Intrinsic disorder is very important in the biological function of several proteins, and is directly linked to their foldability during interaction with their targets. There is a close relationship ...between the intrinsically disordered proteins and the process of carcinogenesis involving viral pathogens. Among these pathogens, we have highlighted the human papillomavirus (HPV) in this study. HPV is currently among the most common sexually transmitted infections, besides being the cause of several types of cancer. HPVs are divided into two groups, called high- and low-risk, based on their oncogenic potential. The high-risk HPV E6 protein has been the target of much research, in seeking treatments against HPV, due to its direct involvement in the process of cell cycle control. To understand the role of intrinsic disorder of the viral proteins in the oncogenic potential of different HPV types, the structural characteristics of intrinsically disordered regions of high and low-risk HPV E6 proteins were analyzed. In silico analyses of primary sequences, prediction of tertiary structures, and analyses of molecular dynamics allowed the observation of the behavior of such disordered regions in these proteins, thereby proving a direct relationship of structural variation with the degree of oncogenicity of HPVs. The results obtained may contribute to the development of new therapies, targeting the E6 oncoprotein, for the treatment of HPV-associated diseases.
Human Antigen Leukocyte-G (
) gene encodes an immune checkpoint molecule that has restricted tissue expression in physiological conditions; however, the gene may be induced in hypoxic conditions by ...the interaction with the hypoxia inducible factor-1 (HIF1). Hypoxia regulatory elements (HRE) located at the
promoter region and at exon 2 are the major HIF1 target sites. Since the G allele of the -964G > A transversion induces higher HLA-G expression when compared to the A allele in hypoxic conditions, here we analyzed HIF1-HRE complex interaction at the pair-atom level considering both -964G > A polymorphism alleles. Mouse HIF2 dimer crystal (Protein Data Bank ID: 4ZPK) was used as template to perform homology modelling of human HIF1 quaternary structure using MODELLER v9.14. Two 3D DNA structures were built from 5'GCRTG'3 HRE sequence containing the -964G/A alleles using x3DNA. Protein-DNA docking was performed using the HADDOCK v2.4 server, and non-covalent bonds were computed by DNAproDB server. Molecular dynamic simulation was carried out per 200 ns, using Gromacs v.2019. HIF1 binding in the HRE containing -964G allele results in more hydrogen bonds and van der Waals contact formation than HRE with -964A allele. Protein-DNA complex trajectory analysis revealed that HIF1-HRE-964G complex is more stable. In conclusion, HIF1 binds in a more stable and specific manner at the HRE with G allele.
Laccases are multicopper oxidases (MCOs) with a broad application spectrum, particularly in second-generation ethanol biotechnology and the bioremediation of xenobiotics and other highly recalcitrant ...compounds. Synthetic pesticides are xenobiotics with long environmental persistence, and the search for their effective bioremediation has mobilized the scientific community. Antibiotics, in turn, can pose severe risks for the emergence of multidrug-resistant microorganisms, as their frequent use for medical and veterinary purposes can generate constant selective pressure on the microbiota of urban and agricultural effluents. In the search for more efficient industrial processes, some bacterial laccases stand out for their tolerance to extreme physicochemical conditions and their fast generation cycles. Accordingly, to expand the range of effective approaches for the bioremediation of environmentally important compounds, the prospection of bacterial laccases was carried out from a custom genomic database. The best hit found in the genome of
sp. CB10, a Bacteroidetes isolate obtained from a biomass-degrading bacterial consortium, was subjected to in silico prediction, molecular docking, and molecular dynamics simulation analyses. The putative laccase CB10_180.4889 (Lac_CB10), composed of 728 amino acids, with theoretical molecular mass values of approximately 84 kDa and a pI of 6.51, was predicted to be a new CopA with three cupredoxin domains and four conserved motifs linking MCOs to copper sites that assist in catalytic reactions. Molecular docking studies revealed that Lac_CB10 had a high affinity for the molecules evaluated, and the affinity profiles with multiple catalytic pockets predicted the following order of decreasing thermodynamically favorable values: tetracycline (-8 kcal/mol) > ABTS (-6.9 kcal/mol) > sulfisoxazole (-6.7 kcal/mol) > benzidine (-6.4 kcal/mol) > trimethoprim (-6.1 kcal/mol) > 2,4-dichlorophenol (-5.9 kcal/mol) mol. Finally, the molecular dynamics analysis suggests that Lac_CB10 is more likely to be effective against sulfisoxazole-like compounds, as the sulfisoxazole-Lac_CB10 complex exhibited RMSD values lower than 0.2 nm, and sulfisoxazole remained bound to the binding site for the entire 100 ns evaluation period. These findings corroborate that LacCB10 has a high potential for the bioremediation of this molecule.
