rasi RNAs a class of small regulatory RNAs

rasi. RNAs: a class of small regulatory RNAs Jasmine Lau 4 May 2009

The Family of small RNAs Small noncoding regulatory RNAs Shorter length 20 -25 nucleotides small interfering RNAs (si. RNAs) micro. RNAs (mi. RNAs) rasi. RNAs = Repeat-associated small interfering RNAs Longer length 24 -27 nucleotides piwi-interacting RNAs (pi. RNAs) Other pi. RNAs sequence-specific gene silencing (RNAi mechanism) forms RNA-protein complexes with Piwi subfamily proteins sequences correspond to genomic repeats and transposons rasi. RNAs

First hints of rasi. RNAs 2001 2002 2003 2004 2005 Trypanosoma brucei • parasitic protist species • causes African trypanosomiasis (sleeping sickness) Abundant 24 -26 nucleotide long si. RNA-like RNAs Represented 2 retrotransposons in the genome Both sense and antisense polarity Regulatory mechanism to silence retrotransposon transcription and mobilization? 2006 2007 2008 Present http: //www. sanger. ac. uk/Projects/T_brucei/Tryp. cartoon. shtml

First hints of rasi. RNAs 2001 Trypanosoma brucei 2002 • parasitic protist species • causes African trypanosomiasis (sleeping sickness) 2003 2004 2005 2006 2007 2008 Present Abundant 24 -26 nucleotide long si. RNA-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 si. RNA 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?

rasi. RNAs: new class of RNAs 2001 2002 2003 2004 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 or transposons 2005 Sequences correspond to genomic repeats/transposons Class of RNAs called repeat-associated si. RNAs (rasi. RNAs) 2006 Also described in zebrafish, fission yeast 2007 2008 Present Sense & antisense orientation of repetitive sequence elements: • Long Terminal Repeat (LTR) and non-LTR retrotransposons • DNA transposons • Satellite and microsatellite sequences • Complex repeats

rasi. RNAs: subclass of pi. RNAs 2001 Longer length 24 -27 nucleotides 2002 2003 piwi-interacting RNAs (pi. RNAs) 2004 2005 2006 2007 2008 Present 24 -26 nucleotide long RNAs BUT lack transposon or repetitive seqeuences Other pi. RNAs rasi. RNAs

rasi. RNA biogenesis Argonaute protein family Ago subfamily Ago 1 Associate with mi. RNAs Ago 2 Associate with si. RNAs Dicer-dependent production to produce double-stranded precursor Piwi subfamily Ago 3 Aubergine (Aub) Piwi Associate with rasi. RNAs, present in germline cells and early embryos Dicer-independent production

rasi. RNA biogenesis 5’ PIWI/ AUB Zucchini/ squash? 5’ 3’ A U Antisense transcript: 3’ 10 complementary nucleotides 5’ Uridine at 5’ terminal end 2’-O-Me ? AGO 3 Adenine at nucleotide 10 AGO 3 Sense transcript: AGO 3 3’ Pimet 5’ A 3’ A AGO 3 5’ 3’ A U PIWI/ AUB 3’ 5’ 5’ 3’ A U 2’-O-Me 3’ U PIWI/ AUB 5’ Pimet 3’ U ? Zucchini/ squash? PIWI/ AUB 5’ 3’ 5’

What do rasi. RNAs do? Present in fission yeast, plants, fly, zebrafish Functions mainly in the germline and early embryo development in fly and zebrafish, found at very low levels in the adult fly. Maintaining genomic stability • 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 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 rasi. RNAs work Fission yeast : Gene silencing and establishing heterochromatin Dicer cleaves long ds. RNA Transcriptional silencing complex in nucleus Ago 1, Chp 1 and Tas 3 Binds DNA/ chromatin, induce DNA and histone methylation Plants : Similar mechanism to yeast (above), also repress repetitive transposable elements Ago 4 -dependent mechanisms Induce systemic gene silencing, Histone H 3 lysine-9 methylation, DNA methylation Direct chromatin modifications

How rasi. RNAs work Drosophila melanogaster: rasi. RNAs associate with piwi Argonaute proteins, and RNA helicases (Spindle-E and Armitage) Induce histone H 3 Lysine 9 methylation and association of HP 1 chromatin binding protein with heterochromatin Downstream effector mechanisms and pathways are not yet known Literature between pi. RNAs and rasi. RNAs are merging

Summary: rasi. RNAs = Repeat-associated small interfering RNAs sequence-specific gene silencing (RNAi mechanism) Longer length 24 -27 nucleotides piwi-interacting RNAs (pi. RNAs) forms RNA-protein complexes with Piwi subfamily proteins sequences correspond to genomic repeats and transposons Other pi. RNAs rasi. RNAs