar NPP in the rotifer Brachionus plicatilis and other invertebrates. Moreover, three bioinformatics algorithms and manual verification were used to purify 13,778 sequences, generating a database with 719 neuropeptide precursors. Phylogenetic trees with the DNA-LM parameters showed that some ACP, CRZ, AKH2 and two NPP share nodes with GnRH from vertebrates and some of this neuropeptide had nodes in invertebrates. Whereas the phylogenetic tree with standard parameters do not showed previous node pattern. Robinson-Foulds metric corroborates the differences among phylogenetic trees. Homology relationship showed four putative orthogroups; AKH4, CRZ, and protostomes GnRH had individual group. This is the first demonstration of NPP in species and would explain the evolution neuropeptide families by the DNA-LM.

Development of synthetic modulator enabling long-term propagation and neurogenesis of human embryonic stem cell-derived neural progenitor cells.

Neural progenitor cells (NPCs) are essential for in vitro drug screening and cell-based therapies for brain-related disorders, necessitating well-defined and reproducible culture systems. Current strategies employing protein growth factors pose challenges in terms of both reproducibility and cost. In this study, we developed a novel DNA-based modulator to regulate FGFR signaling in NPCs, thereby facilitating the long-term maintenance of stemness and promoting neurogenesis. This DNA-based FGFR-agonist effectively stimulated FGFR1 phosphorylation and activated the downstream ERK signaling pathway in human embryonic stem cell (HESC)-derived NPCs. We replaced the basic fibroblast growth factor (bFGF) in the culture medium with our DNA-based FGFR-agonist to artificially modulate FGFR signaling in NPCs. Utilizing a combination of cell experiments and bioinformatics analyses, we showed that our FGFR-agonist could enhance NPC proliferation, direct migration, and promote neurosphere formation, thus mimicking the functions of bFGF. Notably, transcriptomic analysis indicated that the FGFR-agonist could specifically influence the transcriptional program associated with stemness while maintaining the neuronal differentiation program, closely resembling the effects of bFGF. Furthermore, our culture conditions allowed for the successful propagation of NPCs through over 50 passages while retaining their ability to efficiently differentiate into neurons. Collectively, our approach offers a highly effective method for expanding NPCs, thereby providing new avenues for disease-in-dish research and drug screening aimed at combating neural degeneration.

An intracellular, non-oxidative factor activates in vitro chromatin fragmentation in pig sperm.

BACKGROUND: In vitro incubation of epididymal and vas deferens sperm with Mn2+ induces Sperm Chromatin Fragmentation (SCF), a mechanism that causes double-stranded breaks in toroid-linker regions (TLRs). Whether this mechanism, thought to require the participation of topoisomerases and/or DNAses and thus far only described in epididymal mouse sperm, can be triggered in ejaculated sperm is yet to be elucidated. The current study aimed to determine if exposure of pig ejaculated sperm to divalent ions (Mn2+ and Mg2+) activates SCF, and whether this has any impact on sperm function and survival. For this purpose, sperm DNA integrity was evaluated through the Comet assay and Pulsed Field Gel Electrophoresis (PFGE); sperm motility and agglutination were assessed with computer assisted sperm analysis (CASA); and sperm viability and levels of total reactive oxygen species (ROS) and superoxides were determined through flow cytometry. RESULTS: Incubation with Mn2+/Ca2+ activated SCF in a dose-dependent (P < 0.05) albeit not time-dependent manner (P > 0.05); in contrast, Mg2+/Ca2+ only triggered SCF at high concentrations (50 mM). The PFGE revealed that, when activated by Mn2+/Ca2+ or Mg2+/Ca2+, SCF generated DNA fragments of 33-194 Kb, compatible with the size of one or multiple toroids. Besides, Mn2+/Ca2+ affected sperm motility in a dose-dependent manner (P < 0.05), whereas Mg2+/Ca2+ only impaired this variable at high concentrations (P < 0.05). While this effect on motility was concomitant with an increase of agglutination, neither viability nor ROS levels were affected by Mn2+/Ca2+ or Mg2+/Ca2+ treatments. CONCLUSION: Mn2+/Ca2+ and Mn2+/Ca2+ were observed to induce SCF in ejaculated sperm, resulting in DNA cleavage at TLRs. The activation of this mechanism by an intracellular, non-oxidative factor sheds light on the events taking place during sperm cell death.

Balancing the functions of DNA extracellular traps in intracellular parasite infections: implications for host defense, disease pathology and therapy.

The release of DNA to the extracellular milieu is a biological process referred to as etosis, which is involved in both physiological and pathological functions. Although the release of DNA extracellular traps (ETs) was initially attributed to innate immune cells such as neutrophils, eosinophils, and macrophages, recent studies have shown that T cells, as well as non-immune cells, are capable of releasing ETs. These structures were described primarily for their potential to trap and kill pathogens, presenting an important strategy of host defense. Intriguingly, these functions have been associated with intracellular pathogens such as the parasites Leishmania sp. and Trypanosoma cruzi, causative agents of leishmaniasis and Chagas disease, respectively. These are two devastating tropical diseases that lead to thousands of deaths every year. In an apparent contradiction, ETs can also induce and amplify inflammation, which may lead to worsening disease pathology. This has prompted the concept of targeting ETs' release as a means of controlling tissue destruction to treat human diseases. What is the best approach to prevent disease severity: inducing ETs to kill pathogens or preventing their release? In this Perspective article, we will discuss the importance of understanding ETs released by different cell types and the need to balance their potentially complementary functions. In addition, we will explore other functions of ETs and their translational applications to benefit individuals infected with intracellular parasites and other pathogens. Ultimately, a better understanding of the role of ETs in disease pathogenesis will provide valuable insights into developing novel therapies for human diseases.

