TERRA, telomeric repeat-containing RNA, RNA, LncRNA, Long noncoding RNA, Cancer, noncoding RNA, microRNA, transcription, RNA polymerase

1. lncRNA
Long non-protein-coding RNAs
functional surprises from
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2. LONG NON-PROTEIN-CODING RNAS
(LNCRNAS)
 largest transcript class in the mouse and human
transcriptomes
 large amount of noncoding RNAs :
• microRNAs
• long noncoding RNAs(lncRNAs)
regulation of cellular processes :
cell growth and apoptosis, as well as cancer
progression and metastasis
CRITICAL
ROLE
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3. LONG NON-PROTEIN-CODING RNAS
(LNCRNAS)
 biggest class of ncRNAs
 approximately 10,000 lncRNA genes annotated in
humans.
 The FANTOM3 project, by using physical cDNA
clone analysis alone, identified more than 23,000
lncRNAs
 Transcribed:
• RNA polymerase II
• RNA polymerase III
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5. FUNCTION
 The function of the majority of lncRNAs is still unknown.
 negatively or positively regulating gene expression
 differentiation
 human disease
 regulate protein-coding (pc) gene expression:
 posttranscriptional
 transcriptional
 Paucity of Introns (nuclear localization)
 Low GC content (low expression level)
 Predicted ORFs have poor start codon and contexts
 Significant similarity between lncRNA and 3’-UTR of
mRNA (structural feature + sequence composition)
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6. LOCATION
mRNAs  cytoplasm
lncRNAs  nucleus
 cytoplasm
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7. HUMAN DISEASES:
 Cancers (breast cancer, colorectal cancer, prostate
cancer, hepatocellular carcinoma, leukemia, melanoma
 Alzheimer
 Psoriasis
 Heart disease
 Transient neonatal diabetes mellitus (lncRNA HYMAI)
 Pseudohypoparathyroidism (lncRNA NESP-AS)
 Atheromatosis and atherosclerosis
 Beckwith–Wiedeman syndrome
(lncRNAs H19 and KCNQ1OT1)
 Silver–Russell syndrome (lncRNA H19(
 McCune–Albright syndrome (lncRNA NESP-AS)
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8. LOCATION IN GENOME
 LncRNAs can be categorized according to their
proximity to protein coding genes in the genome,
using this criteria lncRNAs are generally placed into
five categories:
 sense
 antisense
 bidirectional
 intronic
 intergenic
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9. Sense - The lncRNA sequence overlaps
with the sense strand of a protein coding
gene.
Antisense - The lncRNA sequence
overlaps with the antisense strand of a
protein coding gene.
Bidirectional - The lncRNA sequence is
located on the opposite strand from a
protein coding gene whose transcription is
initiated less than 1000 base pairs away.
Intronic - The lncRNA sequence is derived
entirely from within an intron of another
transcript. This may be either a true
independent transcript or a product of pre-
mRNA processing
Intergenic - The lncRNA sequence is not
located near any other protein coding loci
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12. HUMAN CANCER-ASSOCIATED LNCRNAS
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13. TERRA
telomeric repeat-containing RNA
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14. TELOMERES
 Identified in the 1930s
 nucleoprotein structures that protect the ends of
 Chromosomes
 From DNA double-strand breaks (DSBs) and from
degradation
 At the cellular level for genome stability
 At the organismal level, they act as tumour
suppressors and may contribute to ageing
 Human telomeres extend from 9–15 Kb,
 but can be as long as 100 Kb in rodents
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15.  Human and mouse telomeres are composed:
 long tracts of double-stranded G rich TTAGGG
repeats
 complex, shelterin
 The shelterin complex consists of six protein
subunits:
1. Telomeric repeat binding factor-1 (TRF1)
2. TRF2,
3. Repressor activator protein-1 (RAP1)
4. TRF1-interacting protein-2 (TIN2)
5. TINT1/PIP1/PTOP 1 (TPP1)
6. protection of telomeres-1 (POT1)
 regulated the maintenance of telomere length and
protects natural chromosome ends from being
recognized as damaged 15

