• Classical epigenetic systemsClassical epigenetic systems
• Gene silencingGene silencing
• Viral cross-protectionViral cross-protection
• Epigenetics in plant developmentEpigenetics in plant development
• Classical epigenetic systemsClassical epigenetic systems
• Gene silencingGene silencing
• Viral cross-protectionViral cross-protection
• Epigenetics in plant developmentEpigenetics in plant development
• Transposons - change of phase Transposons - change of phase
• Paramutation in maizeParamutation in maize
• Transposons - change of phase Transposons - change of phase
• Paramutation in maizeParamutation in maize
CLASSICAL EPIGENETIC SYSTEMSCLASSICAL EPIGENETIC SYSTEMS CLASSICAL EPIGENETIC SYSTEMSCLASSICAL EPIGENETIC SYSTEMS
• Heritable, but reversibleHeritable, but reversible
• Epimutants differ in theirEpimutants differ in their developmental expression patterns developmental expression patterns
• The transition from active to cryptic (and theThe transition from active to cryptic (and the reverse) takes several plant generations reverse) takes several plant generations
• Heritable, but reversibleHeritable, but reversible
• Epimutants differ in theirEpimutants differ in their developmental expression patterns developmental expression patterns
• The transition from active to cryptic (and theThe transition from active to cryptic (and the reverse) takes several plant generations reverse) takes several plant generations
Changes in Changes in SpmSpm activity phase activity phase Changes in Changes in SpmSpm activity phase activity phase
• McClintock: an inactive transposon wakes up when anMcClintock: an inactive transposon wakes up when an active transposon is present, but segregates unchanged active transposon is present, but segregates unchanged
• Fedoroff: an active element can heritably wake up anFedoroff: an active element can heritably wake up an inactive or a cryptic element inactive or a cryptic element
• The transition from active to cryptic (and the reverse) The transition from active to cryptic (and the reverse) takes several plant generations takes several plant generations
• McClintock: an inactive transposon wakes up when anMcClintock: an inactive transposon wakes up when an active transposon is present, but segregates unchanged active transposon is present, but segregates unchanged
• Fedoroff: an active element can heritably wake up anFedoroff: an active element can heritably wake up an inactive or a cryptic element inactive or a cryptic element
• The transition from active to cryptic (and the reverse) The transition from active to cryptic (and the reverse) takes several plant generations takes several plant generations
Genetic analysis of phase changeGenetic analysis of phase change Genetic analysis of phase changeGenetic analysis of phase change
Paramutation at the Paramutation at the R R locus in maizelocus in maizeParamutation at the Paramutation at the R R locus in maizelocus in maize
• A directed, heritable change in gene expressionA directed, heritable change in gene expression
• r-str-st and and r-mb r-mb termed PARAMUTAGENIC termed PARAMUTAGENIC
• R-r termed PARAMUTABLER-r termed PARAMUTABLE
• Altered expression is heritableAltered expression is heritable
• Partial reversion when homozygousPartial reversion when homozygous
• A paramutable allele can become paramutagenicA paramutable allele can become paramutagenic upon exposure to a paramutagenic allele upon exposure to a paramutagenic allele
• A directed, heritable change in gene expressionA directed, heritable change in gene expression
• r-str-st and and r-mb r-mb termed PARAMUTAGENIC termed PARAMUTAGENIC
• R-r termed PARAMUTABLER-r termed PARAMUTABLE
• Altered expression is heritableAltered expression is heritable
• Partial reversion when homozygousPartial reversion when homozygous
• A paramutable allele can become paramutagenicA paramutable allele can become paramutagenic upon exposure to a paramutagenic allele upon exposure to a paramutagenic allele
Brink, R. A., Styles, E. D. and Axtell, J. D. (1968) Science, 159: 161-170Brink, R. A., Styles, E. D. and Axtell, J. D. (1968) Science, 159: 161-170 Brink, R. A., Styles, E. D. and Axtell, J. D. (1968) Science, 159: 161-170Brink, R. A., Styles, E. D. and Axtell, J. D. (1968) Science, 159: 161-170
