RNA Biology Symposium 2018

SCIENTIFIC PROGRAM

DAY 1 - 13 September 2018

9:00AM 9:30AM REGISTRATION + COFFEE

9:30AM 9:45AM Opening Address:

Daniel TENEN Cancer Science Institute of Singapore

Session 1 | Chair Daniel TENEN Cancer Science Institute of Singapore

9:45AM 10:35AM KEYNOTE TALK:

Ada YONATH Weizmann Institute of Science, Israel

Origin of life, or: what was first the genetic code or its products?

10:35AM 11:05AM Martin JINEK University of Zurich, Switzerland

Structural insights into mRNA 3’ end formation

11:05AM 11:35AM Sudhakar JHA Cancer Science Institute of Singapore

circRNAs in Leukemia

11:35AM 1:45PM LUNCH

12:30PM 1:45PM POSTER SESSION I

Session 2 | Chair Wee Joo CHNG Cancer Science Institute of Singapore

1:45M 2:15PM Daniela RHODES Nanyang Technological University, Singapore

Telomerase structure and recruitment

2:15PM 2:45PM Jeffrey A. CHAO University of Basel, Switzerland

Imaging the life and death of mRNAs in single cells

2:45PM 3:15PM Amy PASQUINELLI University of California San Diego, USA

Short Poly(A) tails are a conserved feature of highly expressed genes

3:15PM 3:30PM SRBA ORAL PRESENTATION

3:30PM 4:00PM COFFEE BREAK

Session 3 | Chair Gene YEO University of California San Diego, USA

4:00PM 4:30PM Ling-Ling CHEN Shanghai Institutes for Biological Sciences, China

Unconventional RNA Form and Function

4:30PM 5:00PM Lei SUN Duke-NUS Medical School, Singapore

Explore the function of non-coding RNAs in adipose tissue

5:00PM 5:15PM SRBA ORAL PRESENTATION

RNA Biology Symposium 2018

SCIENTIFIC PROGRAM

DAY 2 - 14 September 2018

9:00AM 9:30AM ARRIVAL OF REGISTRANTS + COFFEE

Session 4 | Chair Polly CHEN Cancer Science Institute of Singapore

9:30AM 10:00AM Mary O’CONNELL Central European Institute of Technology (CEITEC), Czech Republic

Deciphering a novel role of the RNA editing enzyme Adar in Drosophila development.

10:00AM 10:30AM Meng How TAN

Nanyang Technological University, Singapore

Dynamic RNA editing landscape in mammals

10:30AM 11:00AM Ramanuj DASGUPTA Genome Institute of Singapore, A*STAR, Singapore

Dynamic expression of tRNA-derived small RNAs define cellular states

11:00AM 11:15AM SRBA ORAL PRESENTATION

11:15PM 1:15PM LUNCH WITH EXPERTS

12:00PM 1:15PM POSTER SESSION II

Session 5 | Chair Dahai LUO Nanyang Technological University, Singapore

1:15PM 1:45PM Ralf BARTENSCHLAGER University of Heidelberg, Germany

Counteraction of innate antiviral defense by persistent hepatitis viruses

1:45PM 2:15PM Julien LESCAR Nanyang Technological University, Singapore

The dengue virus replication complex: from RNA replication to protein-protein interactions to evasion of innate immunity

2:15PM 2:45PM Sherry AW Institute for Molecular and Cell Biology, A*STAR, Singapore

MicroRNAs in neurodegeneration: A genetic screen and a Spinach-based sensor

2:45PM 3:00PM SRBA ORAL PRESENTATION

3:00PM 3:30PM COFFEE BREAK

Session 6 | Chair Yue WAN Genome Institute of Singapore, A*STAR, Singapore

3:30PM 4:00PM Leah VARDY Institute of Medical Biology, A*STAR, Singapore

Polyamines: Critical regulators of epidermal function

4:00PM 4:50PM KEYNOTE TALK:

Paul AGRIS Duke University School of Medicine, USA

Small molecule antibacterial agents targeting a novel RNA foiling the emergence of resistance

4:50PM 5.10PM Announcement of Poster Competition Results & Closing Address:

Yue WAN

Genome Institute of Singapore, A*STAR, Singapore

DAY 1 | 13 SEPTEMBER 2018

KEYNOTE TALK 1

Ada YONATH Director, the Kimmelman Center for Biomolecular Assemblies, WIS

The Martin S. and Helen Kimmel Professor of Structural Biology

Department of Structural Biology

Weizmann Institute, Rehovot, Israel

ORIGIN OF LIFE, OR: WHAT WAS FIRST THE GENETIC CODE OR ITS PRODUCTS?

Ribosomes, the universal cellular machines for translation of the genetic code into proteins, possess

spectacular architecture accompanied by inherent mobility, allowing for their smooth performance

as polymerases that translate the genetic code into proteins. The site for peptide bond formation is

located within an almost fully conserved internal semi-symmetrical pocket composed exclusively of

RNA. The high conservation of this region implies its existence irrespective of environmental

conditions and indicates that it may represent an ancient RNA machine, which could be the kernel

around which life originated. Hence, called by us the “proto ribosome”. Recently, the validity of this

suggestion was verified, for the first time, by the laboratory formation of a peptide bond by a

synthetic “proto ribosome”, thus indicating that the vestige of a molecular prebiotic bonding entity

is still functioning in all living cells of all organisms. As the initial dipeptide could be elongated by the

proto ribosome to oligopeptides, those fulfilling crucial tasks in the probiotic world or stabilizing the

proto ribosome, survived and led to the creation of a genetic code, which evolved together with the

proto-ribosome and its products, the proteins.

BIOSKETCH

Ada Yonath focuses on genetic code translation by ribosomes, on antibiotics paralyzing this process,

on antibiotic resistance, on designing novel antibiotics and on origin of life. She graduated from

Hebrew University, earned her PhD from Weizmann Institute (WIS) and completed postdoctoral

studies at CMU and MIT, USA. In 1971 she established the first biological-crystallography laboratory

in Israel, which was the only lab of this kind in the country for almost a decade. Since then, she has

been a faculty member and the Director of Kimmelman Center for Biomolecular Structures at WIS.

