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Article ; Online: RNA Post-Transcriptional Modification Mapping Data Analysis Using RNA Framework.

Manfredonia, Ilaria / Incarnato, Danny

Methods in molecular biology (Clifton, N.J.)

2021 Volume 2298, Page(s) 3–13

Abstract: RNA post-transcriptional modifications (PTMs) are progressively gaining relevance in the study of coding-independent functions of RNA. RNA PTMs act as dynamic regulators of several aspects of the RNA physiology, from translation to half-life. Rising ... ...

Abstract	RNA post-transcriptional modifications (PTMs) are progressively gaining relevance in the study of coding-independent functions of RNA. RNA PTMs act as dynamic regulators of several aspects of the RNA physiology, from translation to half-life. Rising interest is supported by the advance of high-throughput techniques enabling the detection of these modifications on a transcriptome-wide scale. To this end, here we illustrate the usefulness of RNA Framework, a comprehensive toolkit for the analysis of RNA PTM mapping experiments, by reanalyzing two published transcriptome-scale datasets of N1-methyladenosine (m
MeSH term(s)	Adenosine/genetics ; Data Analysis ; High-Throughput Nucleotide Sequencing/methods ; Pseudouridine/genetics ; RNA/genetics ; RNA Processing, Post-Transcriptional/genetics ; Sequence Analysis, RNA/methods ; Transcriptome/genetics
Chemical Substances	Pseudouridine (1445-07-4) ; RNA (63231-63-0) ; Adenosine (K72T3FS567)
Language	English
Publishing date	2021-06-03
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-0716-1374-0_1
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Article ; Online: Structure and regulation of coronavirus genomes: state-of-the-art and novel insights from SARS-CoV-2 studies.

Manfredonia, Ilaria / Incarnato, Danny

Biochemical Society transactions

2020 Volume 49, Issue 1, Page(s) 341–352

Abstract: Coronaviruses (CoV) are positive-sense single-stranded RNA viruses, harboring the largest viral RNA genomes known to date. Apart from the primary sequence encoding for all the viral proteins needed for the generation of new viral particles, certain ... ...

Abstract	Coronaviruses (CoV) are positive-sense single-stranded RNA viruses, harboring the largest viral RNA genomes known to date. Apart from the primary sequence encoding for all the viral proteins needed for the generation of new viral particles, certain regions of CoV genomes are known to fold into stable structures, controlling several aspects of CoV life cycle, from the regulation of the discontinuous transcription of subgenomic mRNAs, to the packaging of the genome into new virions. Here we review the current knowledge on CoV RNA structures, discussing it in light of the most recent discoveries made possible by analyses of the SARS-CoV-2 genome.
MeSH term(s)	COVID-19/prevention & control ; COVID-19/virology ; Coronavirus/genetics ; Frameshifting, Ribosomal/genetics ; Genome, Viral/genetics ; Humans ; Models, Molecular ; Nucleic Acid Conformation ; RNA, Ribosomal/chemistry ; RNA, Ribosomal/genetics ; RNA, Viral/chemistry ; RNA, Viral/genetics ; SARS-CoV-2/genetics ; SARS-CoV-2/physiology
Chemical Substances	RNA, Ribosomal ; RNA, Viral
Language	English
Publishing date	2020-12-26
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	184237-7
ISSN	1470-8752 ; 0300-5127
ISSN (online)	1470-8752
ISSN	0300-5127
DOI	10.1042/BST20200670
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Article ; Online: Genome-scale deconvolution of RNA structure ensembles.

Morandi, Edoardo / Manfredonia, Ilaria / Simon, Lisa M / Anselmi, Francesca / van Hemert, Martijn J / Oliviero, Salvatore / Incarnato, Danny

Nature methods

2021 Volume 18, Issue 3, Page(s) 249–252

Abstract: RNA structure heterogeneity is a major challenge when querying RNA structures with chemical probing. We introduce DRACO, an algorithm for the deconvolution of coexisting RNA conformations from mutational profiling experiments. Analysis of the SARS-CoV-2 ... ...