R R gene paramutation in maizegene paramutation in maize R R gene paramutation in maizegene paramutation in maize
Walker, E. L. (1998), Genetics, 148: 1973-1981Walker, E. L. (1998), Genetics, 148: 1973-1981 Walker, E. L. (1998), Genetics, 148: 1973-1981Walker, E. L. (1998), Genetics, 148: 1973-1981
Structure of a paramutagenic R alleleStructure of a paramutagenic R alleleStructure of a paramutagenic R alleleStructure of a paramutagenic R allele
Kermicle, J. L., Eggleston, W. B. and Alleman, M. (1995), Genetics, 141: 361-372Kermicle, J. L., Eggleston, W. B. and Alleman, M. (1995), Genetics, 141: 361-372 Kermicle, J. L., Eggleston, W. B. and Alleman, M. (1995), Genetics, 141: 361-372Kermicle, J. L., Eggleston, W. B. and Alleman, M. (1995), Genetics, 141: 361-372
• The R-st allele contains several highly homologous repeatsThe R-st allele contains several highly homologous repeats
• Paramutagenicity is directly proportional to the number of repeatsParamutagenicity is directly proportional to the number of repeats
• Transcription start sites are methylatedTranscription start sites are methylated
• The R-st allele contains several highly homologous repeatsThe R-st allele contains several highly homologous repeats
• Paramutagenicity is directly proportional to the number of repeatsParamutagenicity is directly proportional to the number of repeats
• Transcription start sites are methylatedTranscription start sites are methylated
Structure of the paramutable R-r alleleStructure of the paramutable R-r allele Structure of the paramutable R-r alleleStructure of the paramutable R-r allele
Walker, E. L. (1998), Genetics, 148: 1973-1981Walker, E. L. (1998), Genetics, 148: 1973-1981 Walker, E. L. (1998), Genetics, 148: 1973-1981Walker, E. L. (1998), Genetics, 148: 1973-1981
Common themes inCommon themes intransposon inactivation and paramutationtransposon inactivation and paramutation
Common themes inCommon themes intransposon inactivation and paramutationtransposon inactivation and paramutation
• Sequence duplication is centralSequence duplication is central
• Promoter sequences are methylatedPromoter sequences are methylated
• Genes/TEs transcriptionally silencedGenes/TEs transcriptionally silenced
• Silencing is heritable, but reversibleSilencing is heritable, but reversible
• Both involve transposon sequencesBoth involve transposon sequences
• Sequence duplication is centralSequence duplication is central
• Promoter sequences are methylatedPromoter sequences are methylated
• Genes/TEs transcriptionally silencedGenes/TEs transcriptionally silenced
• Silencing is heritable, but reversibleSilencing is heritable, but reversible
• Both involve transposon sequencesBoth involve transposon sequences
Gene silencing (co-suppression) by trangenesGene silencing (co-suppression) by trangenes Gene silencing (co-suppression) by trangenesGene silencing (co-suppression) by trangenes
• Transgenes can silence endogenous genesTransgenes can silence endogenous genes
• More transgenes, more gene silencingMore transgenes, more gene silencing
• Inverted repeats are especially effectiveInverted repeats are especially effective
• Silenced genes are often methylatedSilenced genes are often methylated
• Silencing can be heritableSilencing can be heritable
• Silenced genes can be “paramutagenic”Silenced genes can be “paramutagenic”
• Transgenes can silence endogenous genesTransgenes can silence endogenous genes
• More transgenes, more gene silencingMore transgenes, more gene silencing
• Inverted repeats are especially effectiveInverted repeats are especially effective
• Silenced genes are often methylatedSilenced genes are often methylated
• Silencing can be heritableSilencing can be heritable
• Silenced genes can be “paramutagenic”Silenced genes can be “paramutagenic”
Que, Q, Want, H.-Y, and Jorgensen, R.A. (1998). Plant J. 13: 401-9Que, Q, Want, H.-Y, and Jorgensen, R.A. (1998). Plant J. 13: 401-9 Que, Q, Want, H.-Y, and Jorgensen, R.A. (1998). Plant J. 13: 401-9Que, Q, Want, H.-Y, and Jorgensen, R.A. (1998). Plant J. 13: 401-9
Transcriptional and post-transcription silencingTranscriptional and post-transcription silencing (TGS and PTGS)(TGS and PTGS) Transcriptional and post-transcription silencingTranscriptional and post-transcription silencing (TGS and PTGS)(TGS and PTGS)