In 1978 she spent a Sabbatical in the Chicago University, and during 1980-2004 she headed the Max-

Planck-Research-Unit for Ribosome Structure in Hamburg in parallel to her WIS activities. Among

others, she is a member of US-National-Academy-of-Sciences; Israel Academy of Sciences-and-

Humanities; German Academy for Sciences (Leopoldina); European Molecular Biology Organization;

Pontifical (Vatican) Academy of Sciences. She holds honorary doctorates from over 20 universities

worldwide, in Iarael, USA, Latin America, Europe and the Far East.

Her awards include the Israel Prize; Linus Pauling Gold Medal; Albert Einstein World Award for

Excellence; UNESCO-L'Oréal Award; Wolf Prize; Louisa Gross Horwitz Prize; Erice Peace Prize; Indian

Prime-minister medal and the Nobel Prize for Chemistry.

Martin JINEK Professor

Department of Biochemistry

University of Zurich, Switzerland

STRUCTURAL INSIGHTS INTO MRNA 3’ END FORMATION

3' polyadenylation is a key step in eukaryotic mRNA biogenesis. In mammalian cells, this process is

dependent on the recognition of the hexanucleotide AAUAAA motif in the pre-mRNA

polyadenylation signal by the cleavage and polyadenylation specificity factor (CPSF) complex. A core

CPSF complex comprising CPSF160, WDR33, CPSF30 and Fip1 is sufficient for AAUAAA motif

recognition, yet the molecular interactions underpinning its assembly and mechanism of PAS

recognition are not understood. Based on cross-linking-coupled mass spectrometry, crystal

structure of the CPSF160-WDR33 subcomplex and biochemical assays, we defined the molecular

architecture of the core human CPSF complex, identifying specific domains involved in inter-subunit

interactions and RNA binding. We subsequently used cryo-electron microscopy to determine the 3.1

Å-resolution structure of the core CPSF complex bound to the AAUAAA hexanucleotide. Collectively,

these studies reveal the molecular interactions responsible for sequence-specific recognition of the

polyadenylation signal hexamer by the mammalian CPSF complex and provide a rationale for the

mechanistic differences between mammalian and yeast mRNA polyadenylation.

BIOSKETCH

Martin Jinek is an Assistant Professor in the Department of Biochemistry at the University of Zurich.

His research focuses explores two main topics – (i) CRISPR-Cas systems and their use as a genome

editing technology, and (ii) RNA processing and modification pathways in eukaryotic gene

expression. Martin Jinek studied Natural Sciences at the University of Cambridge and obtained his

PhD from the European Molecular Biology Laboratory in Heidelberg. His postdoctoral research with

Prof. Jennifer Doudna at the University of California, Berkeley, led to the discovery of the

biochemical function of the CRISPR-associated endonuclease Cas9 and was pivotal for establishing

CRISPR-Cas9 genome editing. Since starting his research group at the University of Zurich in 2013,

Martin Jinek has used structural and biochemical approaches to study the molecular mechanisms of

CRISPR-Cas genome editor nucleases as well as macromolecular complexes involved in eukaryotic

mRNA metabolism. In recognition of his work, Martin Jinek has been awarded an ERC Starting Grant

(2013), the EMBL John Kendrew Young Scientist Award (2014) and the Friedrich Miescher Award of

the Swiss Society for Molecular and Cellular Biosciences (2015). He is an EMBO Young Investigator

and in 2017 became an International Research Scholar of the Howard Hughes Medical Institute.

Sudhakar JHA Principal Investigator

Cancer Science Institute of Singapore

National University of Singapore, Singapore

CIRCRNAS IN LEUKEMIA

Circular RNAs (circRNAs) are noncoding RNAs generated as a result of ligation between a

downstream splice donor to an upstream splice acceptor. CircRNAs are speculated to have varied

functions ranging from microRNA regulation, cell proliferation, parental gene expression and

regulation of RNA-binding proteins. Circular RNA biology remains a relatively under explored field

with exciting opportunities in the context of cancer biology and their potential as prognostic and

diagnostic tools. Leukemia is a cancer of the blood cells which arises in the bone marrow and results

in an excessive accumulation of white blood cells in the blood stream. Leukemia affects both

children and adults and may be chronic or acute. In this conference, I will discuss identification of

circular RNA generated from the Additional Sex Combs like-1 (ASXL1) gene, one of the genes

implicated in leukemia. CircASXL1 is expressed in variety of AML and CML leukemia cell lines.

Interestingly, depletion of circASXL1 leads to changes in activity of enzymes involved epigenetic

pathways by regulating ASXL1 function.

BIOSKETCH

Dr. Sudhakar Jha is a Principal Investigator at Cancer Science Institute of Singapore, and Assistant

Professor in Department of Biochemistry, YLL School of Medicine at the National University of

Singapore. His group is interested in understanding the regulation of chromatin remodeling

complexes and their role in cancer prevention and intervention (Mol Cell 2009, 34: 521-533). Dr.

Jha’s group focuses on identifying the role of TIP60, a lysine acetyltransferase in transcription (J Mol

Cell Biol 2016, 85: 384–399) and DNA damage response pathway (Mol Cell Biol 2008, 28: 2690-2700

and Mol Cell Biol 2013, 33: 1164-74). Among various regulators of TIP60, Dr. Jha’s group has

discovered human papillomavirus (HPV) E6 and Adenovirus (AdV) oncogenes to destabilize TIP60

(Mol Cell 2010, 38: 700-711, Oncogene 2013, 32: 5017-25 and Oncogene 2016, 35:2062-74). In

addition, his group has also identified a new cellular regulator of TIP60 and have demonstrated its

role and significance in epithelial-mesenchymal transition and breast cancer progression

(Oncotarget 2015, 6:41290-306 and J Mol Cell Biol 2016, 85: 384–399).

