microRNAmicroRNAmicroRNAmicroRNA
Cass JabaraCass JabaraCass JabaraCass Jabara
Exam QuestionsExam QuestionsExam QuestionsExam Questions
What are microRNAs, and what is What are microRNAs, and what is their primary function?their primary function?
Where are microRNAs commonly Where are microRNAs commonly found in the human genome, and found in the human genome, and how does this correlate to disease?how does this correlate to disease?
What are microRNAs, and what is What are microRNAs, and what is their primary function?their primary function?
Where are microRNAs commonly Where are microRNAs commonly found in the human genome, and found in the human genome, and how does this correlate to disease?how does this correlate to disease?
BackgroundBackgroundBackgroundBackground
miRNA - miR - microRNAmiRNA - miR - microRNAmiRNA - miR - microRNAmiRNA - miR - microRNA
EukaryotesEukaryotes S.S. RNAS.S. RNA 19- to 25-nt19- to 25-nt Non-codingNon-coding
Regulates gene expressionRegulates gene expression
Evolutionary conservedEvolutionary conserved
EukaryotesEukaryotes S.S. RNAS.S. RNA 19- to 25-nt19- to 25-nt Non-codingNon-coding
Regulates gene expressionRegulates gene expression
Evolutionary conservedEvolutionary conserved
≠ ≠ mRNA!mRNA!
Eukaryotic GenomeEukaryotic GenomeEukaryotic GenomeEukaryotic Genome
1/3 of genes1/3 of genes regulatedregulated
222 human miRNAs
ProcessingProcessingProcessingProcessing
DNADNA
pri-miRNApri-miRNA
pre-miRNApre-miRNAExportin 5-induced exportExportin 5-induced export
NUCLEUSNUCLEUS CYTOPLASM CYTOPLASM
transcriptiontranscription
Drosha processingDrosha processingDicer processingDicer processing
maturematuremiRNAmiRNA
RISCRISC
miRNA FunctionmiRNA FunctionmiRNA FunctionmiRNA Function
3’3’ 5’5’AAAAAAAAAAAAAAAAAAAAAA5’5’ 3’3’ mRNAmRNA
TranslationTranslation
miRNA FunctionsmiRNA FunctionsmiRNA FunctionsmiRNA Functions
Cellular division - coordinationCellular division - coordination Developmental apoptosisDevelopmental apoptosis Stress resistanceStress resistance Fat metabolismFat metabolism
Positive or negative regulationPositive or negative regulation
Cellular division - coordinationCellular division - coordination Developmental apoptosisDevelopmental apoptosis Stress resistanceStress resistance Fat metabolismFat metabolism
Positive or negative regulationPositive or negative regulation
Primary LiteraturePrimary LiteraturePrimary LiteraturePrimary Literature
Novel miRNA IDNovel miRNA IDNovel miRNA IDNovel miRNA ID
BioinformaticsBioinformatics Scan for conserved hairpin structureScan for conserved hairpin structure
Conserved, repetitive, protein codingConserved, repetitive, protein codingThermodynamic stabilityThermodynamic stabilityExpression of predicted miRNAs - Expression of predicted miRNAs -
microarraysmicroarraysValidate microarray hits via cloning and Validate microarray hits via cloning and
sequencing, fish via capture biotinylated sequencing, fish via capture biotinylated oligonucleotidesoligonucleotides
Placenta, testis, thymus, brain, prostatePlacenta, testis, thymus, brain, prostate
BioinformaticsBioinformatics Scan for conserved hairpin structureScan for conserved hairpin structure
Conserved, repetitive, protein codingConserved, repetitive, protein codingThermodynamic stabilityThermodynamic stabilityExpression of predicted miRNAs - Expression of predicted miRNAs -
microarraysmicroarraysValidate microarray hits via cloning and Validate microarray hits via cloning and
sequencing, fish via capture biotinylated sequencing, fish via capture biotinylated oligonucleotidesoligonucleotides
Placenta, testis, thymus, brain, prostatePlacenta, testis, thymus, brain, prostate
Bentwich et. al, 2005
Oligonucleotide CaptureOligonucleotide CaptureOligonucleotide CaptureOligonucleotide Capture
miRNA ClustersmiRNA ClustersmiRNA ClustersmiRNA Clusters
Cluster 1 - Chromosome 19 - placenta onlyCluster 1 - Chromosome 19 - placenta only 54 miRNAs - largest cluster discovered54 miRNAs - largest cluster discovered Expression parallels human embryonic stem Expression parallels human embryonic stem
cellscells Conserved in primates onlyConserved in primates only
Cluster 2 - X Chromosome - testis onlyCluster 2 - X Chromosome - testis only 10 miRNAs10 miRNAs
10 conserved in primates only10 conserved in primates only 7 conserved in dog7 conserved in dog 4 conserved in mouse and rat4 conserved in mouse and rat
Cluster 1 - Chromosome 19 - placenta onlyCluster 1 - Chromosome 19 - placenta only 54 miRNAs - largest cluster discovered54 miRNAs - largest cluster discovered Expression parallels human embryonic stem Expression parallels human embryonic stem