• Silencing can be transcriptional, post-transcriptional or bothSilencing can be transcriptional, post-transcriptional or both
• TGS is associated with promoter methylationTGS is associated with promoter methylation
• PTGS is associated with coding sequence methylationPTGS is associated with coding sequence methylation
• Promotor methylation is not required for initiation of silencingPromotor methylation is not required for initiation of silencing
• Methylation is required for the maintenance of silencingMethylation is required for the maintenance of silencing
• Silencing can be transcriptional, post-transcriptional or bothSilencing can be transcriptional, post-transcriptional or both
• TGS is associated with promoter methylationTGS is associated with promoter methylation
• PTGS is associated with coding sequence methylationPTGS is associated with coding sequence methylation
• Promotor methylation is not required for initiation of silencingPromotor methylation is not required for initiation of silencing
• Methylation is required for the maintenance of silencingMethylation is required for the maintenance of silencing
Gene silencing and viral resistanceGene silencing and viral resistance Gene silencing and viral resistanceGene silencing and viral resistance
• Viral infection confers immunity to further infectionViral infection confers immunity to further infection
• Transgenic plants expressing coat protein are resistantTransgenic plants expressing coat protein are resistant
• Viral infection confers immunity to further infectionViral infection confers immunity to further infection
• Transgenic plants expressing coat protein are resistantTransgenic plants expressing coat protein are resistant
Ratcliff, F., Harrison, B. D. and Baulcombe, D. C. (1997). Science 276: 1558-1560Ratcliff, F., Harrison, B. D. and Baulcombe, D. C. (1997). Science 276: 1558-1560Ratcliff, F., Harrison, B. D. and Baulcombe, D. C. (1997). Science 276: 1558-1560Ratcliff, F., Harrison, B. D. and Baulcombe, D. C. (1997). Science 276: 1558-1560
Viral resistance is RNA-mediatedViral resistance is RNA-mediated Viral resistance is RNA-mediatedViral resistance is RNA-mediated
• Transgene-induced resistance resembles PTGSTransgene-induced resistance resembles PTGS
• Resistance is mediated by RNAResistance is mediated by RNA
• Virus infection can result in co-suppression Virus infection can result in co-suppression
• Transgene-induced resistance resembles PTGSTransgene-induced resistance resembles PTGS
• Resistance is mediated by RNAResistance is mediated by RNA
• Virus infection can result in co-suppression Virus infection can result in co-suppression
Ratcliff, F., Harrison, B. D. and Baulcombe, D. C. (1997). Science 276: 1558-1560Ratcliff, F., Harrison, B. D. and Baulcombe, D. C. (1997). Science 276: 1558-1560Ratcliff, F., Harrison, B. D. and Baulcombe, D. C. (1997). Science 276: 1558-1560Ratcliff, F., Harrison, B. D. and Baulcombe, D. C. (1997). Science 276: 1558-1560
PVXPVX
W22W22
PVX. PVX. W22W22
Gene silencing: a systemic signalGene silencing: a systemic signal Gene silencing: a systemic signalGene silencing: a systemic signal
Voinnet, O., and Baulcombe, D. C. (1997). Nature 389: 553Voinnet, O., and Baulcombe, D. C. (1997). Nature 389: 553Voinnet, O., and Baulcombe, D. C. (1997). Nature 389: 553Voinnet, O., and Baulcombe, D. C. (1997). Nature 389: 553
The systemic gene silencing signal is RNAThe systemic gene silencing signal is RNA The systemic gene silencing signal is RNAThe systemic gene silencing signal is RNA
• Non-overlapping gene fragments cross-silenceNon-overlapping gene fragments cross-silence
• RNA moves between cells in plantsRNA moves between cells in plants
• Plants encode RNA-dependent RNA polymerasesPlants encode RNA-dependent RNA polymerases
• Non-overlapping gene fragments cross-silenceNon-overlapping gene fragments cross-silence
• RNA moves between cells in plantsRNA moves between cells in plants
• Plants encode RNA-dependent RNA polymerasesPlants encode RNA-dependent RNA polymerases
TGS and PTGS: is there a relationship?TGS and PTGS: is there a relationship? TGS and PTGS: is there a relationship?TGS and PTGS: is there a relationship?
