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Glioblastoma (GBM), a highly lethal form of adult malignant gliomas with little clinical advancement, raises the need for alternative therapeutic approaches. Lipid-soluble vitamins have gained attention in malignant brain tumors owing to their pleiotropic properties and their anti-cancer potential have been reported in a number of human GBM cell lines. The aim of this paper is to systematically review and describe the roles of various biomarkers regulated by lipid-soluble vitamins, such as vitamins A, D, E, and K, in the pathophysiology of GBM. Briefly, research articles published between 2005 and 2021 were systematically searched and selected from five databases (Scopus, PubMed, Ovid MEDLINE, EMBASE via Ovid, and Web of Science) based on the study's inclusion and exclusion criteria. In addition, a number of hand-searched research articles identified from Google Scholar were also included for the analysis. A total of 40 differentially expressed biomarkers were identified from the 19 eligible studies. The results from the analysis suggest that retinoids activate cell differentiation and suppress the biomarkers responsible for stemness in human GBM cells. Vitamin D appears to preferentially modulate several cell cycle biomarkers, while vitamin E derivatives seem to predominantly modulate biomarkers related to apoptosis. However, vitamin K1 did not appear to induce any significant changes to the Raf/MEK/ERK signaling or apoptotic pathways in human GBM cell lines. From the systematic analysis, 12 biomarkers were identified that may be of interest for further studies, as these were modulated by one or two of these lipid-soluble vitamins.

The aim of the present study was to determine the genetic aberrations and novel transcripts, particularly the fusion transcripts, involved in the pathogenesis of low-grade and anaplastic oligodendroglioma. In the present study, tissue samples were obtained from patients with oligodendroglioma and additionally from archived tissue samples from the Brain Tumor Tissue Bank of the Brain Tumor Foundation of Canada. Six samples were obtained, three of which were low-grade oligodendroglioma and the other three anaplastic oligodendroglioma. DNA and RNA were extracted from each tissue sample. The resulting genomic DNA was then hybridized using the Agilent CytoSure 4×180K oligonucleotide array. Human reference DNA and samples were labeled using Cy3 cytidine 5'-triphosphate (CTP) and Cy5 CTP, respectively, while human Cot-1 DNA was used to reduce non-specific binding. Microarray-based comparative genomic hybridization data was then analyzed for genetic aberrations using the Agilent Cytosure Interpret software v3.4.2. The total RNA isolated from each sample was mixed with oligo dT magnetic beads to enrich for poly(A) mRNA. cDNAs were then synthesized and subjected to end-repair, poly(A) addition and connected using sequencing adapters using the Illumina TruSeq RNA Sample Preparation kit. The fragments were then purified and selected as templates for polymerase chain reaction amplification. The final library was constructed with fragments between 350-450 base pairs and sequenced using deep transcriptome sequencing on an Illumina HiSeq 2500 sequencer. The array comparative genomic hybridization revealed numerous amplifications and deletions on several chromosomes in all samples. However, the most interesting result was from the next generation sequencing, where one anaplastic oligodendroglioma sample was demonstrated to have five novel fusion genes that may potentially serve a critical role in tumor pathogenesis and progression.

The aims of the present study were to undertake gene expression profiling of the blood of glioma patients to determine key genetic components of signaling pathways and to develop a panel of genes that could be used as a potential blood-based biomarker to differentiate between high and low grade gliomas, non-gliomas and control samples. In this study, blood samples were obtained from glioma patients, non-glioma and control subjects. Ten samples each were obtained from patients with high and low grade tumours, respectively, ten samples from non-glioma patients and twenty samples from control subjects. Total RNA was isolated from each sample after which first and second strand synthesis was performed. The resulting cRNA was then hybridized with the Agilent Whole Human Genome (4x44K) microarray chip according to the manufacturer's instructions. Universal Human Reference RNA and samples were labeled with Cy3 CTP and Cy5 CTP, respectively. Microarray data were analyzed by the Agilent Gene Spring 12.1V software using stringent criteria which included at least a 2-fold difference in gene expression between samples. Statistical analysis was performed using the unpaired Student's t-test with a p<0.01. Pathway enrichment was also performed, with key genes selected for validation using droplet digital polymerase chain reaction (ddPCR). The gene expression profiling indicated that were a substantial number of genes that were differentially expressed with more than a 2-fold change (p<0.01) between each of the four different conditions. We selected key genes within significant pathways that were analyzed through pathway enrichment. These key genes included regulators of cell proliferation, transcription factors, cytokines and tumour suppressor genes. In the present study, we showed that key genes involved in significant and well established pathways, could possibly be used as a potential blood-based biomarker to differentiate between high and low grade gliomas, non-gliomas and control samples.