Daniela RHODES Professor, School of Biological Sciences and

School of Chemical and Biomedical Engineering

Director, NTU Institute of Structural Biology

Nanyang Technological University, Singapore

TELOMERASE STRUCTURE AND RECRUITMENT

Telomeres, the protein/DNA complexes that cap the ends of eukaryotic chromosomes are essential

for genomic stability and cell viability. The mechanism conserved throughout eukaryotes for

telomere maintenance is by the specialized reverse transcriptase enzyme telomerase that consists

of an RNA subunit TR containing the templet for telomeric DNA synthesis and the catalytic protein

subunit TERT that. Telomerase is expressed during early development and remains active in specific

germ -line cells and toti-potent embryonic stem-cells, but is undetected in most normal somatic cells

leading to telomere shortening, replicative senescence and aging Reactivation of telomerase is a key

requisite in over 90% of human cancers to attain unlimited cell proliferation. Therefore, telomerase

is implicated in both cancer and aging. To obtain a full understanding of the mechanisms of both

telomerase action and recruitment, structural information is required. I will describe our efforts in

determining the three-dimensional structure of both human telomerase and the telomerase

recruitment complex using single-particle cryo-EM.

BIOSKETCH

Daniela Rhodes is a structural biologist and has made important contributions to understanding

chromosome structure and function. She spent most of her scientific career at the world-renowned

MRC Laboratory of Molecular Biology in Cambridge, UK where she obtaining her PhD in 1982 under

the guidance of Nobel Prize winner, Sir Aaron Klug. Since 2011 she holds a joint professorship with

the School of Biological Sciences and the School of Chemical and Biomedical Engineering at Nanyang

Technological University, as well as being the director of the NTU Institute of Structural Biology. Her

scientific achievements have been recognized by being elected: Official Fellow Clare Hall, Cambridge,

UK (1992); EMBO Member (1996); Fellow of the Royal Society, UK (2007) ; Member of the Academia

Europaea (2011) and Ponte d’Oro Prize (2011)

Jeffrey A.CHAO Group Leader

Friedrich Miescher Institute for Biomedical Research, Basel

IMAGING THE LIFE AND DEATH OF MRNAS IN SINGLE CELLS.

After transcription, an mRNA's fate is determined by an orchestrated series of events (processing,

export, localization, translation and degradation) that is regulated both temporally and spatially

within the cell. In order to more completely understand these processes and how they are coupled,

it is necessary to be able to observe these events as they occur on single molecules of mRNA in real-

time in living cells. To expand the scope of questions that can be addressed by RNA imaging, we are

developing multi-color RNA biosensors that allow that status of a single mRNA molecules (e.g.

translation or degradation) to be directly visualized and quantified.

In order to image the first round of translation, we have developed TRICK (translating RNA imaging

by coat protein knock-off) which relies on the detection of two fluorescent signals that are placed

within the coding sequence and the 3′UTR. In this approach, an untranslated mRNA is dual labeled

and the fluorescent label in the coding sequence is displaced by the ribosome during the first round

of translation resulting in translated mRNAs being singly labeled. A conceptually similar approach

was used for single-molecule imaging of mRNA decay, where dual-colored mRNAs identify intact

transcripts, while a single-colored stabilized decay intermediate marked degraded transcripts

(TREAT, 3′ RNA end accumulation during turnover). We are using these tools to characterize

localized translation and degradation during normal cell growth and stress.

BIOSKETCH

Jeff Chao obtained his PhD from The Scripps Research Institute in La Jolla, CA where he worked with

James Williamson on the structure and function of RNA-protein complexes. His postdoctoral studies

with Robert Singer at Albert Einstein College of Medicine in Bronx, NY focused on characterization

of mRNPs involved in RNA localization and developing fluorescent microscopy techniques for

imaging single mRNAs. In 2013, he established his own group at the Friedrich Miescher Institute for

Biomedical Research in Basel, Switzerland. His group has recently described fluorescent imaging

methodologies that enable the time and location of the first round of translation (TRICK) and

degradation (TREAT) of single mRNAs within a living cell to be measured. His laboratory continues

to investigate the mechanisms that control post-transcriptional regulation in the cytoplasm.

Amy PASQUINELLI Professor

Division of Biological Sciences

University of California, San Diego, USA

SHORT POLY(A) TAILS ARE A CONSERVED FEATURE OF HIGHLY EXPRESSED GENES

The poly(A) tails appended to the 3’ ends of most eukaryotic mRNAs play important roles in

translation and stability. However, recent genome-wide studies concluded that poly(A) tail length

was generally not associated with translational efficiency in non-embryonic cells. To investigate if

poly(A) tail size might be coupled to gene expression in an intact organism, we used an adapted

TAIL-seq protocol to measure poly(A) tails in larval stage Caenorhabditis elegans. Surprisingly, we

found that well-expressed transcripts contain relatively short, well-defined tails that would likely

accommodate only 1-2 poly(A) binding proteins (PABPs). This attribute appears dependent on

translational efficiency, as transcripts enriched for optimal codons and ribosome association had the

shortest tail sizes, while non-coding RNAs retained long tails. Across eukaryotes, short tails were a

feature of abundant and well-translated mRNAs. However, for these genes and almost all others,

we were still able to detect transcripts with tail lengths consistent with the very long (>200 nt) poly(A)

tails synthesized on nascent mRNAs. The finding that genes with the highest frequencies of optimal

codons were represented by mRNAs that spanned the entire range of detectable tail sizes, but were

strongly biased for short tailed species, suggests that well-expressed mRNAs undergo poly(A) tail

shortening to an optimal length, which we refer to as pruning. The hallmarks of pruning are that

poly(A) tails are well-defined and relatively short, while tails on mRNAs enriched for suboptimal

codons are more heterogeneous and less defined, showing a spread across the range of possible

sizes. Although this seems to contradict the dogma that deadenylation induces translational

inhibition and mRNA decay, it instead suggests that well-expressed mRNAs accumulate with pruned

tails that accommodate a minimal number of PABPs, which may be ideal for protective and

translational functions.

BIOSKETCH

Dr. Amy Pasquinelli is a Professor of Biology at the University of California, San Diego. The overall

goal of research in the Pasquinelli lab is to understand how post-transcriptional regulation of gene

expression contributes to organismal development and viability. To date, we have primarily used C.

elegans as a model animal system to investigate how microRNAs (miRNAs) and other non-coding

RNAs (ncRNAs) are expressed and regulate gene expression in an endogenous context. The

pathways being studied are broadly conserved throughout animal phylogeny and relevant to

understanding the role of ncRNAs in human development and disease. Dr. Pasquinelli received a B.A.

in Biology from Bucknell University, a Ph.D. in Biomolecular Chemistry from the University of

Wisconsin, Madison, and did postdoctoral training at Harvard Medical School. Research in the

Pasquinelli lab has been funded by the National Institutes of Health (NIH), American Federation for

Aging Research (AFAR), Keck, Searle Scholar, V, Peter Gruber and Emerald Foundations.