Wassenegger, M., Heimes, S., Reidel, L., and Sanger, H. L. (1994) Cell 76: 567-76.Wassenegger, M., Heimes, S., Reidel, L., and Sanger, H. L. (1994) Cell 76: 567-76.Wassenegger, M., Heimes, S., Reidel, L., and Sanger, H. L. (1994) Cell 76: 567-76.Wassenegger, M., Heimes, S., Reidel, L., and Sanger, H. L. (1994) Cell 76: 567-76.
P35S PSTVd cDNA pAnos P35S PSTVd cDNA pAnos P35S PSTVd cDNA pAnos P35S PSTVd cDNA pAnos
Replication competent Replication competent Replication competent Replication competent Replication incompetent Replication incompetent Replication incompetent Replication incompetent
Transcription onlyTranscription only
No replicationNo replication
No methylationNo methylation
Transcription onlyTranscription only
No replicationNo replication
No methylationNo methylation
TranscriptionTranscription
ReplicationReplication
MethylationMethylation
TranscriptionTranscription
ReplicationReplication
MethylationMethylation
microRNAs and silencingRNAs in plantsmicroRNAs and silencingRNAs in plants microRNAs and silencingRNAs in plantsmicroRNAs and silencingRNAs in plants
Mallory, A. C., and Vaucheret, H. (2004) Current Opinion in Plant Biology, 7:120-125.Mallory, A. C., and Vaucheret, H. (2004) Current Opinion in Plant Biology, 7:120-125.Mallory, A. C., and Vaucheret, H. (2004) Current Opinion in Plant Biology, 7:120-125.Mallory, A. C., and Vaucheret, H. (2004) Current Opinion in Plant Biology, 7:120-125.
microRNAs and silencingRNAs in animalsmicroRNAs and silencingRNAs in animals microRNAs and silencingRNAs in animalsmicroRNAs and silencingRNAs in animals
Mallory, A. C., and Vaucheret, H. (2004) Current Opinion in Plant Biology, 7:120-125.Mallory, A. C., and Vaucheret, H. (2004) Current Opinion in Plant Biology, 7:120-125.Mallory, A. C., and Vaucheret, H. (2004) Current Opinion in Plant Biology, 7:120-125.Mallory, A. C., and Vaucheret, H. (2004) Current Opinion in Plant Biology, 7:120-125.
