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Comparative analysis of mRNA isoform expression in cardiac hypertrophy and development reveals multiple post-transcriptional regulatory modules.

Park JY, Li W, Zheng D, Zhai P, Zhao Y, Matsuda T, Vatner SF, Sadoshima J, Tian B - PLoS ONE (2011)

Bottom Line: Previous studies have shown that the expression pattern of a group of genes in hypertrophied heart induced by pressure overload resembles that at the embryonic stage of heart development, a phenomenon known as activation of the "fetal gene program".Genes with functions in certain pathways, such as cell adhesion and cell morphology, are more likely to be regulated by alternative splicing.Moreover, we found 3'UTRs of mRNAs were generally shortened through alternative cleavage and polyadenylation in hypertrophy, and microRNA target genes were generally de-repressed, suggesting coordinated mechanisms to increase mRNA stability and protein production during hypertrophy.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Molecular Medicine, University of Medicine and Dentistry of New Jersey, Newark, New Jersey, United States of America.

ABSTRACT
Cardiac hypertrophy is enlargement of the heart in response to physiological or pathological stimuli, chiefly involving growth of myocytes in size rather than in number. Previous studies have shown that the expression pattern of a group of genes in hypertrophied heart induced by pressure overload resembles that at the embryonic stage of heart development, a phenomenon known as activation of the "fetal gene program". Here, using a genome-wide approach we systematically defined genes and pathways regulated in short- and long-term cardiac hypertrophy conditions using mice with transverse aortic constriction (TAC), and compared them with those regulated at different stages of embryonic and postnatal development. In addition, exon-level analysis revealed widespread mRNA isoform changes during cardiac hypertrophy resulting from alternative usage of terminal or internal exons, some of which are also developmentally regulated and may be attributable to decreased expression of Fox-1 protein in cardiac hypertrophy. Genes with functions in certain pathways, such as cell adhesion and cell morphology, are more likely to be regulated by alternative splicing. Moreover, we found 3'UTRs of mRNAs were generally shortened through alternative cleavage and polyadenylation in hypertrophy, and microRNA target genes were generally de-repressed, suggesting coordinated mechanisms to increase mRNA stability and protein production during hypertrophy. Taken together, our results comprehensively delineated gene and mRNA isoform regulation events in cardiac hypertrophy and revealed their relations to those in development, and suggested that modulation of mRNA isoform expression plays an importance role in heart remodeling under pressure overload.

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Regulation of 3′UTR isoforms in cardiac hypertrophy and development.(A) Schematic of APA and our method to detect 3′UTR length changes using microarray probes. A hypothetical gene contains three polyA sites, resulting in three 3′UTR isoforms. The 3′UTR regions upstream and downstream of the first polyA site are called constitutive and alternative UTRs, or cUTR and aUTR, respectively. Probes mapped to upstream and downstream of each polyA site were used to calculate the RUD score, which reflects the relative expression of 3′UTR isoforms. The RUD score correlates with 3′UTR length. (B) Comparison of ratio of probe intensity for the downstream region of polyA site to that of upstream region (D/U) of polyA site between Sham and 1 W TAC (LVH3). A total of 4,112 genes were examined. Genes with significant regulation of 3′UTR length (P<0.1, T-test) are colored, with red for genes with 3′UTR lengthened in 1 W TAC and green for those with 3′UTR shortened. (C) Global RUD changes in cardiac development and hypertrophy. The median RUD of all genes in each sample was plotted to represent RUD of the sample. All embryonic samples were compared to E10.5, and TAC samples were compared to Sham. Error bars are standard error of the mean (SEM) based on multiple samples. (D) Venn diagram showing numbers of genes with UTR lengthened or shortened in development and hypertrophy. We selected genes with consistent regulation in at least 4 out of 6 ED samples and in at least 2 out of 3 LVH samples. Fisher's exact test was used to assess significance of the overlap between development and hypertrophy. (E) qRT-PCR analysis of genes with shortened 3′UTRs in TAC. Two sets of PCR primers were designed to target regions upstream and downstream of the first polyA site as shown in (A). The difference in their ratio was used to indicate 3′UTR length changes. PCR primer sequences are listed in Table S2, and gene structures are shown in Figure S6.
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pone-0022391-g005: Regulation of 3′UTR isoforms in cardiac hypertrophy and development.(A) Schematic of APA and our method to detect 3′UTR length changes using microarray probes. A hypothetical gene contains three polyA sites, resulting in three 3′UTR isoforms. The 3′UTR regions upstream and downstream of the first polyA site are called constitutive and alternative UTRs, or cUTR and aUTR, respectively. Probes mapped to upstream and downstream of each polyA site were used to calculate the RUD score, which reflects the relative expression of 3′UTR isoforms. The RUD score correlates with 3′UTR length. (B) Comparison of ratio of probe intensity for the downstream region of polyA site to that of upstream region (D/U) of polyA site between Sham and 1 W TAC (LVH3). A total of 4,112 genes were examined. Genes with significant regulation of 3′UTR length (P<0.1, T-test) are colored, with red for genes with 3′UTR lengthened in 1 W TAC and green for those with 3′UTR shortened. (C) Global RUD changes in cardiac development and hypertrophy. The median RUD of all genes in each sample was plotted to represent RUD of the sample. All embryonic samples were compared to E10.5, and TAC samples were compared to Sham. Error bars are standard error of the mean (SEM) based on multiple samples. (D) Venn diagram showing numbers of genes with UTR lengthened or shortened in development and hypertrophy. We selected genes with consistent regulation in at least 4 out of 6 ED samples and in at least 2 out of 3 LVH samples. Fisher's exact test was used to assess significance of the overlap between development and hypertrophy. (E) qRT-PCR analysis of genes with shortened 3′UTRs in TAC. Two sets of PCR primers were designed to target regions upstream and downstream of the first polyA site as shown in (A). The difference in their ratio was used to indicate 3′UTR length changes. PCR primer sequences are listed in Table S2, and gene structures are shown in Figure S6.