SRBA ORAL PRESENTATION

UTILIZATION OF A NOVEL TECHNIQUE TO IDENTIFY RNA-BINDING PROTEINS

Larry Ng1,2, Shweta Jadhav1, Dennis Kappei1 and Sudhakar Jha1,2

1Cancer Science Institute of Singapore, 2Department of Biochemistry, Yong Lin Lee School of Medicine,

National University of Singapore, Singapore

Ribonucleic acid (RNA)-protein interactions are a set of highly dynamic and intricate intracellular

network that play key roles in cellular processes such as mRNA splicing, translation and degradation.

Disruption of the RNA-protein interaction network can lead to serious health consequences, often

leading to diseases such as cancer. Over the years, various in vivo and in vitro strategies were

developed to purify and identify RNA-binding proteins (RBPs), with the purpose of deciphering

mechanisms regulating messenger RNA (mRNA) or the processes regulated by the mRNA in the

context of diseases. However, some existing strategies face challenges in terms of data reliability

and comprehensiveness of RBPs identification. It is therefore crucial to explore and develop a more

robust, comprehensive and reliable approach to purify and identify RBPs. In this study, we

developed dCas9 RNA immunoprecipitation (dCaRIP), which is method that couples a modified

CRISPR-dCas9 system and immunoprecipitation to identify RBPs associated with an RNA transcript

of interest.

Ling-Ling.CHEN Professor

Institute of Biochemistry and Cell Biology

Shanghai Institutes for Biological Sciences

Chinese Academy of Sciences, China

UNCONVENTIONAL RNA FORM AND FUNCTION

Long noncoding RNAs (lncRNAs) comprise different types of RNA polymeraseII - derived noncoding

transcripts with sizes that are greater than 200 nt in length. While a large proportion of lncRNAs look

like mRNAs, a number of lncRNAs form their ends in unusual ways. We have been working on the

biogenesis and functional implication of distinct lncRNA species with small nucleolar RNA (snoRNA)

ends, including the box C/D snoRNA-ended lncRNAs (sno-lncRNAs), the box H/ACA snoRNA-ended

lncRNAs (such as SLERT), and the 5’ snoRNA-ended and 3’ polyadenylated lncRNAs (SPAs). I will

update our current understanding of these sno-processed lncRNAs with a focus on the

recently characterized SLERT and SPAs.

BIOSKETCH

Dr. Ling-Ling Chen carried out the doctoral and post-doctoral work at the University of Connecticut

Health Center, USA from 2004 and 2010. She has been the Principal Investigator at the Shanghai

Institute of Biochemistry and Cell Biology (SIBCB), Chinese Academy of Sciences (CAS) since 2011

and has selected as an HHMI international research scholar since 2017. Ling-Ling Chen’s group has

identified several unconventional types of long noncoding RNA (lncRNA) species by developing

methods to explore the non-polyadenylated transcriptomes. Her group now studies the biogenesis

and functional significance of these lncRNAs in different cellular contexts and relevant human

diseases.

Lei SUN Associate Professor

Cardiovascular & Metabolic Disorders Programme

Duke-NUS Medical School, Singapore

EXPLORE THE FUNCTION OF NON-CODING RNAS IN ADIPOSE TISSUE

Modern sedentary lifestyle and consumption of calorie-dense food are precipitating a rapid growing

population of metabolic diseases such as obesity, type 2 diabetes and heart diseases. It is predicted,

for the first time, that the current generation will have a shorter life-span than previous one.

Understanding the molecular mechanisms underlying these metabolic diseases is urgently needed

for us to develop novel therapeutic strategies. Our studies are revealing a long non-coding RNAs

(lncRNAs)-mediated regulatory network governing the development and function of adipose tissue

at various physiological and pathological conditions. We have depicted comprehensive non-coding

transcriptomes in adipose tissues from different depots, during browning as well as obesity, and

have identified several key lncRNAs that can regulate adipocytes’ function. These studies have

advanced our fundamental understanding of adipocyte biology and opened new avenues to improve

metabolic health.

BIOSKETCH

Dr SUN Lei is an Associate Professor in the program of Cardiovascular and Metabolic Disorders,

Duke-NUS Medical School. He received his B.S degree from Beijing University in 2001 and Ph.D in

Biochemistry from Case Western University in 2008. From 2008 to 2012, he underwent postdoctoral

training in the lab of Harvey Lodish at the Whitehead Institute in Boston, MA. In 2012, he was

awarded an NRF fellowship award in Singapore and joined the faculty at Duke-NUS. His research

focuses on the RNA-regulatory network governing the development and function of major metabolic

organs such as adipose and liver at various physiological and pathological conditions.

SRBA ORAL PRESENTATION

A NOVEL AND EVOLUTIONARY CONSERVED LNCRNA IS ESSENTIAL FOR POSITIVE

REGULATION OF NFΚB

Bilal Unal1,2 , Semih Can Akincilar1, Lele Wu1, Eun Myoung Shin1,3, Surendar Aramugam1, Zahra

Eslami-s1, Ambarnil Ghosh1, Anandhkumar Raju, Hannah Lee Foon Swa4, Manikandan Lakshmanan1,

Jayantha Gunaratne4,5, Yong Jae Shin6,7, Yeri Lee6, Jason K. Sa6, Yunmi Kim6, Do-Hyun Nam7,8, Vinay

Tergaonkar1,2,3,9*

1Laboratory of NFκB Signalling, Institute of Molecular and Cell Biology (IMCB), A*STAR (Agency for Science, Technology and Research), Singapore 138673, Singapore. 2Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore (NUS),