The Arabidopsis hyl1 mutation The Arabidopsis hyl1 mutation
wildtypewildtype
hyl1hyl1
0.6 µM ABA0.6 µM ABANo ABANo ABA
The hyl1 mutation affects miRNA levelsThe hyl1 mutation affects miRNA levels
wtwt hyl1hyl1hen1-1hen1-1 11 33
ARF8ARF8
SCL6-IIISCL6-III
rRNArRNA
MYB33MYB33
35S::HYL135S::HYL1
wtwt hyl1hyl1hen1-1hen1-1 11 33
35S::HYL135S::HYL1
miR167miR167
miR171miR171
tRNA +5S rRNAtRNA +5S rRNA
miR159miR159
UBQ1UBQ1DCL1DCL1
wtwt hyl1hyl1 hen1-1hen1-1 wtwt hyl1hyl1 hen1-1hen1-1
The hyl1 mutation affects mRNA stabilityThe hyl1 mutation affects mRNA stability
B.
Time (hrs)Time (hrs)
% in
itial
val
ue%
initi
al v
alue
10
100
00 44 88 1212
MYB33MYB33
hyl1hyl1
wtwt
35S::HYL135S::HYL1
50
30
hyl1hyl1
wtwt
35S::HYL135S::HYL1
SCL6-III SCL6-III
00 44 88 1212
hyl1hyl1
wtwt
35S::HYL135S::HYL1
00 44 88 1212
ANP1ANP1
hyl1hyl1
wtwt
35S::HYL135S::HYL1
ARF8 ARF8
00 44 88 1212
HYL1 is in
nuclear bodies
HYL1 is in
nuclear bodies
• TnpA and TnpD are required for transpositionTnpA and TnpD are required for transposition
• TnpA is also a weak transcription factorTnpA is also a weak transcription factor
• TnpA and TnpD are required for transpositionTnpA and TnpD are required for transposition
• TnpA is also a weak transcription factorTnpA is also a weak transcription factor
SpmSpm has one gene, has one gene, but codes for two proteinsbut codes for two proteins
SpmSpm has one gene, has one gene, but codes for two proteinsbut codes for two proteins
TranspositionTransposition
active active SpmSpmTnpATnpA
promoterpromoter
TnpD
TnpDmRNA
TnpAmRNA
TnpD
• Promoter methylated, element inactivePromoter methylated, element inactive
• Methylation of GC-rich sequence confers heritabilityMethylation of GC-rich sequence confers heritability
• Reversed by Reversed by SpmSpm-encoded TnpA-encoded TnpA
• Promoter methylated, element inactivePromoter methylated, element inactive
• Methylation of GC-rich sequence confers heritabilityMethylation of GC-rich sequence confers heritability
• Reversed by Reversed by SpmSpm-encoded TnpA-encoded TnpA
Changes in Changes in SpmSpm activity phase activity phase Changes in Changes in SpmSpm activity phase activity phase
Methylated siteMethylated siteUnmethylated siteUnmethylated site
cryptic cryptic SpmSpm active active SpmSpmTnpATnpA
promoterpromoter
GC-rich sequenceGC-rich sequence
• TnpA is a weak transcription factor TnpA is a weak transcription factor
• TnpA binds unmethylated and hemimethylated DNATnpA binds unmethylated and hemimethylated DNA
• TnpA promotes active demethylation TnpA promotes active demethylation
• TnpA is a weak transcription factor TnpA is a weak transcription factor
• TnpA binds unmethylated and hemimethylated DNATnpA binds unmethylated and hemimethylated DNA
• TnpA promotes active demethylation TnpA promotes active demethylation
Molecular mechanism of Molecular mechanism of Spm Spm activationactivation Molecular mechanism of Molecular mechanism of Spm Spm activationactivation
Methyl groupMethyl grouppromoterpromoter
replicationreplication
TnpATnpA
TnpATnpATnpATnpA
Transposon silencing: the chromatin connection Transposon silencing: the chromatin connection Transposon silencing: the chromatin connection Transposon silencing: the chromatin connection
transpositiontranspositiontranspositiontransposition
silencingsilencingsilencingsilencing
mRNAmRNA
siRNAs?siRNAs?siRNAs?siRNAs?
siRNAssiRNAsDNA methylaseDNA methylasehistone deacetylasehistone deacetylasechromatin remodeling proteins chromatin remodeling proteins
siRNAssiRNAsDNA methylaseDNA methylasehistone deacetylasehistone deacetylasechromatin remodeling proteins chromatin remodeling proteins
The story of papaya ringspot virus
The story of papaya ringspot virus
http://www.apsnet.org/education/feature/papaya/Top.htm
http://www.apsnet.org/education/feature/papaya/Top.htm
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are needed to see this picture.