Mentions: Regulation of 3′UTRs by alternative cleavage and polyadenylation (APA) has recently been found to be associated with cell proliferation, oncogenic transformation, and development [34], [35], [36]. To examine how 3′UTRs are regulated in hypertrophy, we took advantage of the fact that a large number of Affymetrix GeneChip probes hybridize to constitutive 3′UTRs (cUTRs) and alternative 3′UTRs (aUTRs) (Figure 5A). cUTRs and aUTRs are defined by polyA sites located in 3′UTRs. We used a score, named relative expression of mRNA isoforms using distal polyA sites (RUD), to indicate the 3′UTR length as illustrated in Figure 5A. RUD was based on comparison of probe intensities for the upstream and downstream regions of a polyA site; a higher score indicates relatively higher abundance of mRNA isoforms resulting from usage of promoter-distal polyA sites, and thus longer 3′UTRs.


Comparative analysis of mRNA isoform expression in cardiac hypertrophy and development reveals multiple post-transcriptional regulatory modules.

Park JY, Li W, Zheng D, Zhai P, Zhao Y, Matsuda T, Vatner SF, Sadoshima J, Tian B - PLoS ONE (2011)

Regulation of 3′UTR isoforms in cardiac hypertrophy and development.(A) Schematic of APA and our method to detect 3′UTR length changes using microarray probes. A hypothetical gene contains three polyA sites, resulting in three 3′UTR isoforms. The 3′UTR regions upstream and downstream of the first polyA site are called constitutive and alternative UTRs, or cUTR and aUTR, respectively. Probes mapped to upstream and downstream of each polyA site were used to calculate the RUD score, which reflects the relative expression of 3′UTR isoforms. The RUD score correlates with 3′UTR length. (B) Comparison of ratio of probe intensity for the downstream region of polyA site to that of upstream region (D/U) of polyA site between Sham and 1 W TAC (LVH3). A total of 4,112 genes were examined. Genes with significant regulation of 3′UTR length (P<0.1, T-test) are colored, with red for genes with 3′UTR lengthened in 1 W TAC and green for those with 3′UTR shortened. (C) Global RUD changes in cardiac development and hypertrophy. The median RUD of all genes in each sample was plotted to represent RUD of the sample. All embryonic samples were compared to E10.5, and TAC samples were compared to Sham. Error bars are standard error of the mean (SEM) based on multiple samples. (D) Venn diagram showing numbers of genes with UTR lengthened or shortened in development and hypertrophy. We selected genes with consistent regulation in at least 4 out of 6 ED samples and in at least 2 out of 3 LVH samples. Fisher's exact test was used to assess significance of the overlap between development and hypertrophy. (E) qRT-PCR analysis of genes with shortened 3′UTRs in TAC. Two sets of PCR primers were designed to target regions upstream and downstream of the first polyA site as shown in (A). The difference in their ratio was used to indicate 3′UTR length changes. PCR primer sequences are listed in Table S2, and gene structures are shown in Figure S6.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3142162&req=5