CTCF and Its Multi-Partner Network for Chromatin Regulation.

Architectural proteins are essential epigenetic regulators that play a critical role in organizing chromatin and controlling gene expression. CTCF (CCCTC-binding factor) is a key architectural protein responsible for maintaining the intricate 3D structure of chromatin. Because of its multivalent properties and plasticity to bind various sequences, CTCF is similar to a Swiss knife for genome organization. Despite the importance of this protein, its mechanisms of action are not fully elucidated. It has been hypothesized that its versatility is achieved through interaction with multiple partners, forming a complex network that regulates chromatin folding within the nucleus. In this review, we delve into CTCF's interactions with other molecules involved in epigenetic processes, particularly histone and DNA demethylases, as well as several long non-coding RNAs (lncRNAs) that are able to recruit CTCF. Our review highlights the importance of CTCF partners to shed light on chromatin regulation and pave the way for future exploration of the mechanisms that enable the finely-tuned role of CTCF as a master regulator of chromatin.

The chromatin - triple helix connection.

Mammalian genomes are extensively transcribed, producing a large number of coding and non-coding transcripts. A large fraction of the nuclear RNAs is physically associated with chromatin, functioning in gene activation and silencing, shaping higher-order genome organisation, such as involvement in long-range enhancer-promoter interactions, transcription hubs, heterochromatin, nuclear bodies and phase transitions. Different mechanisms allow the tethering of these chromatin-associated RNAs (caRNA) to chromosomes, including RNA binding proteins, the RNA polymerases and R-loops. In this review, we focus on the sequence-specific targeting of RNA to DNA by forming triple helical structures and describe its interplay with chromatin. It turns out that nucleosome positioning at triple helix target sites and the nucleosome itself are essential factors in determining the formation and stability of triple helices. The histone H3-tail plays a critical role in triple helix stabilisation, and the role of its epigenetic modifications in this process is discussed.

The Arabidopsis DNA glycosylase MBD4L repairs the nuclear genome in vivo.

DNA glycosylases remove mispaired or modified bases from DNA initiating the base excision repair (BER) pathway. The DNA glycosylase MBD4 (methyl-CpG-binding domain protein 4) has been functionally characterized in mammals, but not yet in plants, where it is called MBD4-like (MBD4L). Mammalian MBD4 and Arabidopsis recombinant MBD4L excise U and T mispaired with G, as well as 5-fluorouracil (5-FU) and 5-bromouracil (5-BrU) in vitro. Here, we investigate the ability of Arabidopsis MBD4L to remove some of these substrates from the nuclear genome in vivo in coordination with uracil DNA glycosylase (AtUNG). We found that mbd4l mutants are hypersensitive to 5-FU and 5-BrU, as they displayed smaller size, less root growth, and higher cell death than control plants in both media. Using comet assays, we determined BER-associated DNA fragmentation in isolated nuclei and observed reduced DNA breaks in mbd4l plants under both conditions, but particularly with 5-BrU. The use of ung and ung x mbd4l mutants in these assays indicated that both MBD4L and AtUNG trigger nuclear DNA fragmentation in response to 5-FU. Consistently, we here report the nuclear localization of AtUNG based on the expression of AtUNG-GFP/RFP constructs in transgenic plants. Interestingly, MBD4L and AtUNG are transcriptionally coordinated but display not completely overlapping functions. MBD4L-deficient plants showed reduced expression of BER genes and enhanced expression of DNA damage response (DDR) gene markers. Overall, our findings indicate that Arabidopsis MBD4L is critical for maintaining nuclear genome integrity and preventing cell death under genotoxic stress conditions.

Pancreatic islet remodeling in cotadutide-treated obese mice.

Obesity and type 2 diabetes mellitus (T2DM) cause morphofunctional alterations in pancreatic islet alpha and beta cells. Therefore, we hypothesize that the new GLP-1/Glucagon receptor dual agonist cotadutide may benefit islet cell arrangement and function. Twelve-week-old C57BL/6 male mice were fed a control diet (C, 10 % kJ fat) or a high-fat diet (HF, 50 % kJ fat) for ten weeks. Then, the animals were divided into four groups for an additional 30 days and daily treated with subcutaneous cotadutide (30 nmol/kg) or vehicle: C, CC (control+cotadutide), HF, and HFC (high-fat+cotadutide). Cotadutide led to weight loss and reduced insulin resistance in the HFC group, increasing insulin receptor substrate 1 and solute carrier family 2 gene expressions in isolated islets. Also, cotadutide enhanced transcriptional factors related to islet cell transdifferentiation, decreasing aristaless-related homeobox and increasing the paired box 4 and 6, pancreatic and duodenal homeobox 1, v-maf musculoaponeurotic fibrosarcoma oncogene family protein A, neurogenin 3, and neurogenic differentiation 1. In addition, cotadutide improved the proliferating cell nuclear antigen, NK6 homeobox 1, B cell leukemia/lymphoma 2, but lessening caspase 3. Furthermore, cotadutide mitigated the endoplasmic reticulum (ER) stress-responsive genes, reducing transcription factor 4, DNA-damage-inducible transcript 3, and growth arrest and DNA-damage-inducible 45. In conclusion, our data demonstrated significant beneficial actions of cotadutide in DIO mice, such as weight loss, glycemic control, and insulin resistance improvement. In addition, cotadutide counteracted the pathological adaptive cellular arrangement of the pancreatic islet in obese mice, improving the markers of the transdifferentiating pathway, proliferation, apoptosis, and ER stress.