We have identified four mutations in Xenopus TFIIIA that increase the stability of TFIIIA-5 S rRNA gene complexes. In each case, the mutation has a relatively modest effect on equilibrium binding affinity. In three cases, these equilibrium binding effects can be ascribed primarily to decreases in the rate constant for protein-DNA complex dissociation. In the fourth case, however, a substitution of phenylalanine for the wild-type leucine at position 148 in TFIIIA results in much larger compensating changes in the kinetics of complex assembly and dissociation. The data support a model in which a relatively unstable population of complexes with multi-component dissociation kinetics forms rapidly; complexes then undergo a slow conformational change that results in very stable, kinetically homogeneous TFIIIA-DNA complexes. The L148F mutant protein acts as a particularly potent transcriptional activator when it is fused to the VP16 activation domain and expressed in yeast cells. Substitution of L148 to tyrosine or tryptophan produces an equally strong transcriptional activator. Substitution to histidine results in genetic and biochemical effects that are more modest than, but similar to, those observed with the L148F mutation. We propose that an amino acid with a planar side chain at position 148 can intercalate between adjacent base pairs in the intermediate element of the 5 S rRNA gene. Intercalation occurs slowly but results in a very stable DNA-protein complex. These results suggest that transcriptional activation by a cis-acting sequence element is largely dependent on the kinetic, rather than the thermodynamic, stability of the complex formed with an activator protein. Thus, transcriptional activation is dependent in large part on the lifetime of the activator-DNA complex rather than on binding site occupancy at steady state. Introduction of intercalating amino acids into zinc finger proteins may be a useful tool for producing artificial transcription factors with particularly high in vivo activity.

Semicarbazide-sensitive amine oxidase (SSAO) catalyzes formation of methylglyoxal (MG) from aminoacetone; MG then reacts with proteins to form advanced glycation end products or AGEs. Because of its potential to generate MG, SSAO may contribute to AGE-associated vascular complications of aging and diabetes. We developed a method to measure SSAO activity in bovine aortic smooth muscle cells (BASMC) based on the oxidation of 2',7'-dichlorofluorescin by hydrogen peroxide and horseradish peroxidase. The SSAO activity was completely inhibited by 10 mM semicarbazide. Argpyrimidine is a readily detectable fluorescent product of the reaction between MG and arginine. Cell lysates incubated with aminoacetone formed argpyrimidine in a reaction that was inhibited by 20 mM semicarbazide. Immunostaining of tissue sections showed that aminoacetone-treated rats (normal as well as diabetic) formed more argpyrimidine in aortic smooth muscle than untreated controls. We believe that SSAO can enhance AGE synthesis in the macrovasculature of diabetic individuals by production of MG.