Singapore 117597, Singapore. 3Cancer Science Institute of Singapore, Singapore 117599, Singapore 4Laboratory of Translational Biomedical Proteomics, Institute of Molecular and Cell Biology (IMCB), A*STAR (Agency for Science, Technology and Research), Singapore 138673, Singapore. 5Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore (NUS), Singapore 117597, Singapore 6Research Institute for Future Medicine, Samsung Medical Center, Seoul 06351, Korea 7Department of Neurosurgery, Samsung Medical Center, Seoul 06351, Korea 8Department of Health Sciences and Technology (SAIHST), Sungkyunkwan University School of Medicine, Seoul 06351, Korea 9Centre for Cancer Biology (University of South Australia and SA Pathology), Adelaide, SA 5000, Australia

Many proteins that regulate inflammation downstream of master transcription factors like NFκB

have been identified and some targeted successfully. However, only about two percent of the

human genome is transcribed into RNAs which make proteins while a majority of the remaining has

now been shown to encode various classes of non-coding RNAs. Long non-coding RNAs (lncRNAs)

constitute a significant proportion of non-coding RNAs, and their function in regulation of NFκB

biology is largely unexplored. Here, using the first genetic screen to identify NFκB specific lncRNAs,

we report the identification of a novel and evolutionary conserved lncRNA designated lncRNA-GM

(in mice) or lncRNA-LOC (in humans). CRISPR-Cas9 mediated deletion of conserved NFκB binding

sites in the promoters of these lncRNAs mitigates NFκB responses, downstream of a number of

physiologically relevant stimuli. Loss of lncRNA-GM reduces tumorigenesis and metastasis in mice

and higher levels of lncRNA-LOC predicts poorer survival in human glioblastomas. Targeting lncRNA-

LOC causes loss of p65 phosphorylation and activation leading to genome-wide reduced p65

occupancy on subsets of target sites such as those of IL-8 and ICAM-1, cytokines essential for

inflammation and cancer progression. lncRNA-LOC primarily localizes in the nucleus and directly

interacts with ATP-dependent RNA helicase DHX15 protein. LOC lncRNA-DHX15 complex sequesters

a key negative regulator of p65 activation, namely Wip1 phosphatase. Activation of lncRNA-LOC by

NFκB controls the length of the time p65 can remain phosphorylated and hence productively engage

in transcription. This positive feed-forward loop between NFκB and lncRNA-LOC highlights the

importance of lncRNAs as modulators of phosphatases in inflammatory responses.

DAY 2 | 14 SEPTEMBER 2018

Mary O’CONNELL ERA Chair

Central European Institute of Technology (CEITEC)

Masaryk University, Czech Republic

DECIPHERING A NOVEL ROLE OF THE RNA EDITING ENZYME ADAR IN DROSOPHILA

DEVELOPMENT

One of the most prevalent type of RNA editing is the conversion of adenosine to inosine in double-

stranded RNAs that is mediated by adenosine deaminase acting on RNA (ADAR) enzymes. A→I RNA

editing can lead to a codon change as the nucleoside inosine (I) is interpreted as guanosine (G) by

the cellular machines, resulting in a diversification of protein function. The ADAR family of proteins

is present in all metazoans. In Drosophila, a single Adar is present at the tip of X chromosome and is

an orthologue of vertebrate ADAR2. In spite of major progress in the identification of editing sites,

little is known about the regulatory mechanism of ADAR proteins in normal development and in

disease. In this present study, we performed a genetic screen that have uncovered a novel role of

Adar in regulating ecdysone signaling which is a crucial regulator of Drosophila development.

Ubiquitous expression of Adar with act5c-Gal4 results in pupal lethality. Tissue specific over-

expression of Adar in the Prothoracic Gland (PG) with phm-Gal4 shows a significant delay in pupation,

due to a complete blocking of ecdysone synthesis and signaling. These defects may be due to either

aberrant RNA editing or RNA binding by ADAR protein. The lethality caused by ubiquitous expression

of Adar can be rescued by blocking ecdysone synthesis and signaling. We hypothesize that Adar

expression in Drosophila is a prerequisite to regulate ecdysone signaling during metamorphosis.

Currently, we are dissecting regulation of the ecdysone pathway by Adar and pursuing loss of

functions studies with Adar RNAi lines to decipher role of Adar in metamorphosis of Drosophila.

BIOSKETCH

Mary O’ Connell did her undergraduate at University College Galway, followed by a PhD at Albert

Einstein College of Medicine, New York. Her first postdoctoral work was at MIT with Prof. Nancy

Hopkins. She then went to the laboratory of Prof. Walter Keller at the Biozentrum in Basel where

she purified and cloned ADAR1 and ADAR2. She was a Group Leader at the MRC Human Genetics

Unit, Edinburgh from 1997-2013. From 2014, she has been ERA Chair at CEITEC, Masaryk University,

Czech Republic. She was elected EMBO member in 2017.

Meng How TAN Assistant Professor

School of Chemical and Biomedical Engineering

Nanyang Technological University, Singapore

Dynamic RNA Editing Landscape in Mammals

RNA can be post-transcriptionally modified in more than a hundred different ways. One of the most

prevalent modifications in the epitranscriptome is the inosine, which is typically interpreted by

cellular machineries as a guanosine. Inosines are formed by a deamination reaction on adenosines

and this reaction is carried out the conserved ADAR family of enzymes. Although there are three

ADAR genes encoded in mammalian genomes, only ADAR1 and ADAR2 are catalytically active. In

this talk, I will describe some of our work on understanding the adenosine-to-inosine (A-to-I) RNA

editing landscape in mammals. Over the past few years, we have systematically profiled hundreds

of normal and diseased samples and uncovered numerous editing patterns across tissues and over

development. Since ADAR1 and ADAR2 alone cannot possibly account for all the patterns that we

observed, we postulate that there must be other trans-acting factors that help shape the editing

landscape. So far, we have characterized two RNA-binding proteins that perform non-canonical

functions in regulating editing levels in mammals. Our research serves to deepen our understanding

of A-to-I RNA editing and the epitranscriptome in general.

BIOSKETCH

Meng How Tan is currently an Assistant Professor in the School of Chemical and Biomedical

Engineering at Nanyang Technological University (NTU) as well as a Senior Research Scientist in the

Genome Institute of Singapore at Agency for Science Technology and Research (A*STAR). Prior to

setting up his laboratory in Singapore, he received a B.S. degree in mechanical engineering and a

B.A. degree in economics from University of California, Berkeley, a M.S. degree in aeronautics from

California Institute of Technology, a M.S. degree in biomedical engineering from NTU, and a Ph.D. in

developmental biology from Stanford University. He also performed postdoctoral research on

genomics and stem cells with Jin Billy Li, Mylene Yao, and Wing Hung Wong at Stanford University.