QuickTime™ and aTIFF (Uncompressed) decompressor
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Papaya ringspot virusPapaya ringspot virus
http://www.apsnet.org/education/feature/papaya/Top.htmhttp://www.apsnet.org/education/feature/papaya/Top.htm
1940s: PRS virus discovered in Hawaii1940s: PRS virus discovered in Hawaii
1950s: Oahu’s papaya industry wiped out1950s: Oahu’s papaya industry wiped out
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1960s: Papaya industry moves to Puna district1960s: Papaya industry moves to Puna district
Papaya ringspot virusPapaya ringspot virus
TGSTGSTGSTGS
• NoNo• NoNo
Papaya ringspot virusPapaya ringspot virus
1991: First transgenic PRSV-resistant papaya plant1991: First transgenic PRSV-resistant papaya plant
1980s: PRSV-resistance project started under direction of Dennis Gonsalves1980s: PRSV-resistance project started under direction of Dennis Gonsalves
1992: PRSV discovered in Puna district1992: PRSV discovered in Puna district
1992: First field trials PRSV-resistant papaya plants1992: First field trials PRSV-resistant papaya plants
1994: USDA granted permission for large scale field trials1994: USDA granted permission for large scale field trials
1995-97: Approvals for release from USDA, EPA, FDA1995-97: Approvals for release from USDA, EPA, FDA
1992-1977: PRVS spread; many farmers went out of business1992-1977: PRVS spread; many farmers went out of business
1998: Seeds released, free of charge, to growers1998: Seeds released, free of charge, to growers
2000: Papaya industry bounced back; crop back to pre-1995 levels2000: Papaya industry bounced back; crop back to pre-1995 levels
Papaya ringspot virusPapaya ringspot virus
http://www.apsnet.org/education/feature/papaya/Top.htmhttp://www.apsnet.org/education/feature/papaya/Top.htm
Papaya ringspot
virus
Papaya ringspot
virus
Epigenetic mechanisms: plantEpigenetic mechanisms: plantevolution, defense and development evolution, defense and development
Epigenetic mechanisms: plantEpigenetic mechanisms: plantevolution, defense and development evolution, defense and development
• Gene silencing is a response to gene duplication Gene silencing is a response to gene duplication (evolution of duplicated genes; transposon control)(evolution of duplicated genes; transposon control)
• Gene silencing is a response to gene overexpression Gene silencing is a response to gene overexpression (dosage compensation)(dosage compensation)
• Gene silencing is a defense response Gene silencing is a defense response (viral cross protection; rapid environmental responses)(viral cross protection; rapid environmental responses)
• Epigenetic mechanisms are used in plant development Epigenetic mechanisms are used in plant development (JAW miRNA in leaf morphogenesis)(JAW miRNA in leaf morphogenesis)
• Gene silencing is a response to gene duplication Gene silencing is a response to gene duplication (evolution of duplicated genes; transposon control)(evolution of duplicated genes; transposon control)
• Gene silencing is a response to gene overexpression Gene silencing is a response to gene overexpression (dosage compensation)(dosage compensation)
• Gene silencing is a defense response Gene silencing is a defense response (viral cross protection; rapid environmental responses)(viral cross protection; rapid environmental responses)
• Epigenetic mechanisms are used in plant development Epigenetic mechanisms are used in plant development (JAW miRNA in leaf morphogenesis)(JAW miRNA in leaf morphogenesis)