Angela Gallo - Academia.edu (original) (raw)

Papers by Angela Gallo

Nature Communications, 2021

The maintenance of genomic stability requires the coordination of multiple cellular tasks upon th... more The maintenance of genomic stability requires the coordination of multiple cellular tasks upon the appearance of DNA lesions. RNA editing, the post-transcriptional sequence alteration of RNA, has a profound effect on cell homeostasis, but its implication in the response to DNA damage was not previously explored. Here we show that, in response to DNA breaks, an overall change of the Adenosine-to-Inosine RNA editing is observed, a phenomenon we call the RNA Editing DAmage Response (REDAR). REDAR relies on the checkpoint kinase ATR and the recombination factor CtIP. Moreover, depletion of the RNA editing enzyme ADAR2 renders cells hypersensitive to genotoxic agents, increases genomic instability and hampers homologous recombination by impairing DNA resection. Such a role of ADAR2 in DNA repair goes beyond the recoding of specific transcripts, but depends on ADAR2 editing DNA:RNA hybrids to ease their dissolution.

Cancer is driven by alterations of the genomic information, which carries mutations in key genes ... more Cancer is driven by alterations of the genomic information, which carries mutations in key genes providing selective advantage for clonal multiplication of cancer cells. However, mutations within DNA are not the only source for cell alteration. RNA molecules are targets of a series of post-transcriptional modifications, such as splicing and RNA editing, that can affect sequence, structure and stability. The most common type of RNA editing in humans converts Adenosine in RNA targets into Inosine (A-to-I) and is catalyzed by two adenosine deaminases that act on dsRNA (ADARs) family of enzymes (ADAR and ADARB1). Inosines are subsequently interpreted as guanosines by several cellular proteins and could ultimately lead to a genomic mutations (A-to-I/G). At present, it has been estimated that over 4 millions editing sites exist in our transcriptome involving coding and non-coding RNAs. These huge amounts of Inosine at RNA level are necessary for our survival and their levels is highly regulated in different tissues and during development. Considering the importance of ADAR activity in our cells we believe that if ADAR are not well regulated they may contribute to cancer on set and/or progression. The advent of high-throughput RNA sequencing has enabled identification of RNA editing sites and global analyses of cancer transcriptomes demonstrate that ADAR-mediated RNA editing dynamically contributes to genetic alterations in cancer, including high-grade gliomas. Glioblastoma (GBM) is one of the most common and aggressive primary brain tumor in humans and despite advances in understanding the molecular mechanisms underlying these tumors, current treatments are ineffective. In order to elucidate the glioma-specific RNA editing signature, we analyzed 146 RNA-Seq of primary glioblastomas from the TCGA dataset compared to 132 normal brain cortex RNA-Seq from the GTEx database and purified pools of normal cortex astrocytes. A-to-I editing events has been detected using a collection of more than 4 million annotated edited substrates and the REDItools suite of python scripts with stringent filters. By means of the Cuffquant/Cuffdiff tools, we have also compared global transcriptome profiles and ADAR genes expression patterns at gene level isoform level and using IHC at protein level. We found that a general down regulation of editing events at both recoding and non -recoding (Alus) sites that correlate with a down expression of ADARB1 enzyme. No differences were observed with ADAR expression. Overall, we observed a strong editing landscape perturbation in glioblastoma that could be important for identifying the most effective target genes for possible therapeutic intervention. Citation Format: Angela Gallo, Alessandro Silvestris, Valeriana Cesarini, Valentina Tassinari, Nicolò Mangraviti, Ernesto Picardi, Graziano Pesole. Deciphering inosinome in glioblastoma versus normal cortex [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1395. doi:10.1158/1538-7445.AM2017-1395