Abstract	RNA structure heterogeneity is a major challenge when querying RNA structures with chemical probing. We introduce DRACO, an algorithm for the deconvolution of coexisting RNA conformations from mutational profiling experiments. Analysis of the SARS-CoV-2 genome using dimethyl sulfate mutational profiling with sequencing (DMS-MaPseq) and DRACO, identifies multiple regions that fold into two mutually exclusive conformations, including a conserved structural switch in the 3' untranslated region. This work may open the way to dissecting the heterogeneity of the RNA structurome.
MeSH term(s)	3' Untranslated Regions/genetics ; Algorithms ; COVID-19 ; Genome, Viral/genetics ; Humans ; Mutation/drug effects ; Mutation/genetics ; Nucleic Acid Conformation ; RNA, Viral/chemistry ; RNA, Viral/genetics ; SARS-CoV-2/genetics ; Sulfuric Acid Esters/pharmacology
Chemical Substances	3' Untranslated Regions ; RNA, Viral ; Sulfuric Acid Esters ; dimethyl sulfate (JW5CW40Z50)
Language	English
Publishing date	2021-02-22
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2169522-2
ISSN	1548-7105 ; 1548-7091
ISSN (online)	1548-7105
ISSN	1548-7091
DOI	10.1038/s41592-021-01075-w
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Article ; Online: Subcellular Characterization of Nicotinamide Adenine Dinucleotide Biosynthesis in Metastatic Melanoma by Using Organelle-Specific Biosensors.

Gaudino, Federica / Manfredonia, Ilaria / Managò, Antonella / Audrito, Valentina / Raffaelli, Nadia / Vaisitti, Tiziana / Deaglio, Silvia

Antioxidants & redox signaling

2019 Volume 31, Issue 15, Page(s) 1150–1165

Abstract: Aim: ...

Abstract	Aim:
MeSH term(s)	Biosensing Techniques/methods ; Cell Line, Tumor ; Humans ; Lentivirus/genetics ; Melanoma/metabolism ; Microscopy, Confocal ; Mitochondria/metabolism ; NAD/metabolism ; Organelles/metabolism ; Time-Lapse Imaging
Chemical Substances	NAD (0U46U6E8UK)
Language	English
Publishing date	2019-08-27
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1483836-9
ISSN	1557-7716 ; 1523-0864
ISSN (online)	1557-7716
ISSN	1523-0864
DOI	10.1089/ars.2019.7799
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Article ; Online: Tumors carrying BRAF-mutations over-express NAMPT that is genetically amplified and possesses oncogenic properties.

Audrito, Valentina / Moiso, Enrico / Ugolini, Filippo / Messana, Vincenzo Gianluca / Brandimarte, Lorenzo / Manfredonia, Ilaria / Bianchi, Simonetta / De Logu, Francesco / Nassini, Romina / Szumera-Ciećkiewicz, Anna / Taverna, Daniela / Massi, Daniela / Deaglio, Silvia

Journal of translational medicine

2022 Volume 20, Issue 1, Page(s) 118

Abstract: Background: Nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in nicotinamide adenine dinucleotide (NAD) biosynthesis, is up-regulated in several cancers, including metastatic melanoma (MM). The BRAF oncogene is mutated in ... ...