cellscells Conserved in primates onlyConserved in primates only
Cluster 2 - X Chromosome - testis onlyCluster 2 - X Chromosome - testis only 10 miRNAs10 miRNAs
10 conserved in primates only10 conserved in primates only 7 conserved in dog7 conserved in dog 4 conserved in mouse and rat4 conserved in mouse and rat
SignificanceSignificanceSignificanceSignificance Human miRNA total >800Human miRNA total >800
(vs current knowledge of 222)(vs current knowledge of 222) Conserved only 400-500Conserved only 400-500
Significant miRNA evolution unique to Significant miRNA evolution unique to primatesprimates 89 Novel miRNAs discovered89 Novel miRNAs discovered
53 not conserved under primates53 not conserved under primates
Primate-specific developmental Primate-specific developmental expression indicates important role in expression indicates important role in mammalian complexitymammalian complexity
Human miRNA total >800Human miRNA total >800 (vs current knowledge of 222)(vs current knowledge of 222) Conserved only 400-500Conserved only 400-500
Significant miRNA evolution unique to Significant miRNA evolution unique to primatesprimates 89 Novel miRNAs discovered89 Novel miRNAs discovered
53 not conserved under primates53 not conserved under primates
Primate-specific developmental Primate-specific developmental expression indicates important role in expression indicates important role in mammalian complexitymammalian complexity
miRNA and CancermiRNA and CancermiRNA and CancermiRNA and Cancer
Novel tumorigenesis gene classNovel tumorigenesis gene classe.g. 13q14 deletions - e.g. 13q14 deletions - miR-15amiR-15a, , miR-miR-
16a16aB cell chronic lymphocytic leukemiaB cell chronic lymphocytic leukemiaMantle cell lymphomaMantle cell lymphomaMultiple myelomaMultiple myelomaProstate cancerProstate cancer
Novel tumorigenesis gene classNovel tumorigenesis gene classe.g. 13q14 deletions - e.g. 13q14 deletions - miR-15amiR-15a, , miR-miR-
16a16aB cell chronic lymphocytic leukemiaB cell chronic lymphocytic leukemiaMantle cell lymphomaMantle cell lymphomaMultiple myelomaMultiple myelomaProstate cancerProstate cancer
miRNA LocationsmiRNA LocationsmiRNA LocationsmiRNA Locations
HPV Integration SitesHPV Integration Sites
FRAs - Fragile SitesFRAs - Fragile Sites Loci prone to metaphase breakageLoci prone to metaphase breakage Rearrangement or deletions in cancerRearrangement or deletions in cancer
CAGR - Cancer-associated genomic regionsCAGR - Cancer-associated genomic regions LOH - loss of heterozygosity (e.g. p53, RB)LOH - loss of heterozygosity (e.g. p53, RB)
HOX - homeobox clustersHOX - homeobox clusters Developmental gene regulationDevelopmental gene regulation
HPV Integration SitesHPV Integration Sites
FRAs - Fragile SitesFRAs - Fragile Sites Loci prone to metaphase breakageLoci prone to metaphase breakage Rearrangement or deletions in cancerRearrangement or deletions in cancer
CAGR - Cancer-associated genomic regionsCAGR - Cancer-associated genomic regions LOH - loss of heterozygosity (e.g. p53, RB)LOH - loss of heterozygosity (e.g. p53, RB)
HOX - homeobox clustersHOX - homeobox clusters Developmental gene regulationDevelopmental gene regulation
oncogenes
Calin et. al, 2003
SummarySummarySummarySummary
microRNAs - 19-25nt - regulates microRNAs - 19-25nt - regulates gene expressiongene expression
Over 800 predicted, many primate Over 800 predicted, many primate conserved, may play role in conserved, may play role in complexitycomplexity
Positive and negative regulation, Positive and negative regulation, novel tumorigenesis gene classnovel tumorigenesis gene class
microRNAs - 19-25nt - regulates microRNAs - 19-25nt - regulates gene expressiongene expression
Over 800 predicted, many primate Over 800 predicted, many primate conserved, may play role in conserved, may play role in complexitycomplexity
Positive and negative regulation, Positive and negative regulation, novel tumorigenesis gene classnovel tumorigenesis gene class
ReferencesReferencesReferencesReferences Bentwich, I., Avniel, A., Karov, Y., Aharonov, R., Gilad, S., Barad, O., Bentwich, I., Avniel, A., Karov, Y., Aharonov, R., Gilad, S., Barad, O.,
Barzilai, A., Einat, P., Einav., E., Meiri., E., Sharon, E., Spector, Y., Barzilai, A., Einat, P., Einav., E., Meiri., E., Sharon, E., Spector, Y., Bentwich, Z. (2005) Identification of hundreds of conserved and Bentwich, Z. (2005) Identification of hundreds of conserved and nonconserved human microRNAs. nonconserved human microRNAs. NatureNature 77: 766-770.: 766-770.
