Chemical Inhibition of Apurinic-Apyrimidinic Endonuclease 1 Redox and DNA Repair Functions Affects the Inflammatory Response via Different but Overlapping Mechanisms.

The presence of oxidized DNA lesions, such as 7,8-dihydro-8-oxoguanine (8-oxoG) and apurinic/apyrimidinic sites (AP sites), has been described as epigenetic signals that are involved in gene expression control. In mammals, Apurinic-apyrimidinic endonuclease 1/Redox factor-1 (APE1/Ref-1) is the main AP endonuclease of the base excision repair (BER) pathway and is involved in active demethylation processes. In addition, APE1/Ref-1, through its redox function, regulates several transcriptional factors. However, the transcriptional control targets of each APE1 function are not completely known. In this study, a transcriptomic approach was used to investigate the effects of chemical inhibition of APE1/Ref-1 redox or DNA repair functions by E3330 or methoxyamine (MX) in an inflammatory cellular model. Under lipopolysaccharide (LPS) stimulation, both E3330 and MX reduced the expression of some cytokines and chemokines. Interestingly, E3330 treatment reduced cell viability after 48 h of the treatment. Genes related to inflammatory response and mitochondrial processes were downregulated in both treatments. In the E3330 treatment, RNA processing and ribosome biogenesis genes were downregulated, while they were upregulated in the MX treatment. Furthermore, in the E3330 treatment, the cellular stress response was the main upregulated process, while the cellular macromolecule metabolic process was observed in MX-upregulated genes. Nuclear respiratory factor 1 (NRF1) was predicted to be a master regulator of the downregulated genes in both treatments, while the ETS transcription factor ELK1 (ELK1) was predicted to be a master regulator only for E3330 treatment. Decreased expression of ELK1 and its target genes and a reduced 28S/18S ratio were observed, suggesting impaired rRNA processing. In addition, both redox and repair functions can affect the expression of NRF1 and GABPA target genes. The master regulators predicted for upregulated genes were YY1 and FLI1 for the E3330 and MX treatments, respectively. In summary, the chemical inhibition of APE1/Ref-1 affects gene expression regulated mainly by transcriptional factors of the ETS family, showing partial overlap of APE1 redox and DNA repair functions, suggesting that these activities are not entirely independent. This work provides a new perspective on the interaction between APE1 redox and DNA repair activity in inflammatory response modulation and transcription.

XPA deficiency affects the ubiquitin-proteasome system function.

Xeroderma pigmentosum complementation group A (XPA), is defective in xeroderma pigmentosum patients, causing pre-disposition to skin cancer and neurological abnormalities, which is not well understood. Here, we analyzed the XPA-deficient cells transcriptional profile under oxidative stress. The imbalance in of ubiquitin-proteasome system (UPS) gene expression was observed in XPA-deficient cells and the involvement of nuclear factor erythroid 2-related factor-2 (NFE2L2) was indicated. Co-immunoprecipitation assays showed the interaction between XPA, apurinic-apyrimidinic endonuclease 1 (APE1) and NFE2L2 proteins. Decreased NFE2L2 protein expression and proteasome activity was also observed in XPA-deficient cells. The data suggest the involvement of the growth arrest and DNA-damage-inducible beta (GADD45ß) in NFE2L2 functions. Similar results were obtained in xpa-1 (RNAi) Caenorhabditis elegans suggesting the conservation of XPA and NFE2L2 interactions. In conclusion, stress response activation occurs in XPA-deficient cells under oxidative stress; however, these cells fail to activate the UPS cytoprotective response, which may contribute to XPA patient's phenotypes.

A comprehensive analysis of core polyadenylation sequences and regulation by microRNAs in a set of cancer predisposition genes.