Currently, his laboratory is interested in understanding how biological information hardwired in the

genome of living cells can be permanently or transiently altered at both the DNA and RNA levels

Ramanuj DASGUPTA Senior Investigator

Genome Institute of Singapore

A*Star, Singapore

DYNAMIC EXPRESSION OF TRNA-DERIVED SMALL RNAS DEFINE CELLULAR STATES

Transfer RNA (tRNA)-derived small RNAs, referred to as tsRNAs or tRFs (tRNA-fragments) have

recently emerged as important regulators of protein translation and shown to have diverse

biological functions. However, the underlying cellular and molecular mechanisms of tsRNA function

in the context of dynamic cell-state transitions remain unclear. In this study we report the

identification of a set of tsRNAs (31-33nt 5’ halves) that are upregulated upon RNA-induced

differentiation of mouse embryonic stem cells (mESCs). Mechanistic analyses revealed primary

functions of tsRNAs in regulating polysome assembly and translation. Notably, interactome studies

with differentially enriched tsRNAs revealed a switch in associations with ‘effector’ RNPs and ‘target’

mRNAs in different cell-states. We demonstrate that a specific pool of tsRNAs can interact with

effector RNPs to influence the expression of the pluripotency-promoting factors, such as cMyc,

thereby providing a mechanistic basis for how tsRNAs may modulate cell-states in mESCs. Finally,

tsRNA expression analyses in distinct, heterologous cell and tissue models of stem/transformed

versus differentiated/normal states reveal that tsRNA-mediated regulation of protein translation

may represent a global biological phenomenon associated with cell-state transitions.

BIOSKETCH

Dr DasGupta is a Senior Investigator at the Genome Institute of Singapore (Cancer Therapeutics and

Stratified Oncology), and holds an adjunct Associate Professor position at CSI-NUS. He also holds an

Associate Professor position at the New York University Cancer Institute, New York. The major focus

in the DasGupta laboratory is to implement “Phenotype-driven Precision Oncology” in the clinic by

establishing the next-generation of HTS-amenable patient-personalized cancer models to identify

novel therapeutic opportunities, and biomarkers. The overall research goal is to define the function,

and underlying mechanisms of intra-tumor heterogeneity (ITH) and tumor evolution in the

acquisition of treatment resistant, and metastatic phenotypes. A wide variety of functional genomic

approaches are employed including single-cell/bulk transcriptomics, epigenomics, and exome-seq

to address questions related to how individual cells evolve/acquire metastatic and resistant

phenotypes. In addition, the DasGupta lab is also interested in the identification and functional

characterization of non-coding RNAs (ncRNAs), and associated RNA-binding proteins (RNPs) in the

regulation of oncogenic signaling pathways, as well as in defining specific developmental cell-states

both in normal and cancer stem cells (CSCs).

SRBA ORAL PRESENTATION

THE HUMAN TRNA EPITRANSCRIPTOME: A PIVOT FOR MODULATION BY BOTH HOST AND

VIRUS

Cheryl Chan1, Thomas J. Begley2, Peter C. Dedon1,3

1Singapore-MIT Alliance for Research and Technology, Singapore 138602; 2College of Nanoscale Science and

Engineering, State University of New York, Albany, NY 12203; The RNA Institute, College of Arts and Science,

University at Albany, SUNY, Albany, NY 12222; 3Department of Biological Engineering, Massachusetts

Institute of Technology, Cambridge, MA 02139.

Emerging evidence points to the central role of the tRNA epitranscriptome – the collection of

ribonucleoside modifications of tRNA – in cellular translational adaptation to various cell stresses.

This is pertinent during virus infection stress in human cells where reprogramming the human tRNA

epitranscriptome is required to translate cellular stress response proteins, as well as viral proteins

by accommodating the inherent mismatch in the virus and host codon usage. Using a dengue

serotype 2 virus infection model of human liver Huh-7 cells in culture, we profiled changes in more

than 30 ribonucleoside modifications of human tRNA during infection by liquid chromatography

coupled with tandem mass spectrometry. We observed an increase in wobble uridine mcm5s2 and

ncm5 modification levels during dengue virus infection, consistent with the virus’ need to

preferentially decode A-ending cognate codons – a signature of dengue virus codon usage.

Conversely, a marked decrease in the levels of the AUA-decoding wobble f5C modification was found

to be associated with the host’s defense to limit virus translation during infection. RNAi-mediated

depletion of the f5C writer ALKBH1 reduced viral protein translation while restoration of f5C levels

by ALKBH1 transient overexpression enhanced viral protein translation and production of new

infectious viruses. Further, codon usage analysis of deregulated proteins during dengue virus

infection revealed enrichment of A/T- and G/C-ending codon subsets involved in virulence and host

defense respectively. Together, these findings provide insights to the functional roles of human

tRNA epitranscriptome reprogramming during dengue virus infection and offer potential targets for

host-directed antiviral therapeutics.

Ralf BARTENSCHLAGER Professor, Dr

Department for Infectious Diseases, Molecular Virology

University of Heidelberg, Germany

COUNTERACTION OF INNATE ANTIVIRAL DEFENSE BY PERSISTENT HEPATITIS VIRUSES

Infections with the Hepatitis B and C virus (HBV, HCV) are a major risk factor for chronic liver disease,

with both viruses having a high propensity to establish persistence. While HCV is a positive-strand

RNA virus replicating in the cytoplasm in membranous replication organelles, HBV is a pararetrovirus,

replicating its pregenomic RNA via reverse transcription within the nucleocapsid. To establish

persistence, both viruses have developed efficient strategies to overcome innate antiviral immunity.

In the case of HCV we found that it blocks the interferon activation pathway via MAVS by proteolytic

cleavage of this signaling molecule. However, it still induces a strong interferon response in vitro

and in vivo and is highly sensitive to the antiviral program induced by this cytokine. Moreover, HCV

does not actively suppress the TLR3 signaling pathway but keeps the TLR3-induced response low via

release of exosomes containing viral replication intermediates. In contrast, HBV is a prototypic

“stealth” virus passively bypassing the interferon system at all levels of sensors and antiviral

effectors. This might be the result of the long-term coevolution of HBV with its host over geologic

eras.