Journal of Neuro-Oncology

Purpose Patient-derived cancer cell lines can be very useful to investigate genetic as well as ep... more Purpose Patient-derived cancer cell lines can be very useful to investigate genetic as well as epigenetic mechanisms of transformation and to test new drugs. In this multi-centric study, we performed genomic and transcriptomic characterization of a large set of patient-derived glioblastoma (GBM) stem-like cells (GSCs). Methods 94 (80 I surgery/14 II surgery) and 53 (42 I surgery/11 II surgery) GSCs lines underwent whole exome and trascriptome analysis, respectively. Results Exome sequencing revealed TP53 as the main mutated gene (41/94 samples, 44%), followed by PTEN (33/94, 35%), RB1 (16/94, 17%) and NF1 (15/94, 16%), among other genes associated to brain tumors. One GSC sample bearing a BRAF p.V600E mutation showed sensitivity in vitro to a BRAF inhibitor. Gene Ontology and Reactome analysis uncovered several biological processes mostly associated to gliogenesis and glial cell differentiation, S − adenosylmethionine metabolic process, mismatch repair and methylation. Comparison of...

Cell Reports, 2020

Highlights d CD27 dull and CD27 bright MBCs share their VH repertoire but have different function... more Highlights d CD27 dull and CD27 bright MBCs share their VH repertoire but have different functions d CD27 dull MBCs are the long-lived substrate of selected and specific CD27 bright MBCs d The interplay between CD27 dull and CD27 bright MBCs preserves B cell memory d In pregnancy, MBCs decline, but persisting CD27 dull MBCs re-expand after delivery

Genome Biology, 2021

Background N6-methyladenosine (m6A) and adenosine-to-inosine (A-to-I) RNA editing are two of the ... more Background N6-methyladenosine (m6A) and adenosine-to-inosine (A-to-I) RNA editing are two of the most abundant RNA modification events affecting adenosines in mammals. Both these RNA modifications determine mRNA fate and play a pivotal role in tumor development and progression. Results Here, we show that METTL3, upregulated in glioblastoma, methylates ADAR1 mRNA and increases its protein level leading to a pro-tumorigenic mechanism connecting METTL3, YTHDF1, and ADAR1. We show that ADAR1 plays a cancer-promoting role independently of its deaminase activity by binding CDK2 mRNA, underlining the importance of ADARs as essential RNA-binding proteins for cell homeostasis as well as cancer progression. Additionally, we show that ADAR1 knockdown is sufficient to strongly inhibit glioblastoma growth in vivo. Conclusions Hence, our findings underscore METTL3/ADAR1 axis as a novel crucial pathway in cancer progression that connects m6A and A-to-I editing post-transcriptional events.

Adenosine deaminases that act on dsRNA (ADARs) are enzymes that target double-stranded regions of... more Adenosine deaminases that act on dsRNA (ADARs) are enzymes that target double-stranded regions of RNA converting adenosines into inosines (A-to-I editing) thus contributing to genome complexity and fine regulation of gene expression. It has been described that a member of the ADAR family, ADAR1, can target viruses and affect their replication process. Here we report evidence showing that ADAR1 stimulates human immuno deficiency virus type 1 (HIV-1) replication by using both editing-dependent and editing-independent mechanisms. We show that over-expression of ADAR1 in HIV-1 producer cells increases viral protein accumulation in an editing-independent manner. Moreover, HIV-1 virions generated in the presence of over-expressed ADAR1 but not an editing-inactive ADAR1 mutant are released more efficiently and display enhanced infectivity, as demonstrated by challenge assays performed with T cell lines and primary CD4 + T lymphocytes. Finally, we report that ADAR1 associates with HIV-1 RNAs and edits adenosines in the 5' untranslated region (UTR) and the Rev and Tat coding sequence. Overall these results suggest that HIV-1 has evolved mechanisms to take advantage of specific RNA editing activity of the host cell and disclose a stimulatory function of ADAR1 in the spread of HIV-1.