Abstract	Background: Nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in nicotinamide adenine dinucleotide (NAD) biosynthesis, is up-regulated in several cancers, including metastatic melanoma (MM). The BRAF oncogene is mutated in different cancer types, among which MM and thyroid carcinoma (THCA) are prominent. Drugs targeting mutant BRAF are effective, especially in MM patients, even though resistance rapidly develops. Previous data have linked NAMPT over-expression to the acquisition of BRAF resistance, paving the way for therapeutic strategies targeting the two pathways. Methods: Exploiting the TCGA database and a collection of MM and THCA tissue microarrays we studied the association between BRAF mutations and NAMPT expression. BRAF wild-type (wt) cell lines were genetically engineered to over-express the BRAF V600E construct to demonstrate a direct relationship between over-activation of the BRAF pathway and NAMPT expression. Responses of different cell line models to NAMPT (i)nhibitors were studied using dose-response proliferation assays. Analysis of NAMPT copy number variation was performed in the TCGA dataset. Lastly, growth and colony forming assays were used to study the tumorigenic functions of NAMPT itself. Results: The first finding of this work is that tumor samples carrying BRAF-mutations over-express NAMPT, as demonstrated by analyzing the TCGA dataset, and MM and THC tissue microarrays. Importantly, BRAF wt MM and THCA cell lines modified to over-express the BRAF V600E construct up-regulated NAMPT, confirming a transcriptional regulation of NAMPT following BRAF oncogenic signaling activation. Treatment of BRAF-mutated cell lines with two different NAMPTi was followed by significant reduction of tumor growth, indicating NAMPT addiction in these cells. Lastly, we found that several tumors over-expressing the enzyme, display NAMPT gene amplification. Over-expression of NAMPT in BRAF wt MM cell line and in fibroblasts resulted in increased growth capacity, arguing in favor of oncogenic properties of NAMPT. Conclusions: Overall, the association between BRAF mutations and NAMPT expression identifies a subset of tumors more sensitive to NAMPT inhibition opening the way for novel combination therapies including NAMPTi with BRAFi/MEKi, to postpone and/or overcome drug resistance. Lastly, the over-expression of NAMPT in several tumors could be a key and broad event in tumorigenesis, substantiated by the finding of NAMPT gene amplification.
MeSH term(s)	Carcinogenesis/genetics ; Cell Line, Tumor ; DNA Copy Number Variations ; Humans ; Melanoma/pathology ; Mutation/genetics ; Nicotinamide Phosphoribosyltransferase/genetics ; Nicotinamide Phosphoribosyltransferase/metabolism ; Oncogenes ; Proto-Oncogene Proteins B-raf/genetics
Chemical Substances	Nicotinamide Phosphoribosyltransferase (EC 2.4.2.12) ; BRAF protein, human (EC 2.7.11.1) ; Proto-Oncogene Proteins B-raf (EC 2.7.11.1)
Language	English
Publishing date	2022-03-10
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2118570-0
ISSN	1479-5876 ; 1479-5876
ISSN (online)	1479-5876
ISSN	1479-5876
DOI	10.1186/s12967-022-03315-9
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Article ; Online: Tumors carrying BRAF-mutations over-express NAMPT that is genetically amplified and possesses oncogenic properties

Valentina Audrito / Enrico Moiso / Filippo Ugolini / Vincenzo Gianluca Messana / Lorenzo Brandimarte / Ilaria Manfredonia / Simonetta Bianchi / Francesco De Logu / Romina Nassini / Anna Szumera-Ciećkiewicz / Daniela Taverna / Daniela Massi / Silvia Deaglio

Journal of Translational Medicine, Vol 20, Iss 1, Pp 1-

2022 Volume 13

Abstract: Abstract Background Nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in nicotinamide adenine dinucleotide (NAD) biosynthesis, is up-regulated in several cancers, including metastatic melanoma (MM). The BRAF oncogene is mutated in ... ...

Abstract	Abstract Background Nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in nicotinamide adenine dinucleotide (NAD) biosynthesis, is up-regulated in several cancers, including metastatic melanoma (MM). The BRAF oncogene is mutated in different cancer types, among which MM and thyroid carcinoma (THCA) are prominent. Drugs targeting mutant BRAF are effective, especially in MM patients, even though resistance rapidly develops. Previous data have linked NAMPT over-expression to the acquisition of BRAF resistance, paving the way for therapeutic strategies targeting the two pathways. Methods Exploiting the TCGA database and a collection of MM and THCA tissue microarrays we studied the association between BRAF mutations and NAMPT expression. BRAF wild-type (wt) cell lines were genetically engineered to over-express the BRAF V600E construct to demonstrate a direct relationship between over-activation of the BRAF pathway and NAMPT expression. Responses of different cell line models to NAMPT (i)nhibitors were studied using dose–response proliferation assays. Analysis of NAMPT copy number variation was performed in the TCGA dataset. Lastly, growth and colony forming assays were used to study the tumorigenic functions of NAMPT itself. Results The first finding of this work is that tumor samples carrying BRAF-mutations over-express NAMPT, as demonstrated by analyzing the TCGA dataset, and MM and THC tissue microarrays. Importantly, BRAF wt MM and THCA cell lines modified to over-express the BRAF V600E construct up-regulated NAMPT, confirming a transcriptional regulation of NAMPT following BRAF oncogenic signaling activation. Treatment of BRAF-mutated cell lines with two different NAMPTi was followed by significant reduction of tumor growth, indicating NAMPT addiction in these cells. Lastly, we found that several tumors over-expressing the enzyme, display NAMPT gene amplification. Over-expression of NAMPT in BRAF wt MM cell line and in fibroblasts resulted in increased growth capacity, arguing in favor of ...
Keywords	NAMPT ; Oncogene ; Transformation ; BRAF oncogenic signaling ; MAPK ; NAMPT inhibitors ; Medicine ; R
Subject code	616
Language	English
Publishing date	2022-03-01T00:00:00Z
Publisher	BMC
Document type	Article ; Online
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Article ; Online: Genome-wide mapping of SARS-CoV-2 RNA structures identifies therapeutically-relevant elements.