BIOSKETCH

Ralf Bartenschlager is molecular biologist by training and interested in the complexities of the

interactions between viruses and their host cells. His work centers on hepatitis viruses, notably

hepatitis C and B virus (HCV and HBV, respectively) and comparative analyses with flaviviruses

(Dengue and Zikavirus). One research direction in the Bartenschlager lab deals with the strategies

used by HCV and HBV to establish persistence with a focus on the innate antiviral defense. Another

direction centers on the cell biology of the replication cycle of these viruses, how they exploit host

cell factors and pathways for efficient replication and how this relates to virus – host evolution.

Finally, knowledge gained from these studies is used to develop novel antiviral strategies, focusing

on host cell dependency factors that hold promise for the development of broad-spectrum antiviral

drugs.

Julien LESCAR Associate Professor

School of Biological Sciences

Nanyang Technological University, Singapore

THE DENGUE VIRUS REPLICATION COMPLEX: FROM RNA REPLICATION TO PROTEIN-

PROTEIN INTERACTIONS TO EVASION OF INNATE IMMUNITY

Viruses from the Flavivirus family are the causative agents of major or emerging public health

problems such as dengue fever, Zika, Japanese encephalitis, West Nile encephalitis or Yellow fever.

A better understanding of how flavivirus replicate is likely to stimulate the design of antiviral

therapies. During flavivirus replication, RNA synthesis is mediated by a dynamic multi-protein

assembly attached to the endoplasmic reticulum membrane, named the replication complex (RC).

The RC is composed of both viral and host-cell proteins and assemble within vesicles near the

nucleus. At the heart of the flavivirus RC lies the large NS5 methyl-transferase polymerase. NS5 is a

large and dynamic protein that represents a key target for antiviral design. We will present a new

crystal structure of the full-length NS5 protein from Dengue virus serotype 2 that has implications

for the possible evolution of the flavivirus family. We will also present an overview of our recent

drug discovery efforts targeting NS5. A summary of what we know about the network of interactions

established by NS2B-NS3, NS4B and NS5 (their “interactome”) will be given. This leads to a working

model of part of the RC that can be refined and tested in the near future.

BIOSKETCH

Julien Lescar (NTU) is a structural biologist working in Singapore for 15 years. He established the

laboratory of X-ray crystallography at NTU. He is actively involved in research programs on infectious

diseases caused by RNA viruses (DENV, ZIKV and CHIKV). He has been collaborating with the Novartis

Institute for Tropical Diseases, Duke-NUS and SMART on structure-based drug discovery programs

for antiviral compounds, especially against enzymes involved in RNA modification. His group

published several first structures for the RNA helicase and RdRp from DENV.

Sherry AW Group Leader

Institute of Molecular and Cell Biology

A*Star, Singapore

MICRORNAS IN NEURODEGENERATION: A GENETIC SCREEN AND A SPINACH-BASED

SENSOR

My lab studies the molecular mechanisms that underlie neurodegeneration, using the fruitfly

Drosophila. Drosophila has a million-fold fewer brain cells than humans, yet share 60% of our genes.

Despite the differences in anatomy and scale between human and fly brains, Drosophila disease

models have turned out to be good paradigms for human neurological disorders, and fly models

have made invaluable contributions to our understanding of the mechanisms underlying many

neurodegenerative diseases. Loss of Dicer2 is associated with neurodegeneration, implicating a key

role for microRNAs in neuroprotection. In order to identify microRNAs that function in

neuroprotection, we carried out an in vivo Drosophila screen of microRNA knockout animals. We

identified a microRNA, mir-263a, that plays a glioprotective role by regulating glutamate receptors

in astrocytic and ensheathing glia. In the absence of mir-263a, levels of the glutamate receptors

Nmdar1, Nmdar2 and Grik are elevated, causing an increase in glial death, and a concomitant

increase in movement deficits. At the end of my talk, I will also briefly outline the challenges for

visualising realtime changes in levels of microRNAs, and describe our efforts to develop a direct,

genetically encodeable Spinach-based microRNA sensor, Pandan.

BIOSKETCH

Sherry Aw obtained her BS in Biochemistry from the University of Wisconsin-Madison and

completed her doctoral studies at Harvard Medical School. Her work focuses on understanding the

mechanisms underlying neurodegenerative diseases, including elucidating the roles of microRNAs.

By carrying out a screen for microRNA mutants that exhibit defective motor function in the aging fly,

she identified novel glioprotective and neuroprotective microRNAs. In addition, she led the

development of a Spinach RNA-based microRNA sensor, Pandan, and a state-of-the-art optical fly

tracking system. She is a co-inventor on two patents, and was awarded the L'Oréal-UNESCO

Singapore For Women in Science National Fellowship in Life Sciences 2017.

SRBA ORAL PRESENTATION

CHARACTERISATION OF IMMUNOMODULATING RNA AS A RIG-I-LIKE RECEPTOR AGONISTS

FOR DENGUE VIRUS THERAPY

Victor Ho1,3, Yong Hui Yee 2, Luo Dahai 2 and Katja Fink 1,3

1 Singapore Immunology Network, A*STAR 2 LKC School of Medicine, Nanyang Technological University 3School of Biological Science, Nanyang Technological University

Dengue is a growing problem globally owing to failure in preventing the spread of the virus. Dengue

virus (DENV) replication can be blocked through activation of innate immune responses using RIG‐I‐

like receptor agonists which are double-stranded RNA containing a triphosphate group on the 5’ end.

Using the smallest dsRNA ligand that can activate RIG‐I signaling (3p10L), we demonstrated that this

molecule is capable of priming human epithelial A549 cells and human monocytic U937 cells into an

anti‐viral state through type I interferon signaling activation. Modifications to dsRNA sequences

resulted in differences in activation of type I interferon signaling, leading to the identification of

3p10LG9, a dsRNA ligand more potent than 3p10L in activating RIG-I dependent type I interferon

response in cell lines as well as inducing these cells into an anti‐viral state. Both 3p10LG9 and 3p10L

have also been shown to potently inhibit DENV replication in primary human dendritic cells isolated

from human skin samples (skin DCs) obtained from healthy donors. When injected with a cationic

polymer, both dsRNA ligands were observed to induce type I interferon production in mice. Overall,

results suggest that 3p10L and 3p10LG9 can activate antiviral responses and confer short-term

protection against DENV. Current work involves investigating the potential of 3p10LG9 to be used

as a vaccine adjuvant and its ability to enhance long-term protection through the adaptive immune

response.