RNA editing is an important co/post-transcriptional molecular process able to modify RNAs by nucl... more RNA editing is an important co/post-transcriptional molecular process able to modify RNAs by nucleotide insertions/deletions or substitutions. In human, the most common RNA edit-ing event involves the deamination of adenosine (A) into inosine (I) through the adenosine deaminase acting on RNA proteins. Although A-to-I editing can occur in both coding and non-coding RNAs, recent findings, based on RNA-seq experiments, have clearly demon-strated that a large fraction of RNA editing events alter non-coding RNAs sequences including untranslated regions of mRNAs, introns, long non-coding RNAs (lncRNAs), and low molecular weight RNAs (tRNA, miRNAs, and others). An accurate detection of A-to-I events occurring in non-coding RNAs is of utmost importance to clarify yet unknown functional roles of RNA editing in the context of gene expression regulation and mainte-nance of cell homeostasis. In the last few years, massive transcriptome sequencing has been employed to identify putative RNA editi...

Biomolecules

Background: Epitranscriptomic mechanisms, such as A-to-I RNA editing mediated by ADAR deaminases,... more Background: Epitranscriptomic mechanisms, such as A-to-I RNA editing mediated by ADAR deaminases, contribute to cancer heterogeneity and patients’ stratification. ADAR enzymes can change the sequence, structure, and expression of several RNAs, affecting cancer cell behavior. In glioblastoma, an overall decrease in ADAR2 RNA level/activity has been reported. However, no data on ADAR2 protein levels in GBM patient tissues are available; and most data are based on ADARs overexpression experiments. Methods: We performed IHC analysis on GBM tissues and correlated ADAR2 levels and patients’ overall survival. We silenced ADAR2 in GBM cells, studied cell behavior, and performed a gene expression/editing analysis. Results: GBM tissues do not all show a low/no ADAR2 level, as expected by previous studies. Although, different amounts of ADAR2 protein were observed in different patients, with a low level correlating with a poor patient outcome. Indeed, reducing the endogenous ADAR2 protein in G...

(A) Sequence of an intronic portion of OPHN1 pre-m... more (A) Sequence of an intronic portion of OPHN1 pre-mRNA (intron 9–10), with AluJo region in blue and adenosines undergoing editing in red capital letters. In grey, primers used for PCR amplifications (AluJo Fw and AluJo Rev, Table S1, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0091351#pone.0091351.s001" target="_blank">File S1</a>). (B) Chromatogram of the AluJo region isolated from human brain cDNA, showing the newly identified editing sites, named as 1 to 14 and represented as a double peak of adenosine (green) and guanosine (black). The same positions (1–14) in the corresponding human brain gDNA sequence are only adenosine (Figure S1 in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0091351#pone.0091351.s001" target="_blank">File S1</a>).</p

For each tissue both the gDNA and the corresponding cDNA are shown. Arrows indicate sele... more For each tissue both the gDNA and the corresponding cDNA are shown. Arrows indicate selected edited positions (site 1, 2, 8, 9, 10 and 11) and the corresponding editing levels of the sequence chromatograms are reported above each site as percentages (%). Editing appears as a double peak of adenosine (green) and guanosine (black) in cDNA sequences, whereas only adenosines are present in the gDNA. A representative experiment out of three is shown. Editing levels at all the AluJo editing sites as found in human brain, spinal cord, skin as well as kidney and thyroid are reported in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0091351#pone-0091351-t001" target="_blank">Table 1</a>.</p

Next Generation Sequencing Workshop (Third Edition), 2011

Figure S1-S12 with figure legends. (PDF 1404 kb)

Table S5. GBM reclustering editing-based INO-1 and INO-2 in female patients. (XLSX 29 kb)

Table S4. Editing-based GBM reclustering. (XLSX 31 kb)