Manfredonia, Ilaria / Nithin, Chandran / Ponce-Salvatierra, Almudena / Ghosh, Pritha / Wirecki, Tomasz K / Marinus, Tycho / Ogando, Natacha S / Snijder, Eric J / van Hemert, Martijn J / Bujnicki, Janusz M / Incarnato, Danny

Nucleic acids research

2020 Volume 48, Issue 22, Page(s) 12436–12452

Abstract: SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome, whose outbreak caused the ongoing COVID-19 pandemic. The ability of coronaviruses to rapidly evolve, adapt, and cross species barriers makes the development of ... ...

Abstract	SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome, whose outbreak caused the ongoing COVID-19 pandemic. The ability of coronaviruses to rapidly evolve, adapt, and cross species barriers makes the development of effective and durable therapeutic strategies a challenging and urgent need. As for other RNA viruses, genomic RNA structures are expected to play crucial roles in several steps of the coronavirus replication cycle. Despite this, only a handful of functionally-conserved coronavirus structural RNA elements have been identified to date. Here, we performed RNA structure probing to obtain single-base resolution secondary structure maps of the full SARS-CoV-2 coronavirus genome both in vitro and in living infected cells. Probing data recapitulate the previously described coronavirus RNA elements (5' UTR and s2m), and reveal new structures. Of these, ∼10.2% show significant covariation among SARS-CoV-2 and other coronaviruses, hinting at their functionally-conserved role. Secondary structure-restrained 3D modeling of these segments further allowed for the identification of putative druggable pockets. In addition, we identify a set of single-stranded segments in vivo, showing high sequence conservation, suitable for the development of antisense oligonucleotide therapeutics. Collectively, our work lays the foundation for the development of innovative RNA-targeted therapeutic strategies to fight SARS-related infections.
MeSH term(s)	5' Untranslated Regions/genetics ; Algorithms ; Antiviral Agents/chemistry ; Antiviral Agents/metabolism ; Antiviral Agents/therapeutic use ; Base Sequence ; Binding Sites/genetics ; COVID-19/epidemiology ; COVID-19/prevention & control ; COVID-19/virology ; Conserved Sequence/genetics ; Genome, Viral/genetics ; Humans ; Models, Molecular ; Nucleic Acid Conformation ; Pandemics ; RNA, Viral/chemistry ; SARS-CoV-2/drug effects ; SARS-CoV-2/genetics ; SARS-CoV-2/physiology
Chemical Substances	5' Untranslated Regions ; Antiviral Agents ; RNA, Viral
Keywords	covid19
Language	English
Publishing date	2020-11-09
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkaa1053
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Article ; Online: Genome-wide mapping of therapeutically-relevant SARS-CoV-2 RNA structures

Manfredonia, Ilaria / Nithin, Chandran / Ponce-Salvatierra, Almudena / Ghosh, Pritha / Wirecki, Tomasz K. / Marinus, Tycho / Ogando, Natacha S. / Snider, Eric J. / van Hemert, Martijn J. / Bujnicki, Janusz M. / Incarnato, Danny

bioRxiv

Abstract: SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome of ∼30 kb, whose outbreak caused the still ongoing COVID-19 pandemic. The ability of coronaviruses to rapidly evolve, adapt, and cross species barriers makes the ... ...

