rasiRNAs: a class of small regulatory RNAs

Jasmine Lau

4 May 2009

Small noncoding regulatory RNAs

Longer length 24-27 nucleotides

small interfering RNAs

(siRNAs)

microRNAs (miRNAs)

Shorter length 20-25 nucleotides

piwi-interacting RNAs (piRNAs)

Other piRNAs

rasiRNAs

The Family of small RNAs

rasiRNAs= Repeat-associated small interfering RNAs

sequence-specific gene silencing (RNAi mechanism)

forms RNA-protein complexes with Piwi subfamily proteins

sequences correspond to genomic repeats and transposons

First hints of rasiRNAsTrypanosoma brucei • parasitic protist species• causes African trypanosomiasis (sleeping sickness)

2001

2002

2003

2004

2005

2006

2007

2008

Present

Abundant 24-26 nucleotide long siRNA-like RNAs Represented 2 retrotransposons in the genome Both sense and antisense polarity

Regulatory mechanism to silence retrotransposon transcription and mobilization?

http://www.sanger.ac.uk/Projects/T_brucei/Tryp.cartoon.shtml

First hints of rasiRNAsTrypanosoma brucei • parasitic protist species• causes African trypanosomiasis (sleeping sickness) 2002

2001

2003

2004

2005

2006

2007

2008

Present

Abundant 24-26 nucleotide long siRNA-like RNAs Represented 2 retrotransposons in the genome Both sense and antisense polarity

Regulatory mechanism to silence retrotransposon transcription and mobilization?

Nicotiana and Arabidopsis thaliana plants

24-26 nucleotide long endogenous siRNA species Represented 2 retrotransposons in Nicotiana genome, 1 retrotransposon in Arabidopsis genome Both sense and antisense polarity

Linked to DNA methylation and systemic silencing Mechanism to keep retrotransposon silent?

rasiRNAs: new class of RNAs

2003

2001

2002

2004

2005

2006

2007

2008

Present

Drosophila melanogaster small RNA profiling

2 distinct classes of small RNAs:(i) 21-23 nt RNAs(ii) 24-26 nt RNAs

Sequences correspond to genomic repeats/transposons Class of RNAs called repeat-associated siRNAs (rasiRNAs)

Sequences correspond to genomic repeats or transposons

Also described in zebrafish, fission yeast

Sense & antisense orientation of repetitive sequence elements:• Long Terminal Repeat (LTR) and non-LTR retrotransposons• DNA transposons• Satellite and microsatellite sequences• Complex repeats

rasiRNAs: subclass of piRNAs

2006

2001

2002

2004

2005

2003

2007

2008

Present

Longer length 24-27 nucleotides

piwi-interacting RNAs (piRNAs)

Other piRNAs

rasiRNAs

24-26 nucleotide long RNAsBUT lack transposon or repetitive seqeuences

rasiRNA biogenesisArgonaute

protein family

Piwi subfamily

Ago1

Ago subfamily

Ago2 Ago3Aubergine

(Aub)Piwi

Associate with rasiRNAs, present in germline cells and early embryos

Associate with miRNAs

Associate with siRNAs

Dicer-dependent production to produce

double-stranded precursor

Dicer-independent production

rasiRNA biogenesisSense transcript:

Antisense transcript:

5’3’

AGO3

5’ 3’A

PIWI/ AUB

3’ 5’U

Adenine at nucleotide 10

Uridine at 5’ terminal end

UA

PIWI/ AUB

5’3’ U

5’ 3’

PIWI/ AUB

5’3’ U

10 complementary nucleotides

5’ 3’A

AGO3

5’ 3’A

AGO3

5’3’ U

? PIWI/ AUB

A5’ 3’

?AGO3

Zucchini/ squash?

Zucchini/ squash?

2’-O-Me

Pimet

Pimet

2’-O-Me

What do rasiRNAs do?

• Silencing and suppressing mobilization of retrotransposons and other selfish genetic transposable elements• Guide transcriptional and post-transcriptional gene silencing of repetitive sequence elements• Guide chromatin silencing to establish and maintain heterochromatin

Maintaining genomic stability

Present in fission yeast, plants, fly, zebrafishFunctions mainly in the germline and early embryo development in fly and zebrafish, found at very low levels in the adult fly.

Also in particular for flies:• Maintaining germ cell self-renewal• Establishing axis polarity of the early embryo• Regulating pole cell formation and oocyte maturation• Maintaining telomeres

How rasiRNAs work

Fission yeast : Gene silencing and establishing heterochromatin

Dicer cleaves long dsRNA

Transcriptional silencing complex in nucleus Ago1, Chp1 and Tas3

Binds DNA/ chromatin, induce DNA and histone methylation

Plants : Similar mechanism to yeast (above), also repress repetitive transposable elements

Ago4-dependent mechanisms

Induce systemic gene silencing, Histone H3 lysine-9 methylation, DNA methylation Direct chromatin modifications

How rasiRNAs work

Drosophila melanogaster:

rasiRNAs associate with piwi Argonaute proteins, and RNA helicases (Spindle-E and Armitage)

Induce histone H3 Lysine 9 methylation and association of HP1 chromatin binding protein with heterochromatin

Downstream effector mechanisms and pathways are not yet known Literature between piRNAs and rasiRNAs are merging

Longer length 24-27 nucleotides

piwi-interacting RNAs (piRNAs)

Other piRNAs

rasiRNAs

Summary: rasiRNAs

rasiRNAs= Repeat-associated small interfering RNAs

sequence-specific gene silencing (RNAi mechanism)

forms RNA-protein complexes with Piwi subfamily proteins

sequences correspond to genomic repeats and transposons