Basic Science, 2019

Background /Aim: Adenosine to inosine RNA editing is an essential posttranscriptional RNA modific... more Background /Aim: Adenosine to inosine RNA editing is an essential posttranscriptional RNA modification catalysed by Abstract BS47 Figure 2 Medin in deposited in the ECM by exosomes. A) Deposits of medin are observed in ECM from cultured VMSCs (white arrows). Fibronectic staining shows the presence of ECM. B) Medin deposits colocalise with exosome marker, CD63. Treatment with calcium and phosphate (+CaP) increases the size of medin deposits. Treatment with an exosome secretion inhibitor, spiroepoxide (+SOM), decreases the deposit size. D) The area of each medin aggregate was measured using ImageJ

Cancer Research, 2016

This study was supported by AIRC. Glioblastoma (or grade IV astrocytomas) is a highly heterogeneo... more This study was supported by AIRC. Glioblastoma (or grade IV astrocytomas) is a highly heterogeneous and deadly malignant brain tumor and one of the most incurable form of cancer in human that urgently needs new therapeutic targets. There is great interest in elucidating all the molecular basis and functional importance of genetics and epigenetic mechanisms occurring in glioblastoma. In our laboratory, we investigate whether essential post-transcriptional event (such as the A-to-I RNA editing) is a possible molecular mechanism involved in the appearance/progression of glioblastoma. A-to-I RNA editing modifies the nucleotide sequence of RNA target molecules. The ADARs (Adenosine Deaminases that Act on RNA) are the enzymes responsible for this conversion acting on pre-mRNAs or non-coding RNAs (such as microRNA). Inosine, is recognized by the translation and splicing machineries, as Guanosine. Therefore, ADARs can play a direct and critical role in generating alternative protein isoform...

PLoS ONE, 2012

RNA editing is a post-transcriptional process occurring in a wide range of organisms. In human br... more RNA editing is a post-transcriptional process occurring in a wide range of organisms. In human brain, the A-to-I RNA editing, in which individual adenosine (A) bases in pre-mRNA are modified to yield inosine (I), is the most frequent event. Modulating gene expression, RNA editing is essential for cellular homeostasis. Indeed, its deregulation has been linked to several neurological and neurodegenerative diseases. To date, many RNA editing sites have been identified by next generation sequencing technologies employing massive transcriptome sequencing together with whole genome or exome sequencing. While genome and transcriptome reads are not always available for single individuals, RNA-Seq data are widespread through public databases and represent a relevant source of yet unexplored RNA editing sites. In this context, we propose a simple computational strategy to identify genomic positions enriched in novel hypothetical RNA editing events by means of a new two-steps mapping procedure requiring only RNA-Seq data and no a priori knowledge of RNA editing characteristics and genomic reads. We assessed the suitability of our procedure by confirming A-to-I candidates using conventional Sanger sequencing and performing RNA-Seq as well as whole exome sequencing of human spinal cord tissue from a single individual.

Leukemia, 2017

Adenosine deaminases acting on RNA (ADARs) are key proteins for hematopoietic stem cell self-rene... more Adenosine deaminases acting on RNA (ADARs) are key proteins for hematopoietic stem cell self-renewal and for survival of differentiating progenitor cells. However, their specific role in myeloid cell maturation has been poorly investigated. Here we show that ADAR1 is present at basal level in the primary myeloid leukemia cells obtained from patients at diagnosis as well as in myeloid U-937 and THP1 cell lines and its expression correlates with the editing levels. Upon phorbol-myristate acetate or Vitamin D3/granulocyte macrophage colony-stimulating factor (GM-CSF)-driven differentiation, both ADAR1 and ADAR2 enzymes are upregulated, with a concomitant global increase of A-to-I RNA editing. ADAR1 silencing caused an editing decrease at specific ADAR1 target genes, without, however, interfering with cell differentiation or with ADAR2 activity. Remarkably, ADAR2 is absent in the undifferentiated cell stage, due to its elimination through the ubiquitin-proteasome pathway, being strongly upregulated at the end of the differentiation process. Of note, peripheral blood monocytes display editing events at the selected targets similar to those found in differentiated cell lines. Taken together, the data indicate that ADAR enzymes play important and distinct roles in myeloid cells.