高级病生第7次课NoncodingRNAanddisease.ppt

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1、Noncoding RNA and Disease,Zhu-Zhong Mei,PhDDepartment of Pathophysiology,Central dogma of biology,protein,DNA,RNA,RNA and the genome-the end of the central dogma,25,000 protein coding genes in humansProtein-coding DNA=12%of genome,The number of protein coding genes cannot justify the evolutionary ga

2、p between eukaryotes,Gene number,Transcript number,25k,15k,20k,6.5k,31k,30k,30k,Increase the complexity by increasing the combinatorial use of exons by alternative splicing,primarytranscript,Alternativesplicing,Function A,Isoform B,Isoform A,Function B,49k?,20k,23k,6.5k,30k,Gene number,Transcript nu

3、mber,Still the number of different transcripts cannot justify the evolutionary gap between eukaryotes,49k,The Old Guard,Holds steadfast to the idea that most of the transcription(like most of the DNA)is“Junk”Key reasoning:Little sequence conservation of non-coding RNA from species to speciesExplanat

4、ion:Transcription of leftover retroviruses and RNA Polymerase infidelity,The Old Guard(holding onto a lack of science),Microarray data of mouse embryonic stem cells identified at least 945 long non-coding RNAs that are distinctly regulated with development and differentiation.(Dinger,M.E.et al.Genom

5、e Res.2008,18:14331445)Further analysis finds ncRNA to be tissue specific.(Furuno et al.2006;Kapranov et al.2002;Nakaya et al.2007;Ravasi et al.2006)And,its transcription responds to environmental stimuli.(Cawley et al.2004;Louro et al.2007;Ravasi et al.2006).,Continuing,Many have upstream transcrip

6、tional binding sites for proteins such as Sp1,c-Myc,p53,and Creb.(Cawley et al.2004;Euskirchen et al.2004)Long ncRNA with a known function,such as Xist,have very little sequence similarity.The same miRNA regulates different genes in different species(share little sequence homology/rapid evolution th

7、eory).these go on and on.,Eukaryotic RNA transcripts,tRNArRNAsnRNAtmRNARnase P RNAvRNAsgRNAsMRP RNASRP RNAsTelomerase RNA,Transcription/chromatin structure regulatorsPost-transcriptional regulatorsProtein function modulatorsRNA/protein localization regulators,RNA Transcripts,miRNAsiRNApiRNAlncRNA,Pr

8、otein-coding mRNA,Non-coding RNA Transcripts,snoRNAs,Housekeeping RNAs,Regulatory RNAs,micro(mi)RNA,Small RNAs(22 nucleotides in mature form).Encoded within regions of non protein-coding DNA or introns of traditional genes.Seed sequence:nucleotide positions 28 from the 5 end of miRNAs,which generall

9、y perfectly basepair with target mRNA.Mediate Post-transcriptional regulation of messenger RNAs.Complex set of proteins required for formation and processing of miRNA.,General knowledge of miRNAs,Found in wide range of plants and animal species.1-5%of predicted genes humans(1000 total).Individual mi

10、RNAs are present at a level of 1,000 molecules/cell with some reaching 50,000.Estimated that 1/3 of protein coding genes are regulated by miRNAs.Cells shorten 3 UTR to dampen miRNA regulation of important genes at certain times.,The discovery of miRNAs,miRNA was first discovered in 1993 by Victor Am

11、bros and Gary Ruvkun(published at the same issue of Cell).The second miRNA Let-7 was discovered in 2000 by Frank Slack as a postdoc at Harvard(Ruvkun lab).,Victor Ambros,Gary Ruvkun,Most genes identified from a mutagenesis screens are protein-coding,but lin-4 encodes a 22-nucleotide non-coding RNA t

12、hat is partially complementary to 7 conserved sites located in the 3-untranslated region(UTR)of the lin-14 gene.,The first discoered miRNA:lin-4,He L et al.,Nature Review in Genetics,2004,5:522-531,lin-4 vs lin-14:the first miR-target couple,Thought to be an oddity not a general phenomenon.,He L et

13、al.,Nature Review in Genetics,2004,5:522-531,Processing and maturation of miRNA,Principles of miRNA-mRNA interaction,Why all the fun for down-regulation?miRNA mediated TNF-alpha upregulation!,Vasudevan S.et al.Science 2007,318:1931-1934,Since each miRNA may regulate as many as 200 different gene tar