Expression of light harvesting II genes and of bacteriochlorophyll and carotenoid biosynthesis genes in Rhodobacter capsulatus is repressed under aerobic growth conditions by the transcription factor CrtJ. In this study, we demonstrate that the crtA-crtI intergenic region contains divergent promoters that initiate transcription 116 base pairs apart, based on primer extension analyses. DNase I protection assays demonstrate that purified CrtJ binds to one palindrome that overlaps the crtA -10 promoter recognition sequence as well as to a second palindrome that overlaps the -35 crtI promoter recognition sequence. Similar analyses also show that the puc promoter region contains two distant CrtJ palindromes, with one near the -35 promoter recognition sequence and the other located 240 base pairs upstream. Gel mobility shift and filter retention assays indicate that CrtJ binds in a cooperative manner to these distantly separated palindromes. In vivo expression assays with puc and crtI promoter reporter plasmids further demonstrate that aerobic repression of puc and crtI expression requires both CrtJ palindromes. These in vitro and in vivo results indicate that aerobic repression of puc, crtA, and crtI expression involves cooperative interactions between CrtJ bound to distant palindromes. A DNA looping model is discussed.

Previous studies demonstrated that bacteriochlorophyll, carotenoid, and light harvesting gene expression in Rhodobacter capsulatus is repressed under aerobic growth conditions by the repressor CrtJ. Isolated CrtJ is known to bind to the palindrome TGTN12ACA, which is present in two copies in the bchC promoter, one of which spans the -35 and the other the -10 sigma-70 recognition sequences. In this study, we demonstrate that CrtJ binds to the two palindromic sites in the bchC promoter in a cooperative manner. The level of cooperativity of CrtJ binding to the -35 palindrome was shown to be 26-fold. A distance of 8 base pairs between the two palindromic sites was shown to be critical for cooperative binding, as evidenced by the disruption of binding that resulted when +6 and +11 base pairs were inserted between the palindromes.

Previous genetic analysis indicated that the photosynthesis gene cluster from Rhodobacter capsulatus coded for the transcription factor, CrtJ, that is responsible for aerobic repression of bacteriochlorophyll, carotenoid, and light harvesting-II gene expression. In this study, we have heterologously overexpressed and purified CrtJ to homogeneity and shown by gel mobility shift assays that CrtJ is biologically active. DNase I footprint analysis confirms molecular genetic studies by showing that CrtJ binds to conserved palindromic sequences that overlap the -10 and -35 promoter regions of the bchC operon. Graphs of the percentage of DNA bound versus protein concentration show sigmoidal curves, which is highly indicative of cooperative binding of CrtJ to the two palindromic sites. A binding constant for interaction of CrtJ with the palindrome that spans the -10 region was calculated to be 4.8 x 10(-9) M, whereas affinity for the palindrome that spans the -35 region was found to be 2.9 x 10(-9) M. Binding of CrtJ to the bchC promoter region was also found to be redox-sensitive, with CrtJ exhibiting a 4.5-fold higher binding affinity under oxidizing versus reducing conditions.

For most species of purple photosynthetic bacteria, the presence of molecular oxygen represses synthesis of carotenoids and bacteriochlorophyll. In this study we characterize a strain of Rhodobacter capsulatus, DB469, which contains a genomic disruption of an open reading frame in the photosynthesis gene cluster termed ORF469. Characterization of the steady-state level of bacteriochlorophyll synthesis demonstrates that disruption of ORF469 results in a 2.5-fold increase in aerobic synthesis of bacteriochlorophyll over that observed with the parent strain. Utilizing reporter plasmids that contain transcriptional fusions of lacZ to various carotenoid and bacteriochlorophyll biosynthesis genes, we also demonstrate that disruption of ORF469 leads to an approximate twofold increase in bacteriochlorophyll and carotenoid gene expression under anaerobic growth conditions. Similar analysis with reporter plasmids that contain translational fusions of lacZ to the puf, puh, and puc operons demonstrates that disruption of ORF469 leads to elevated levels of aerobic transcription of light harvesting-II genes (puc), without affecting light harvesting-I or reaction center gene expression (puf and puh, respectively). Gel mobility analysis demonstrates that DB469 cells lack a DNA-binding protein that interacts with a palindromic sequence in the bchC promoter region. The results of this study indicate that ORF469 codes for a DNA-binding protein that acts as an aerobic repressor of promoters for bacteriochlorophyll, carotenoid, and light harvesting-II gene expression.