Leah VARDY Senior Principal Investigator

Institute of Medical Biology

A*Star, Singapore

POLYAMINES: CRITICAL REGULATORS OF EPIDERMAL FUNCTION

Understanding gene expression control is essential to understanding cellular behavior in normal and

diseased states. The polyamines putrescine, spermidine and spermine, have been shown to play a

regulatory role in controlling gene expression. The polyamines are ubiquitously expressed and

interact predominantly with RNA and to a lesser extent with DNA and protein in the cell. Changes in

polyamine levels and ratios can regulate cellular function by modulating transcription, RNA

processing, translation, RNA stability and protein function. We are exploring the role of controlled

changes in polyamine levels and ratios in the control of cellular behavior in the epidermis. While it

is clear polyamines play an essential role in the skin, their precise function is not well understood.

We are addressing the role of polyamines in wound healing and epidermal barrier formation. We

show that polyamine regulator AMD1 is translationally upregulated on epidermal differentiation.

AMD1 is rate limiting for the conversion of putrescine to spermidine and spermine and its

upregulation drives an increase in spermine at the expense of putrescine in differentiating

keratinocytes. We further show that the upregulation of AMD1 is essential for epidermal

differentiation and barrier formation. We propose that the translational upregulation of AMD1

drives a shift in polyamine levels required for epidermal differentiation. We have performed genome

wide studies to identify the gene expression changes that are downstream of the polyamine shift.

We are currently examining how these polyamine ratio changes influence RNA behavior in

keratinocytes to drive differentiation.

BIOSKETCH

Leah Vardy became a Senior Principal Investigator at the Skin Research Institute of Singapore in April

2018. Prior to this, she spent 10 years with the Institute of Medical Biology in Singapore. Leah

completed her Ph.D at the ICRF in London and her postdoctoral training at the Whitehead Institute

in Cambridge, MA. Her lab is interested in understanding the role of the polyamines in the control

of RNA in the epidermis.

KEYNOTE TALK 2

Paul AGRIS Professor

Department of Medicine

Duke University School of Medicine, USA

SMALL MOLECULE ANTIBACTERIAL AGENTS TARGETING A NOVEL RNA FOILING THE

EMERGENCE OF RESISTANCE

The emergence of multidrug-resistant, pathogenic bacterial infections requires identification of

unique targets and novel antibacterial agents that elude drug resistance. The redundant usage in

Gram-positive bacteria of a well-conserved tRNA-responsive transcriptional regulatory element, the

T-box, makes it an attractive drug target with a low likelihood of developing resistance. Unique to

Gram-positive bacteria and found in the 5’-untranslated region of mRNAs, T-boxes regulate multiple

operons containing the essential aminoacyl-tRNA synthetase and amino acid biosynthesis genes

within the same cell. In silico docking yielded 200 small molecules as potential binders to the

‘Specifier Loop’ of the Bacillus subtilis tyrS T-box. A family of chemically related compounds (PKZ18)

bound the Specifier Loop in vitro (Kd ~24 μM). PKZ18 inhibited growth of Gram-positive bacteria

including clinical isolates of methicillin-resistant Staphylococcus aureus at minimum inhibitory

concentrations (15-64 μg/mL) consistent with common topical antibiotics. Biofilms were inhibited

using one third (150 μg/mL) the concentration of vancomycin. Liquid and solid media assays

indicated that resistance develops at an extremely low mutational frequency (1.21 × 10-10). In

culture, PKZ18 inhibited transcription of the Bacillus subtilis glycyl-tRNA synthetase mRNA and

translation of the S. aureus threonyl-tRNA synthetase protein. Compound transport in a mammalian

membrane model system was very slow, supporting PKZ18’s minimal cytotoxicity in mammalian cell

culture and its lack of toxicity when applied topically to a wound healing model. Core chemistry of

PKZ18 has been identified as being necessary for antibacterial activity. These findings accelerate a

new paradigm for antibiotic drug discovery that impedes the emergence of resistance.

BIOSKETCH

In 2009, the University at Albany recruited Paul F. Agris from North Carolina State University to pilot

the creation of The RNA Institute, an Albany, NY-based large modern laboratory and global alliance

of top genetic scientists and biomedical investigators. Innovator in RNA biology and chemistry, RNA

modification science and applications and expert in nucleic acid design with modified nucleosides,

Agris was a professor of biochemistry and Head of NCSU’s Department of Molecular and Structural

Biochemistry. He founded and led the RNA Society of North Carolina for more than a decade.

Previously, he was an Assistant, Associate and full Professor in the Division of Biological Sciences and

Department of Medicine at the University of Missouri-Columbia. Agris’ own scientific investigations

into the discovery of RNA-based therapeutics to treat many of the most hard-to-contain RNA-based

diseases — including drug-resistant HIV and MRSA — have been supported continuously since 1974.

He has authored some 170 peer reviewed articles in RNA and RNA modification, a number of reviews,

chapters and edited three volumes. He received a Ph.D. in Biochemistry at MIT under Gene Brown,

and conducted postdoctoral research at the Department of Molecular Biophysics and Biochemistry

at Yale University. Dr. Agris’ present position is in the Department of Medicine, Duke University

Medical School.

OUR SPONSORS

Scientific Organizers

Dahai LUO Nanyang Assistant Professor, Lee Kong Chian School of Medicine, NTU

Institute of Structural Biology, NTU Yvonne TAY

Principal Investigator, Cancer Science Institute of Singapore, NUS President’s Assistant Professor, Department of Biochemistry, NUS

Yue WAN

Senior Research Scientist, Genome Institute of Singapore, A*STAR Adjunct Assistant Professor, Department of Biochemistry, NUS

Gene YEO Professor of Cellular and Molecular Medicine, University of California San Diego

Visiting Professor, Department of Physiology, NUS