16.  Single-stranded repeat structures created by 5′ to 3′
exonucleases
 The invasion of the 3′ telomeric overhang into the
duplex telomeric array forms a T-loop TRF2.
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17. THE FOLLOWING ROLES OF
TELOMERES ARE WELL ESTABLISHED
 Regulated the lifespan of cells (Harley et al, 1990;
Allsopp et al, 1992; Levy et al, 1992; Bodnar et al, 1998)
• Telomere shortening occurs at the distal ends
• Telomerase is regulated at individual chromosome ends
• in humans, telomerase is expressed in most tissues only
during the first weeks of embryogenesis (Ulaner and
Giudice, 1997).
• Repression of telomerase in somatic cells is thought to
result in a powerful tumour-suppressive function.
 Telomeres protect natural chromosome ends from
unwanted DNA repair activities
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18. TELOMERIC REPEAT-CONTAINING RNA
(REFERRED TO AS TERRA)
 A noncoding RNA molecule, has recently been found in
mammalian cells
 Forms an integral component of telomeric heterochromatin
 TERRA transcription occurs at most or all chromosome ends
 TERRA regulated by RNA surveillance factors and in
response to changes in telomere length
 Roles in the regulation of telomerase and in orchestrating
chromatin remodelling throughout development and cellular
differentiation
 transcribed by RNA polymerase II (RNAPII). RNAPI and
RNAPIII)
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19.  inhibition of RNAPII by a-amanitin reduces the
abundance of TERRA in both human and mouse
cells
 Actinomycin D treatment resulted in a
50%reduction in TERRA levels within 2.5–3 h
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20.  Approximately 7% of human TERRA molecules are
polyadenylated
 most or all yeast TERRA molecules carry a poly(A)
tail
 all human and most yeast TERRA 5′ ends contain a
7 -methylguanosine (m7G) cap structure
 increase the stabilityof TERRA molecules
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22.  TERRA forms stable G-quadruplexes in vitro and in
vivo
 there is direct evidence that TERRA binds to the
telomeric protein TRF2 by forming an
intramolecular G-quadruplex structure
 TERRA participates in multiple regulatory functions:
 telomerase activity
 Heterochromatinization of telomeres
 cellular differentiation
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23. REGULATION OF TELOMERE LENGTH VIA SEVERAL
PATHWAYS
 TERRA can inhibit telomerase activity to promote
telomere shortening.
 TERRA can promote Exo1-dependent resection at
chromosome ends to initiate telomere shortening.
 the loss of TRF1 increases the recruitment of
telomerase
 Rap1 binds to the telomeres via TRF2 and plays a
negative regulatory role on telomerase recruitment.
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24. HOW DOES TERRA REGULATE TELOMERE
LENGTH DURING THIS PROCESS?
 The shelterin components TRF1 and TRF2 directly
associate with TERRA in multiple cell types in vivo
 It has also been reported that the T-loops can be
deleted by homologous recombination (HR), which
results in the rapid shortening of telomeres.
 TRF2 is proposed to suppress T-loop HR
 The amino-terminal define (GAR) domain of TRF2
has a high affinity for RNA
 Binding of TERRA to TRF2 could alleviate
protection of the t-loop and induce t-loop HDR
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26. TERRA AT TELOMERES
 TERRA is exclusively found in nuclear RNA fraction
 TERRA can be detected using RNA fluorescence in
situ hybridization (RNAFISH) at a subset of
telomeres in interphase cells
 present on human and mouse chromosome ends in
the metaphase of the cell cyclens from human and
mouse cells
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27. TERRA EXPRESSION LEVELS DO NOT CORRELATE WITH
TELOMERE LENGTH AND RADIATION SENSITIVITY IN HUMAN
CANCER CELL LINES
 In human cells, CpG-island promoters drive TERRA
transcription and are regulated by methylation
 suggestion:the amount of TERRA may be related to
telomere length.
 five human cell lines:
 HeLa (cervical carcinoma)
 BRC-230 (breast cancer)
 AKG (gastric cancers)
 GK2 (gastric cancers)
 GM847 (SV40 immortalized skin fibroblasts)
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28.  in cancer cells telomere length is maintained,
usually through telomerase activation
 in about 15% of human cancers, an alternative
telomere lengthening (ALT) mechanism operates
which does not require telomerase activation.
 In ALT cells, telomere maintenance is due to
a recombination based mechanism causing
great intracellular variability of telomere length
 TERRA is regulated during the cell cycle
 being lowest in late S phase
 peaking in early G1
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TERRA
telomerase
3’
3’

30.  regulation of telomerase by TERRA and hnRNPA1
 multifunctional RNA-binding protein
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37.  The Reference Database For Functional Long Noncoding
RNAs
http://www.lncrnadb.org/
 the LncRNADisease database
http://www.cuilab.cn/lncrnadisease
 the latest version of this long non-coding RNA database
contains 113,513 human annotated lncRNAs
http://www.lncipedia.org/
http://deepbase.sysu.edu.cn/chipbase/lncrna.php
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38. THE END
Thanks
Sogand Vahidi
Email: So.vahidii@gmail.com
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