14、gets,overexpression or loss of function of miRNAs may have widespread oncogenic effects because many genes will be dysregulated.Overexpression of some miRNAs can suppress the expression of tumor-suppressor gene targets,whereas loss of function of other miRNAs may allow overexpression of the oncogene

15、s they regulate.Approximately 10%of miRNAs have been found to be either greatly overexpressed or down-regulated in various tumors,and are referred to as oncomirs.,miRNA and disease,Garzon R et al.,Nature Reviews in Drug Discovery,2010,9:775-789,miRNAs as oncogenes and tumor suppressors,Dysregulated

16、expression of miRNAs in tumorigenesis,Cho WC et al.Int J Biochem Cell Biol 2010;Cho WC et al.Mol Cancer.2007;6:60.,Dysregulated expression of miRNAs in tumorigenesis,Cho WC et al.Int J Biochem Cell Biol 2010;Cho WC et al.Mol Cancer.2007;6:60.,miRNA expression pattern changes during oncogenesis,and i

17、s unique for each cancer,miR-34 family as mediator of tumor suppression by p53,Hermeking H et al.,Cancer Cell,2007,12:414-418,Ma L et al.,Nature cell biology,2010,247,miR-9,a MYC/MYCN activated microRNA,regulates E-cadherin and cancer metastasis,Cellular mRNA targets of viral miRNAs,Cullen BR,Nature

18、,2009,457:421-425,Some viruses encode miRNAs,Nobel Prize in Physiology or Medicine 2006,Andrew Z.FireCraig C.Mello,C.elegans,miRNA,Exogenously delivered21-23mer dsRNAActs through RISCInduces homologous target cleavagePerfect sequence matchResults in target degradation,Endogenously produced21-23mer d

19、sRNAActs through RISCInduces homologous target cleavageImperfect sequence matchResults in target degradation or translation arrest,siRNA,A comparison between miRNA and siRNA,A comparison between miRNA and siRNA,He L,Nature Review in Genetics,2004,5:522-531,lncRNA:long non-coding RNA,Transcripts long

20、er than 200 nucleotides that have little or no protein-coding capacity.Long ncRNAs can regulate gene expression through a diversity of mechanisms.,Genomic organiztion of coding and non-coding transcripts,Mercer TR,Nature Reviews in Genetics,2009,10:155-159,Functions of lnRNAs,Mercer TR,Nature Review

21、s in Genetics,2009,10:155-159,ncRNAs mediated X chromosome inactivation,lnRNA HOTAIR reprograms chromatin state to promote cancer metastasis,Gupta RA,Nature,2010,464:1071-1076,Rate limitingstep,amyloid precursor protein,BACE1-AS antisense RNA and Alzheimers Disease,Chr.11,(104 nts complete match),BA

22、CE1-AS upregulates the expression of BACE1,Faghihi MA,Nature Medicine,2008:14,723-730,Dysregulation of BACE1 and A production in Alzheimers disease,St George-Hyslop P,Nature Medicine,2008:14,711-712,Posttranscriptional regulation of PTEN dosage by noncoding RNA,He L,Science Signaling,2010,3:pe39,p53

23、 mRNA controls p53 activity by managing Mdm2 functions,Candeinas MM et al.Nature cell biology,2008,10:1098,Clinical application of non-coding RNAs,Alnylam Pharmaceuticals recently passed phase II human clinical trials showing statistically significant treatment of human RSV with siRNAMany other earl

24、y clinical trials beginningMany siRNAs have worked in animal models such as recently,at Harvards Immune Disease Institute,Ab and polyarginine conjugated siRNAs blocked HIV replication in humanized mice.,Strategies for miRNA based therapies,Garzon R,Nature Reviews in Drug Discovery,2010,9:775-789,The

25、rapeutic silencing of miR-10b inhibits metastasis in a mouse mammary tumor model,Ma L,Nature Biotechnology,2010,28:341-347,Future Directions RNA therapy,SMALL(small enough to use as a therapeutic?)Would function as an entirely new class of therapyGoal:To selectively inhibit viral or oncogenic protei

26、n transcription/translationGoal 1a:antisense small RNA to soak up detrimental RNA(such as miR-155 in lymphomas)Goal 1b:Could be used to regulate metabolic or inflammatory disorders with more understanding,Summary RNAs can,Down-regulate transcription/translationUp-regulate transcription/translationProtect from viruses/transposonsServe in epigenetic inheritance(via histone patterns)Signaling Molecule?,Perhaps the rabbit hole is deeper than what was originally imagined?,RNA world,Most of the RNA transcribed from your genome doesnt make protein.,Thank you,Email:mzhuongQQ.com,