hervs (human endogenous retroviruses) and ltr (long terminal repeat) - like elements are dispersed...
TRANSCRIPT
HERVs (Human endogenous retroviruses) and LTR (long terminal repeat) - like elements are dispersed over 8% of the whole human genome. There are at least 22 independent HERV families within the human genome, which originated from germ-cell infection by the exogenous retrovirus during primate evolution. Elucidation of expression pattern in HERV elements should provide information about fundamental cellular activities and the pathogenesis of multifactorial diseases such as cancer and autoimmune disease. HERV-W env gene is related to multiple sclerosis, and has potential roles for normal differentiation of human villous cytotrophoblast into syncytiotrophoblast. HERV-W env gene was expressed differentially in human tissues. Especially, it was highly expressed in human placenta. This phenomenon indicates HERV-W env gene have the different roles in each tissues. Here, we applied realtime RT-PCR for detection of its expression in various human tissues. We also analysed such amplification using cancer cells and monkey tissues, and discussed in relation to physiological function. ____________________
Retroelement
Retroposon
SINE
Retrotransposon
RetrovirusLINE
RNA intermediate- LTR element + LTR element
- env + env
- RT + RT
gag pol envLTR LTR
Human Alu
ORF1 ORF2P
Poly(A)L1Full-length HERVs/exogenous retrovirus
P
LTR LTR
LTR LTRORF2ORF1Poly(A)
Yeast Ty1/copia/truncated HERVs
Human THE1
Pseudogenes
LTR LTR
provirus
LTR LTR
Enodgenous retrovirus
Exogenous Retroviruses
RetrotransposonsEndogenous Retroviruses
buddingproteins(gag, env)
translation
infection
REVERSE
TRANSCRIPTION
transcription
integrationcDNA
AAA
mRNA
AAA
mRNAtranscription
LTR LTR
re - integration
translation
proteins(gag, env)
REVERSETRANSCRIPTION
cDNA
Particle formation(budding)
LTR LTR
re - integration
Particle formationno
re-infection
LTR LTR
retrotransposonAAAtranscription
mRNA
cDNA
REVERSETRANSCRIPTION
Cellular gene
pseudogene
splicing
AAA
REVERSETRANSCRIPTION
cDNAintegration
transcriptionspliced- mRNA
translation
proteins(gag)
Other region16%
Gene-related Sequence
36%
LINE
20%
SINE13%
Coding sequence3%
Pseudogene1%
HERV element8%
DNA element3%
Primer Design
Twenty Different Human tissue cDNAs
QuantitativeAnalysis
Realtime RT-PCR using SYBR Green I
Extension(II) phase
Extension(I) phase
End of PCR cycle
Annealing phase
SYBR Green I
Detection of fluorescence
Experimental Procedure
relative quantification
normalisation
via onereference
gene
via referencegene index
>3HKG
externalcalibration
curve withoutany reference
gene
without real-time PCRefficiency correction
2(- CP)△△
with real-time PCRefficiency correction
REST, qGeneLC software & etc...
1. Gapdh standard 2. W-env standard
3. Meting graph
5. Tissue melting graph
4. Tissue expression graph
0
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
0.09
0
0.002
0.004
0.006
0.008
0.01
0.012
0.014
0.016
6. Relative quantitative analysis of w-env expression
7. Relative quantitative analysis of w-env expression except for placenta
Yi JM and Kim HM and Kim HS 2004. Expression of the human endogenous retrovirus HERV-W family in various human tissues and cancer cells. J. Gen. Virol. 85: 1203-1210.
Yi JM, Osamu T and Kim HS 2003. Molecular characterization and phylogenetic relationship of HERV-W family in the Macaca fuscata. Arch. Virol. 148(8) 1613-1622.
Plac
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Adre
nal g
lan
d Bone
mar
row Ce
rebe
llum Br
ain Fe
tal b
rai
n Feta
l liv
er Hea
rt Kidn
ey Li
ver Lu
ng Pr
osta
te Sa
livar
y gl
and Sk
elet
al m
uscl
e Spin
al c
ord Te
sti
s Thym
us Th
yroi
d Trac
hea Ute
rus