HLA-G is considered to be an immune checkpoint molecule, a function that is closely linked to the structure and dynamics of the different HLA-G isoforms. Unfortunately, little is known about the ...structure and dynamics of these isoforms. For instance, there are only seven crystal structures of HLA-G molecules, being all related to a single isoform, and in some cases lacking important residues associated to the interaction with leukocyte receptors. In addition, they lack information on the dynamics of both membrane-bound HLA-G forms, and soluble forms. We took advantage of
strategies to disclose the dynamic behavior of selected HLA-G forms, including the membrane-bound HLA-G1 molecule, soluble HLA-G1 dimer, and HLA-G5 isoform. Both the membrane-bound HLA-G1 molecule and the soluble HLA-G1 dimer were quite stable. Residues involved in the interaction with ILT2 and ILT4 receptors (α3 domain) were very close to the lipid bilayer in the complete HLA-G1 molecule, which might limit accessibility. On the other hand, these residues can be completely exposed in the soluble HLA-G1 dimer, due to the free rotation of the disulfide bridge (Cys42/Cys42). In fact, we speculate that this free rotation of each protomer (i.e., the chains composing the dimer) could enable alternative binding modes for ILT2/ILT4 receptors, which in turn could be associated with greater affinity of the soluble HLA-G1 dimer. Structural analysis of the HLA-G5 isoform demonstrated higher stability for the complex containing the peptide and coupled β2-microglobulin, while structures lacking such domains were significantly unstable. This study reports for the first time structural conformations for the HLA-G5 isoform and the dynamic behavior of HLA-G1 molecules under simulated biological conditions. All modeled structures were made available through GitHub (https://github.com/KavrakiLab/), enabling their use as templates for modeling other alleles and isoforms, as well as for other computational analyses to investigate key molecular interactions.
Objective To elucidate the potential mechanisms involved in the physiopathology of endometriosis. We analyzed the differential gene expression profiles of eutopic and ectopic tissues from women with ...endometriosis. Design Prospective laboratory study. Setting University hospital. Patient(s) Seventeen patients in whom endometriosis was diagnosed and 11 healthy fertile women. Intervention(s) Endometrial biopsy specimens from the endometrium of healthy women without endometriosis and from the eutopic and ectopic endometrium tissues of patients with endometriosis were obtained in the early proliferative phase of the menstrual cycle. Main Outcome Measure(s) Six paired samples of eutopic and ectopic tissue were analyzed by subtractive hybridization. To evaluate the expression of genes found by rapid subtraction hybridization methods, we measured CTGF, SPARC, MYC, MMP, and IGFBP1 genes by real-time polymerase chain reaction in all samples. Result(s) This study identified 291 deregulated genes in the endometriotic lesions. Significant expression differences were obtained for SPARC, MYC, and IGFBP1 in the peritoneal lesions and for MMP3 in the ovarian endometriomas. Additionally, significant differences were obtained for SPARC and IGFBP1 between the peritoneal and ovarian lesions. No significant differences were found for the studied genes between the control and the eutopic endometrium. Conclusion(s) This study identified 291 genes with differential expression in endometriotic lesions. The deregulation of the SPARC, MYC, MMP3, and IGFBPI genes may be responsible for the loss of cellular homeostasis in endometriotic lesions.
The function of medullary thymic epithelial cells (mTECs) is associated with thymocyte adhesion, which is crucial for the negative selection of autoreactive thymocytes in the thymus. This process ...represents the root of central tolerance of self-components and prevents the onset of autoimmune diseases. Since thymic epithelia correspond to an important target of donor T cells during the onset of chronic graft-vs-host-disease, mTEC-thymocyte adhesion may have implications for alloimmunity. The
and
genes function as transcriptome controllers in mTECs. The central question of this study is whether there is a mutual relationship between mTEC-thymocyte adhesion and the control of the mTEC transcriptome and whether Aire is involved in this process. Here, we show that
mTEC-thymocyte adhesion causes transcriptome changes in mTECs and upregulates the transcriptional expression of
and
, as well as cell adhesion-related genes such as
or
, among others. Crispr-Cas9-mediated
gene disruption demonstrated that this gene plays a role in the process of mTEC-thymocyte adhesion. Consistent with the nuclear localization signal (NLS) encoded by
exon 3, which was targeted, we demonstrate that
KO
mTECs impair AIRE protein localization in the nucleus. Consequently, the loss of function of
reduced the ability of these cells to adhere to thymocytes. Their transcriptomes differed from their wild-type
counterparts, even during thymocyte adhesion. A set of mRNA isoforms that encode proteins involved in cell adhesion were also modulated during this process. This demonstrates that both thymocyte interactions and
influence transcriptome profiling of mTEC cells.
