jclub 2007 aleksandr tkachenko - ut · genepaint.org hierarchical clustering distance metric...
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Jclub 2007
Aleksandr Tkachenko
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The article:
http://dx.doi.org/10.1371/journal.pgen.0030178
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In Situ Hybridization (ISH)
● Expression of genes is manifested by the production of RNA transcripts within cells
● ISH uses a labeled complementary RNA strand to localize a specific RNA sequence in a portion of tissue
● As few as five mRNA molecules per cell can be detected
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In Situ Hybridization (ISH)
● validation of regulatory networks● discovery of network components
Hence, we need:● automatic, high-throughput technology● tools to manipulate expression data for
individual genes (atlases)
Possible applications:
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Genepaint.orgis a digital atlas of gene expression patterns
in the mouse
Features:
● Enables to retrieve expression patterns by gene ids, anatomical locations or sequence homology
● Provides high-resolution visualization
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Genepaint.org
● Hierarchical controlled vocabulary of majority of anatomical structures (organs, tissues)
● Attributes:
– Type of expression (regional, scattered)
– Intensity (not detected, low, medium, strong)
Annotation of expression patterns
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Genepaint.orgAnnotation of expression patterns
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Genepaint.orgHierarchical Clustering
● Genes● Anatomical regions
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Genepaint.orgHierarchical Clustering
Distance metric
● depends on both an expression intensity and a type of pattern
● satisfies criteria:
– genes that do not express in any of the same anatomical regions are maximally distant
– the distance metric is adaptive to the range of expression strengths for a gene.
– pattern is secondary to intensity
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Genepaint.orgHierarchical Clustering
Results:
tissues that have similar embryonic origin and/or similar physiological functions group together
there are groups of genes with similar expression patterns – can form a part of a common signaling pathway.
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Experiment
Clustering results:
7 meaningful groups of anatomical sections12 groups of genes
Organism: midgestation mouse embryo
Anatomical sections: ≈ 90
Genes: ≈1000
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Experiment
Focus on group containing Pax6.
Hypothesis: if genes demonstrate similar behavior in this cluster, they could form a component of regulatory network. We'll try to identify genes regulated by Pax6.
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Experiment
Verification:
1.Knockout of Pax6 caused a change in expression of 17 genes in neocortex
2.Electromobility shift assays identified 12 genes not previously known as targets of Pax6 regulation.
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Conclusion
In situ hybridization when combined with appropriate data mining tools is an effective method for discovery and validation of regulatory network components