Abstract	SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome of ∼30 kb, whose outbreak caused the still ongoing COVID-19 pandemic. The ability of coronaviruses to rapidly evolve, adapt, and cross species barriers makes the development of effective and durable therapeutic strategies a challenging and urgent need. As for other RNA viruses, genomic RNA structures are expected to play crucial roles in several steps of the coronavirus replication cycle. Despite this, only a handful of functionally conserved structural elements within coronavirus RNA genomes have been identified to date. Here, we performed RNA structure probing by SHAPE-MaP to obtain a single-base resolution secondary structure map of the full SARS-CoV-2 coronavirus genome. The SHAPE-MaP probing data recapitulate the previously described coronavirus RNA elements (5′ UTR, ribosomal frameshifting element, and 3′ UTR), and reveal new structures. Secondary structure-restrained 3D modeling of highly-structured regions across the SARS-CoV-2 genome allowed for the identification of several putative druggable pockets. Furthermore, ∼8% of the identified structure elements show significant covariation among SARS-CoV-2 and other coronaviruses, hinting at their functionally-conserved role. In addition, we identify a set of persistently single-stranded regions having high sequence conservation, suitable for the development of antisense oligonucleotide therapeutics. Collectively, our work lays the foundation for the development of innovative RNA-targeted therapeutic strategies to fight SARS-related infections.
Keywords	covid19
Publisher	BioRxiv; WHO
Document type	Article ; Online
DOI	10.1101/2020.06.15.151647
Database	COVID19

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Article: Genome-wide mapping of SARS-CoV-2 RNA structures identifies therapeutically-relevant elements

Nucleic acids res

Abstract	SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome, whose outbreak caused the ongoing COVID-19 pandemic. The ability of coronaviruses to rapidly evolve, adapt, and cross species barriers makes the development of effective and durable therapeutic strategies a challenging and urgent need. As for other RNA viruses, genomic RNA structures are expected to play crucial roles in several steps of the coronavirus replication cycle. Despite this, only a handful of functionally-conserved coronavirus structural RNA elements have been identified to date. Here, we performed RNA structure probing to obtain single-base resolution secondary structure maps of the full SARS-CoV-2 coronavirus genome both in vitro and in living infected cells. Probing data recapitulate the previously described coronavirus RNA elements (5' UTR and s2m), and reveal new structures. Of these, â¼10.2% show significant covariation among SARS-CoV-2 and other coronaviruses, hinting at their functionally-conserved role. Secondary structure-restrained 3D modeling of these segments further allowed for the identification of putative druggable pockets. In addition, we identify a set of single-stranded segments in vivo, showing high sequence conservation, suitable for the development of antisense oligonucleotide therapeutics. Collectively, our work lays the foundation for the development of innovative RNA-targeted therapeutic strategies to fight SARS-related infections.
Keywords	covid19
Publisher	WHO
Document type	Article
Note	WHO #Covidence: #917707
Database	COVID19

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Article ; Online: Genome-wide mapping of SARS-CoV-2 RNA structures identifies therapeutically-relevant elements

Nucleic Acids Research ; ISSN 0305-1048 1362-4962

2020

Abstract: Abstract SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome, whose outbreak caused the ongoing COVID-19 pandemic. The ability of coronaviruses to rapidly evolve, adapt, and cross species barriers makes the ... ...

Abstract	Abstract SARS-CoV-2 is a betacoronavirus with a linear single-stranded, positive-sense RNA genome, whose outbreak caused the ongoing COVID-19 pandemic. The ability of coronaviruses to rapidly evolve, adapt, and cross species barriers makes the development of effective and durable therapeutic strategies a challenging and urgent need. As for other RNA viruses, genomic RNA structures are expected to play crucial roles in several steps of the coronavirus replication cycle. Despite this, only a handful of functionally-conserved coronavirus structural RNA elements have been identified to date. Here, we performed RNA structure probing to obtain single-base resolution secondary structure maps of the full SARS-CoV-2 coronavirus genome both in vitro and in living infected cells. Probing data recapitulate the previously described coronavirus RNA elements (5′ UTR and s2m), and reveal new structures. Of these, ∼10.2% show significant covariation among SARS-CoV-2 and other coronaviruses, hinting at their functionally-conserved role. Secondary structure-restrained 3D modeling of these segments further allowed for the identification of putative druggable pockets. In addition, we identify a set of single-stranded segments in vivo, showing high sequence conservation, suitable for the development of antisense oligonucleotide therapeutics. Collectively, our work lays the foundation for the development of innovative RNA-targeted therapeutic strategies to fight SARS-related infections.
Keywords	Genetics ; covid19
Language	English
Publisher	Oxford University Press (OUP)
Publishing country	uk
Document type	Article ; Online
DOI	10.1093/nar/gkaa1053
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