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Exon and junction microarrays detect widespread mouse strain- and sex-bias expression differences

ABSTRACT: 71 liver RNA samples from three mouse strains - DBA/2J, C57BL/6J and C3H/HeJ - were profiled using a custom-designed microarray monitoring exon and exon-junction expression of 1,020 genes representing 9,406 exons. Gene expression was calculated via two different methods, using the 3'-most exon probe ("3' gene expression profiling") and using all probes associated with the gene ("whole-transcript gene expression profiling"), while exon expression was determined using exon probes and flanking junction probes that spanned across the neighboring exons("exon expression profiling"). Widespread strain and sex influences were detected using a two-way Analysis of Variance (ANOVA) regardless of the profiling method used. However, over 90% of the genes identified in 3' gene expression profiling or whole transcript profiling were identified in exon profiling, along with 75% and 38% more genes, respectively, showing evidence of differential isoform expression. Overall, 55% and 32% of genes, respectively, exhibited strain- and sex-bias differential gene or exon expression. Keywords: Grouped

ORGANISM(S): Mus musculus

PROVIDER: GSE10736 | GEO | 2008/03/30

SECONDARY ACCESSION(S): PRJNA107511

REPOSITORIES: GEO

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Project description:71 liver RNA samples from three mouse strains - DBA/2J, C57BL/6J and C3H/HeJ - were profiled using a custom-designed microarray monitoring exon and exon-junction expression of 1,020 genes representing 9,406 exons. Gene expression was calculated via two different methods, using the 3'-most exon probe ("3' gene expression profiling") and using all probes associated with the gene ("whole-transcript gene expression profiling"), while exon expression was determined using exon probes and flanking junction probes that spanned across the neighboring exons("exon expression profiling"). Widespread strain and sex influences were detected using a two-way Analysis of Variance (ANOVA) regardless of the profiling method used. However, over 90% of the genes identified in 3' gene expression profiling or whole transcript profiling were identified in exon profiling, along with 75% and 38% more genes, respectively, showing evidence of differential isoform expression. Overall, 55% and 32% of genes, respectively, exhibited strain- and sex-bias differential gene or exon expression. Keywords: Grouped Hybridization material was generated through a random-priming amplification of poly[A]+ purified RNA using primers with a random sequence at the 3' end and a fixed motif at the 5' end that was optimized to generate strand-specific cDNA copies of full-length mRNA transcripts [32]. Since the region used for exon and junction probe selection is constrained to a smaller region, more probes contain sequence with suboptimal characteristics (e.g. high GC content or higher homology to other genes). The hybridization of cDNA, rather than cRNA as commonly done, partially mitigates this issue due to higher specificity and lower background levels [24]. Hybridization conditions were as previously described [33]. All 71 samples were hybridized in a two-channel experiment, where one channel was a common reference, generated by pooling all 71 samples in equal mass. Array hybridizations were done in duplicate with fluor reversal to systematically correct for Cy3/Cy5 dye bias. Array images were processed as described to obtain background noise, single channel intensity and associated measurement error estimates [34]. Expression changes between samples and pool were quantified as mean log10(expression ratio), and associated error.

Project description:Alternative pre-messenger RNA splicing impacts development, physiology, and disease, but its regulation in humans is not well understood, partially due to the limited scale to which the expression of specific splicing events has been measured. We generated the first genome-scale expression compendium of human alternative splicing events using custom whole-transcript microarrays monitoring expression of 24,426 mutually exclusive alternative splicing event pairs in 48 diverse human samples. Over 11,700 genes and 9,500 splicing events were differentially expressed, providing a rich resource for studying splicing regulation. An unbiased, systematic screen of 21,760 4-mer to 7-mer words for cis-regulatory motifs identified 143 RNA 'words' enriched near regulated cassette exons, including six clusters of motifs represented by UCUCU, UGCAUG, UGCU, UGUGU, UUUU, and AGGG, which map to trans-acting regulators PTB, Fox, Muscleblind, CELF/CUG-BP, TIA-1, and hnRNP F/H, respectively. Each cluster showed a distinct pattern of genomic location and tissue specificity. For example, UCUCU occurs 110 to 35 nucleotides preceding cassette exons upregulated in brain and striated muscle but depleted in other tissues. UCUCU and UGCAUG appear to have similar function but independent action, occurring 5' and 3', respectively, of 33% of the cassette exons upregulated in skeletal muscle but co-occurring for only 2%. Keywords: multiple tissue comparison PolyA+ purified RNA pooled from multiple donors of a single human tissue type (e.g. cerebellum) were amplified with random primers and hybridized on a two-color ink-jet oligonucletodie microarray (17 array set) against a common reference pool, comprising ~20 normal adult tissue pools, on custom microarray patterns containing probes to monitor every exon and exon-exon junction in transcript databases, patent databases, and predicted from mouse transcripts. Data were analyzed for gene expression (the average of multiple probes), exon and junction expression, and splice form proportionality (see paper).

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Exon and junction microarrays detect widespread mouse strain- and sex-bias expression differences

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