Vitiligo is the most frequent cause of depigmentation worldwide. Genetic association studies have discovered about 50 loci associated with disease, many with immunological functions. Among them is ...HLA-G, which modulates immunity by interacting with specific inhibitory receptors, mainly LILRB1 and LILRB2. Here we investigated the
and
association with vitiligo risk and evaluated the possible role of interactions between HLA-G and its receptors in this pathogenesis. We tested the association of the polymorphisms of
,
, and
with vitiligo using logistic regression along with adjustment by ancestry. Further, methods based on the multifactor dimensionality reduction (MDR) approach (MDR v.3.0.2, GMDR v.0.9, and MB-MDR) were used to detect potential epistatic interactions between polymorphisms from the three genes. An interaction involving rs9380142 and rs2114511 polymorphisms was identified by all methods used. The polymorphism rs9380142 is an
3'UTR variant (+3187) with a well-established role in mRNA stability. The polymorphism rs2114511 is located in the exonic region of
. Although no association involving this SNP has been reported, ChIP-Seq experiments have identified this position as an EBF1 binding site. These results highlight the role of an epistatic interaction between
and
in vitiligo pathogenesis.
ABSTRACT Fig tree (Ficus carica L.) breeding programs using conventional methods, such as directed crosses, to obtain new cultivars, are unworkable in many countries, including Brazil. Consequently, ...genetic breeding through mutagenesis has emerged as an important line of research that can improve this crop, and be a significant source of information about this species and assist in the implementation of propagation projects and appropriate management. The aim of this study was to verify the existence of epigenetic variability attributable to DNA methylation in irradiated fig selections when compared both to each other and to the main commercial cultivar, “Roxo-de-Valinhos”, which had previously used methylation-sensitive amplified polymorphism (MSAP) and DNA sequencing to detect the position of polymorphic regions, analyzable by bioinformatic tools. The sequencing of DNA, isolated from the differentially methylated sites, makes it possible to observe different patterns of methylation by sequencing the treated DNA with sodium bisulfite in the coding regions of regulatory genes active in the development, and fruit ripening stages. Furthermore, they have been found in the mitochondrial DNA of treatments which regulate the supply of energy in Adenosine triphosphate (ATP) form in plants. Closely related to their development, they justify the different phenotypes found in both fruit and plant growth that have suffered stress due to exposure to gamma radiation. Thus, future studies on gene expression in treatments have emerged as an extremely important strategy for understanding these complex regulatory systems, which may lead to the identification of genes of agricultural interest for the fig tree crop, and allow for manipulation and subsequent propagation of improved crops for commercial purposes.
Objective To identify genes specifically expressed in mammalian oocytes using an in silico subtraction, and to characterize the mRNA patterns of selected genes in oocytes, embryos, and adult tissues. ...Design Comparison between oocyte groups and between early embryo stages. Setting Laboratories of embryo manipulation and molecular biology from Departamento de Genética (FMRP) and Departamento de Ciências Básicas (FZEA) - University of São Paulo. Sample(s) Oocytes were collected from slaughtered cows for measurements, in vitro fertilization, and in vitro embryo culture. Somatic tissue, excluding gonad and uterus tissue, was collected from male and female cattle. Main Outcome Measure(s) Messenger RNA levels of poly(A)-binding protein nuclear-like 1 (Pabpnl1) and methyl-CpG–binding domain protein 3–like 2 (Mbd3l2). Result(s) Pabpnl1 mRNA was found to be expressed in oocytes, and Mbd3l2 transcripts were present in embryos. Quantification of Pabpnl1 transcripts showed no difference in levels between good- and bad-quality oocytes before in vitro maturation (IVM) or between good-quality oocytes before and after IVM. However, Pabpnl1 transcripts were not detected in bad-quality oocytes after IVM. Transcripts of the Mbd3l2 gene were found in 4-cell, 8-cell, and morula-stage embryos, with the highest level observed in 8-cell embryos. Conclusion(s) Pabpnl1 gene expression is restricted to oocytes and Mbd3l2 to embryos. Different Pabpnl1 mRNA levels in oocytes of varying viability suggest an important role in fertility involving the oocyte potential for embryo development.
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