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Research progress of piRNA
evolution
Xuedong Pan
2012-3-28
What is piRNA
• Small RNAs: piRNA, siRNA, miRNA.
• piRNA: derive from repetitive genomic
element, interact with PIWI
AGO
family
protein
Guide RNA
RISC(RNA
induced
silencing
complex)
Germline-specific AGO
PIWI
family( PIWI,
AUB, AGO3)
piRNA
piRISC:
recognize and
silence
complementary
RNA
Biogenesis of small RNAs
Unkown;
important
Dicer
independent
Proved; 5’U
and 10 A
piRNA biogenesis
Long precursor can generate a lot
of piRNAs, thus is piRNA cluster
Not only piwi. Piwi interact with
Tudor-domain proteins directly.
Tudor-domain proteins methylate
arginine in piwi, which differs
piwi with other AGOs. Piwi also
needs Armi.
Putative roles for piRNA during
silencing of protein-coding genes
In drosophila testis, if
Su on ChrY do not
work, Ste on ChrX is
deprepressed, which
leads to infertility.
Piwi expression
driven by TJ. piR
from 3’UTR of tj
together with piwi
suppress Fas3.
In drosophila testis,
AT-chrX code piR, piR
interact with AUB,
and little AGO3, to
suppress vasa
Failure of nos mRNA
deadenylation and
translational
repression by piR
piRNA at different developmental
stages in mouse
Conclusion: different
developmental stage,
different protein and piR
piR enriched in
TE. Binds MILI
and MIWI2
Pachytene: cross over
happens
piR not enriched
in TE. Binds MIWI
and MILI
Piwi family
in mouse:
MIWI2,
MIWI, MILI
TDR: tudor domain
containing proteins.
Bind piwi directly.
Current Evolutionary studies
demonstrate
• piRNA strongly repress retrotransposons, coevole?
• piRNA evolves very fast among species
• piRNA loci locates in low recombination region
• piRNA show a signature of selective constraint in
African populations
Molecular evolution: piRNA evolves
super fast and do not lost
• Rapid repetitive element-mediated expansion of
piRNA clusters in mammalian evolution (09 PNAS,
Assis et. al, U Michigan)
1) 43% of all rodent piR clusters arose after
rodent-primate divergence. While the highest
known expansion rate for olfactory receptors is
33%.
2) Olfactory receptors and miRNAs are lost at the
same rate with which they are acquired. However,
not a single cluster loss was observed for piRNA.
Positive selection most likely
• RE(repetitive element) usually increases
deletions more than insertions.
• 60 cluster acquisitions without a single loss
• Long insertions are unlikely to be neutual
• piRNAs are involved in transposon silencing.
• So, an arms race between expanding families
of mammalian TE and piRNA cluster?
Proteincoding
gene
piRNA evolve by ectopic
recombination
ProteinRepetitive
element
piRNA
cluster
coding
gene
(A) Architecture of a typical cluster-harboring genomic region. Intergenic region between
two protein-coding genes piRNA cluster. The inserted segment is depicted in brackets.
(B) The inserted segment is scanned against the genome to locate the source paralog.
(C) Similarity between cluster- and source paralog-harboring regions includes preceding
REs
(D) Double-stranded break, leads to extension and reannealing of the broken strand
Population dynamics of piRNA and
TE(transposable element) in Drosophila
• Jian Lu and Andrew G. Clark, PNAS 2010
• Activities of a large number of retrotransposons
are severely silenced by piRNAs.
1) quantified expression levels of 32 TE families
2) in ovaries of one wild-type and three piRNA
mutants
• Other reports prove piRNA silence transposons.
piRNA repress
TEs in RNA level
Life cycles of
transposable
elements(TE)
a:
IR/DR: inverted/direct repeats
bind by transposases
b:
Reverse transcription in
cytoplasm, integration in
nucleus by integrases
LTR: long terminal repeat
GAG protein: form virus-like
particles
c:
ORF1 and ORF2 have reverse
transcriptase domain
mRNAs are integrated into
genome by target-primed
reverse transcription
Forward simulation of piRTs and
targetRTs
• Focus on retrotransposon only
• Retrotransposons including piRTs: located
inside piRNA loci; targetRTs: remaining.