pone-0022391-g005: Regulation of 3′UTR isoforms in cardiac hypertrophy and development.(A) Schematic of APA and our method to detect 3′UTR length changes using microarray probes. A hypothetical gene contains three polyA sites, resulting in three 3′UTR isoforms. The 3′UTR regions upstream and downstream of the first polyA site are called constitutive and alternative UTRs, or cUTR and aUTR, respectively. Probes mapped to upstream and downstream of each polyA site were used to calculate the RUD score, which reflects the relative expression of 3′UTR isoforms. The RUD score correlates with 3′UTR length. (B) Comparison of ratio of probe intensity for the downstream region of polyA site to that of upstream region (D/U) of polyA site between Sham and 1 W TAC (LVH3). A total of 4,112 genes were examined. Genes with significant regulation of 3′UTR length (P<0.1, T-test) are colored, with red for genes with 3′UTR lengthened in 1 W TAC and green for those with 3′UTR shortened. (C) Global RUD changes in cardiac development and hypertrophy. The median RUD of all genes in each sample was plotted to represent RUD of the sample. All embryonic samples were compared to E10.5, and TAC samples were compared to Sham. Error bars are standard error of the mean (SEM) based on multiple samples. (D) Venn diagram showing numbers of genes with UTR lengthened or shortened in development and hypertrophy. We selected genes with consistent regulation in at least 4 out of 6 ED samples and in at least 2 out of 3 LVH samples. Fisher's exact test was used to assess significance of the overlap between development and hypertrophy. (E) qRT-PCR analysis of genes with shortened 3′UTRs in TAC. Two sets of PCR primers were designed to target regions upstream and downstream of the first polyA site as shown in (A). The difference in their ratio was used to indicate 3′UTR length changes. PCR primer sequences are listed in Table S2, and gene structures are shown in Figure S6.
Mentions: Regulation of 3′UTRs by alternative cleavage and polyadenylation (APA) has recently been found to be associated with cell proliferation, oncogenic transformation, and development [34], [35], [36]. To examine how 3′UTRs are regulated in hypertrophy, we took advantage of the fact that a large number of Affymetrix GeneChip probes hybridize to constitutive 3′UTRs (cUTRs) and alternative 3′UTRs (aUTRs) (Figure 5A). cUTRs and aUTRs are defined by polyA sites located in 3′UTRs. We used a score, named relative expression of mRNA isoforms using distal polyA sites (RUD), to indicate the 3′UTR length as illustrated in Figure 5A. RUD was based on comparison of probe intensities for the upstream and downstream regions of a polyA site; a higher score indicates relatively higher abundance of mRNA isoforms resulting from usage of promoter-distal polyA sites, and thus longer 3′UTRs.

Bottom Line: Previous studies have shown that the expression pattern of a group of genes in hypertrophied heart induced by pressure overload resembles that at the embryonic stage of heart development, a phenomenon known as activation of the "fetal gene program".Genes with functions in certain pathways, such as cell adhesion and cell morphology, are more likely to be regulated by alternative splicing.Moreover, we found 3'UTRs of mRNAs were generally shortened through alternative cleavage and polyadenylation in hypertrophy, and microRNA target genes were generally de-repressed, suggesting coordinated mechanisms to increase mRNA stability and protein production during hypertrophy.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Molecular Medicine, University of Medicine and Dentistry of New Jersey, Newark, New Jersey, United States of America.

ABSTRACT
Cardiac hypertrophy is enlargement of the heart in response to physiological or pathological stimuli, chiefly involving growth of myocytes in size rather than in number. Previous studies have shown that the expression pattern of a group of genes in hypertrophied heart induced by pressure overload resembles that at the embryonic stage of heart development, a phenomenon known as activation of the "fetal gene program". Here, using a genome-wide approach we systematically defined genes and pathways regulated in short- and long-term cardiac hypertrophy conditions using mice with transverse aortic constriction (TAC), and compared them with those regulated at different stages of embryonic and postnatal development. In addition, exon-level analysis revealed widespread mRNA isoform changes during cardiac hypertrophy resulting from alternative usage of terminal or internal exons, some of which are also developmentally regulated and may be attributable to decreased expression of Fox-1 protein in cardiac hypertrophy. Genes with functions in certain pathways, such as cell adhesion and cell morphology, are more likely to be regulated by alternative splicing. Moreover, we found 3'UTRs of mRNAs were generally shortened through alternative cleavage and polyadenylation in hypertrophy, and microRNA target genes were generally de-repressed, suggesting coordinated mechanisms to increase mRNA stability and protein production during hypertrophy. Taken together, our results comprehensively delineated gene and mRNA isoform regulation events in cardiac hypertrophy and revealed their relations to those in development, and suggested that modulation of mRNA isoform expression plays an importance role in heart remodeling under pressure overload.

Show MeSH
Related in: MedlinePlus