Approaches

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Approaches
Different resources used to identify regulatory
promoter motifs
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Network discovery driven by large-scale
transcriptional profiling
Vast publicly-available Arabidopsis transcript
data sets
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Respiratory chain
MATRIX
UQ
IMS

• Number of genes in Arabidopsis
• cytochrome pathway approx. 150
• AOX 5
• alternative NAD(P)H DHs - 7

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The alternative respiratory pathway was induced
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ModuleFinder - Biclustering
Salicylic acid (100mM) Salicylic acid
(10mM) Rotenone (3hr) Rotenone (12hr) Chlorampheni
col (3 hr) Chloramphenicol (12 hr) Cysteine H2O2 O
ligomycin (1.25mM) Oligomycin (0.125mM) Anoxia
(N2)
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Identification of regulatory motifs
SR2 SR1 SR3 SR4 SR5 SR6 SR6 SR8
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Models of regulation - testable
Model of module promoters
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Leaf development transcript profiles
QRT-PCR to measure transcripts over 2 rosette
leaf development
0 1 2 3 4 5 6
0 1 2 3 4 5 6
0 1 2 3 4 5 6
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Identification of Import Component Regulatory
Elements
Tom20-2 / Tom20-3
Normalised Transcript Abundance
Leaf age (Weeks)
0 1 2 3 4 5 6
Tom9-1 / Tom20-4
Leaf Development
0 1 2 3 4 5 6
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Identification of Import Component Regulatory
Elements
Tom20-2 / Tom20-3
0 1 2 3 4 5 6
Tom9-1 / Tom20-4
0 1 2 3 4 5 6
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Test predictions
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Testing functionality

Measure reporter gene
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Phylogenetic analysis AtAox1c and GmAox2b
motif
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Aox1a - motif stress specific
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Functional analysis of motifs in leaves
Aox 2kb
Aox 2kb ?? 1683-1677
Aox 1kb
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Identification of common active elements
Aox1a
NDB2
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Model of gene expression for Aox1a and NDB2 upon
treatment with rotenone
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Identification of Import Component Regulatory
Elements
Tom20-2 / Tom20-3
0 1 2 3 4 5 6
Tom9-1 / Tom20-4
0 1 2 3 4 5 6
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Identification of Import Component Regulatory
Elements
Tom20-2 / Tom20-3
0 1 2 3 4 5 6
Site II Elements
Tom9-1 / Tom20-4
0 1 2 3 4 5 6
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Promoter deletion/biolistics
Electrophoretic mobility shift assays
Enhancer function





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Motif identification
Oligo nuc extract
Oligo comp nuc extrct
Oligo comp nuc extract
Oligo nuc extract
Oligo
Oligo
Reporter gene activity
20-4 60-mer
20-4 60-mer ?TGGGCT