• Model: largely come from Dolgin and
Charlesworth, Genetics, 2008
Start prameters
For 15000
generations
Selection: fitness is exponential quadratic, decreasing function of
TE copy number.
recombination: uniform distribution of crossover positions
transposition and excision(Poisson process throughout genome)
parameters
• Ne = 10e6, constant-sized
• One chromosome, 40Mb(close to chr2,3)
• One chr one crossover per generation, r =
2.5e-8
 an bn2 / 2
• Fitness of chr: w  e
; n: the
number of retrotransposons; a = 10e-5;
b = 5e-6
• Excision rate is v. v = 0
• For targetRTs, retrotransposition rate is u1 if
piRNA is not expressed in the cell, u2 if piRNA
is expressed.
Model piRNA’s suppression effect
• For targetRTs, retrotransposition rate is u1 if
piRNA is not expressed in the cell, u2 if piRNA
is expressed.
• Four scenarios:
1) piRNAs have no repression effect: u1 = u2
2) 3) 4) piRNAs can reduce retrotransposition
rates to 10%, 1% and 0.1%(u2 = 0.1u1, 0.01u1,
0.001u1)
Model’s other assumptions
• Retrotransposons inserted into piRNAgenerating regions will be suppressed.
• No sequence divergence between paralogous
copies of retrotransposons, so that one piRNA
can potentially repress all retrotransposons.
• Retrotransposons located inside piRNA loci
lose the ability to retrotranspose
• Ectopic recombination is not allowed
Forward simulation for 15000
generations: active piR repress
retrotransposons and increase fitness
A) Number of retrotransposons
B) Fitness costs to the host
* scenario1,2,3,4: piRNA’s repressing capabilities increases
Forward simulation for 15000
generations: if piR is active, piRTs
increases with time
Scenario
IV
Scenario
I
Proportion(%) of all retrotransposons that are piRTs
Scaled parameters: Ne = 500, a = 0.001, b= 0.0005, r = 2.5e-8, u1 = 0.01, v = 0
Frequency spectra
of piRT insertions:
When piRNA takes
effect, piRTs have
a higher
probability to be
fixed
Frequency spectra
of targetRT
insertions
Also has a higher
probability to be
fixed, because their
deleterious effects
are alleviated by
piRNAs
Frequency spectra of piRT and targetRT
from published genomic data:
recombination matters
A vs. B: TE
longer than
500bp
No
recombination
occured
C vs. D: subset
of A vs. B,
recombination
occured
Combination
of 2 datasets
TE and piR like low recombination
regions
• piR loci enriched in ericentromeric or
telomeric heterochromatin
• TEs significantly enriched in low
recombination regions, even in euchromatin
• When recombination occurs, piRTs are
deleterious because they can mediate ectopic
recombination
** piRNA also enriches in centromeric and
telomeric region. Telomeric lacZ insertion
produces abundant piRNAs. Telomere is a
picluster?
Reducing recombination rate of piRNA
loci greatly reduces retrotransposons
Recombination rate = 2.5e-8
Recombination rate = 0
Human piRNAs are under selection in
Africans and Repress TEs
• Sergio Lukic and Kevin Chen, MBE, 2011
• Material and methods:
human piR sequences(Girard et al. 2006)
mouse piR sequences(Lau et al. 2006)
Hapmap phase3 data
methods: derived allele frequency spectrum;
BWA tool, mapping reads
piR evolved rapidly between human
and chimp
• piR region vs. piR flanking region
piR flanking region (1000bp each side of piR);
nucleotide substitution between human and
chimp is not significantly different between
piR region and piR flanking region
Selective constraint in African
populations
ASW, YRI vs. CHB, CEU
Selective constraint in Africans is
consistent with a much higher rate of
transposon insertions in African
compared with non-African
populations(Ewing A, Kazazian H, Genome
Res, 2010)
Interspecies: no constraint
intraspecies: constraint in Africans;
mild constraint in non-African
• Explanation1: strength of selective constraint
my simply differ between these two time
scales.
• Explanation2: interspecies substitution rate,
but not the derived allele frequency
distribution, is affected by mutation rate
biases.
younger TE, more piR targets
Younger TEs are more active, so more
piRNAs which target them remain in
current human genome; the pattern is
the same for mouse
The pattern is the same
in our data
ORF2 of LINE1 is depleted of piRNA
matches
L1-ORF2
functions as
reverse
transcriptase
Red line: number of G/C-nucleotides per base on L1 mRNA
Blue line: density of sense piRNA matches to L1
Green line: density of antisense piRNA matches to L1
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