Two core polyadenylation elements (CPE) located in the 3' untranslated region of eukaryotic pre-mRNAs play an essential role in their processing: the polyadenylation signal (PAS) AAUAAA and the cleavage site (CS), preferentially a CA dinucleotide. Herein, we characterized PAS and CS sequences in a set of cancer predisposition genes (CPGs) and performed an in silico investigation of microRNAs (miRNAs) regulation to identify potential tumor-suppressive and oncogenic miRNAs. NCBI and alternative polyadenylation databases were queried to characterize CPE sequences in 117 CPGs, including 81 and 17 known tumor suppressor genes and oncogenes, respectively. miRNA-mediated regulation analysis was performed using predicted and validated data sources. Based on NCBI analyses, we did not find an established PAS in 21 CPGs, and verified that the majority of PAS already described (74.4%) had the canonical sequence AAUAAA. Interestingly, "AA" dinucleotide was the most common CS (37.5%) associated with this set of genes. Approximately 90% of CPGs exhibited evidence of alternative polyadenylation (more than one functional PAS). Finally, the mir-192 family was significantly overrepresented as regulator of tumor suppressor genes (P < 0.01), which suggests a potential oncogenic function. Overall, this study provides a landscape of CPE in CPGs, which might be useful in development of future molecular analyses covering these frequently neglected regulatory sequences.

Gain of transcription factor binding sites is associated to changes in the expression signature of human brain and testis and is correlated to genes with higher expression breadth.

The gain of transcription factor binding sites (TFBS) is believed to represent one of the major causes of biological innovation. Here we used strategies based on comparative genomics to identify 21,822 TFBS specific to the human lineage (TFBS-HS), when compared to chimpanzee and gorilla genomes. More than 40% (9,206) of these TFBS-HS are in the vicinity of 1,283 genes. A comparison of the expression pattern of these genes and the corresponding orthologs in chimpanzee and gorilla identified genes differentially expressed in human tissues. These genes show a more divergent expression pattern in the human testis and brain, suggesting a role for positive selection in the fixation of TFBS gains. Genes associated with TFBS-HS were enriched in gene ontology categories related to transcriptional regulation, signaling, differentiation/development and nervous system. Furthermore, genes associated with TFBS-HS present a higher expression breadth when compared to genes in general. This biased distribution is due to a preferential gain of TFBS in genes with higher expression breadth rather than a shift in the expression pattern after the gain of TFBS.

Genome-wide identification of cancer/testis genes and their association with prognosis in a pan-cancer analysis.

Cancer/testis (CT) genes are excellent candidates for cancer immunotherapies because of their restrict expression in normal tissues and the capacity to elicit an immune response when expressed in tumor cells. In this study, we provide a genome-wide screen for CT genes with the identification of 745 putative CT genes. Comparison with a set of known CT genes shows that 201 new CT genes were identified. Integration of gene expression and clinical data led us to identify dozens of CT genes associated with either good or poor prognosis. For the CT genes related to good prognosis, we show that there is a direct relationship between CT gene expression and a signal for CD8+ cells infiltration for some tumor types, especially melanoma.

A tool for integrating genetic and mass spectrometry-based peptide data: Proteogenomics Viewer: PV: A genome browser-like tool, which includes MS data visualization and peptide identification parameters.

In this manuscript we describe Proteogenomics Viewer, a web-based tool that collects MS peptide identification, indexes to genomic sequence and structure, assigns exon usage, reports the identified protein isoforms with genomic alignments and, most importantly, allows the inspection of MS2 information for proper peptide identification. It also provides all performed indexing to facilitate global analysis of the data. The relevance of such tool is that there has been an increase in the number of proteogenomic efforts to improve the annotation of both genomics and proteomics data, culminating with the release of the two human proteome drafts. It is now clear that mass spectrometry-based peptide identification of uncharacterized sequences, such as those resulting from unpredicted exon joints or non-coding regions, is still prone to a higher than expected false discovery rate. Therefore, proper visualization of the raw data and the corresponding genome alignments are fundamental for further data validation and interpretation. Also see the video abstract here: http://youtu.be/5NzyRvuk4Ac.