Calin, G.A., Sevignani, C, Dumitru, C.D., Hyslop, T., Noch, E., Yendamuri, Calin, G.A., Sevignani, C, Dumitru, C.D., Hyslop, T., Noch, E., Yendamuri, S., Shimizu, M., Rattan, S., Bullrich, H., Negrini, M., Croce, C.M. (2003) S., Shimizu, M., Rattan, S., Bullrich, H., Negrini, M., Croce, C.M. (2003) Human microRNA genes are frequently located at fragile sites and Human microRNA genes are frequently located at fragile sites and genomic regions involved in cancers. genomic regions involved in cancers. Proc. Natl. Acad. Sci.Proc. Natl. Acad. Sci. 101101: 2999-: 2999-30043004
““microRNA (miRNA) Resource” Ambion. microRNA (miRNA) Resource” Ambion. http://www.ambion.com/techlib/resources/miRNA/index.htmlhttp://www.ambion.com/techlib/resources/miRNA/index.html
““MicroRNA” Wikipedia. MicroRNA” Wikipedia. http://en.wikipedia.org/wiki/MicroRNAhttp://en.wikipedia.org/wiki/MicroRNA
““RNA-induced Silencing Complex” Wikipedia. RNA-induced Silencing Complex” Wikipedia. http://en.wikipedia.org/wiki/RNA-induced_silencing_complexhttp://en.wikipedia.org/wiki/RNA-induced_silencing_complex
Bentwich, I., Avniel, A., Karov, Y., Aharonov, R., Gilad, S., Barad, O., Bentwich, I., Avniel, A., Karov, Y., Aharonov, R., Gilad, S., Barad, O., Barzilai, A., Einat, P., Einav., E., Meiri., E., Sharon, E., Spector, Y., Barzilai, A., Einat, P., Einav., E., Meiri., E., Sharon, E., Spector, Y., Bentwich, Z. (2005) Identification of hundreds of conserved and Bentwich, Z. (2005) Identification of hundreds of conserved and nonconserved human microRNAs. nonconserved human microRNAs. NatureNature 77: 766-770.: 766-770.
Calin, G.A., Sevignani, C, Dumitru, C.D., Hyslop, T., Noch, E., Yendamuri, Calin, G.A., Sevignani, C, Dumitru, C.D., Hyslop, T., Noch, E., Yendamuri, S., Shimizu, M., Rattan, S., Bullrich, H., Negrini, M., Croce, C.M. (2003) S., Shimizu, M., Rattan, S., Bullrich, H., Negrini, M., Croce, C.M. (2003) Human microRNA genes are frequently located at fragile sites and Human microRNA genes are frequently located at fragile sites and genomic regions involved in cancers. genomic regions involved in cancers. Proc. Natl. Acad. Sci.Proc. Natl. Acad. Sci. 101101: 2999-: 2999-30043004
““microRNA (miRNA) Resource” Ambion. microRNA (miRNA) Resource” Ambion. http://www.ambion.com/techlib/resources/miRNA/index.htmlhttp://www.ambion.com/techlib/resources/miRNA/index.html
““MicroRNA” Wikipedia. MicroRNA” Wikipedia. http://en.wikipedia.org/wiki/MicroRNAhttp://en.wikipedia.org/wiki/MicroRNA
““RNA-induced Silencing Complex” Wikipedia. RNA-induced Silencing Complex” Wikipedia. http://en.wikipedia.org/wiki/RNA-induced_silencing_complexhttp://en.wikipedia.org/wiki/RNA-induced_silencing_complex