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A global analysis of the complex landscape of isoforms and regulatory networks of p63 in human cells and tissues.

Sethi I, Romano RA, Gluck C, Smalley K, Vojtesek B, Buck MJ, Sinha S - BMC Genomics (2015)

Bottom Line: Furthermore using unsupervised clustering of human cell lines, tissues and organs, we show that ΔNp63 and TAp63 driven transcriptional networks involve very distinct sets of molecular players, which may underlie their different biological functions.We curate publicly available data generated in part by consortiums such as ENCODE, FANTOM and Human Protein Atlas to delineate the vastly different transcriptomic landscapes of ΔNp63 and TAp63.Our studies help not only in dispelling prevailing myths and controversies on p63 expression in commonly used human cell lines but also augur new isoform- and cell type-specific activities of p63.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Center of Excellence in Bioinformatics and Life Sciences, State University of New York, 701 Ellicott Street, Buffalo, NY, 14203, USA.

ABSTRACT

Background: The transcription factor p63 belongs to the p53/p63/p73 family and plays key functional roles during normal epithelial development and differentiation and in pathological states such as squamous cell carcinomas. The human TP63 gene, located on chromosome 3q28 is driven by two promoters that generate the full-length transactivating (TA) and N-terminal truncated (ΔN) isoforms. Furthermore alternative splicing at the C-terminus gives rise to additional α, β, γ and likely several other minor variants. Teasing out the expression and biological function of each p63 variant has been both the focus of, and a cause for contention in the p63 field.

Results: Here we have taken advantage of a burgeoning RNA-Seq based genomic data-sets to examine the global expression profiles of p63 isoforms across commonly utilized human cell-lines and major tissues and organs. Consistent with earlier studies, we find ΔNp63 transcripts, primarily that of the ΔNp63α isoforms, to be expressed in most cells of epithelial origin such as those of skin and oral tissues, mammary glands and squamous cell carcinomas. In contrast, TAp63 is not expressed in the majority of normal cell-types and tissues; rather it is selectively expressed at moderate to high levels in a subset of Burkitt's and diffuse large B-cell lymphoma cell lines. We verify this differential expression pattern of p63 isoforms by Western blot analysis, using newly developed ΔN and TA specific antibodies. Furthermore using unsupervised clustering of human cell lines, tissues and organs, we show that ΔNp63 and TAp63 driven transcriptional networks involve very distinct sets of molecular players, which may underlie their different biological functions.

Conclusions: In this study we report comprehensive and global expression profiles of p63 isoforms and their relationship to p53/p73 and other potential transcriptional co-regulators. We curate publicly available data generated in part by consortiums such as ENCODE, FANTOM and Human Protein Atlas to delineate the vastly different transcriptomic landscapes of ΔNp63 and TAp63. Our studies help not only in dispelling prevailing myths and controversies on p63 expression in commonly used human cell lines but also augur new isoform- and cell type-specific activities of p63.

No MeSH data available.


Related in: MedlinePlus

Transcription Factors predicted to coordinate with ΔNp63 and TAp63 based on expression profiles (a) Heatmap depicts hierarchical clustering of fold change in expression (over median) of 867 TFs across 40 cell-types (54 experiments). Trimmed dendrogram (correlation > = 0.6) highlights the transcriptional regulators with similar expression pattern to (b) ΔNp63 and (c) TAp63, across a subset of cell-lines (depicted by yellow dotted rectangle). The specific p63 isoforms are shown in red
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Fig3: Transcription Factors predicted to coordinate with ΔNp63 and TAp63 based on expression profiles (a) Heatmap depicts hierarchical clustering of fold change in expression (over median) of 867 TFs across 40 cell-types (54 experiments). Trimmed dendrogram (correlation > = 0.6) highlights the transcriptional regulators with similar expression pattern to (b) ΔNp63 and (c) TAp63, across a subset of cell-lines (depicted by yellow dotted rectangle). The specific p63 isoforms are shown in red

Mentions: Cooperative interactions between TFs constitute an important and indispensable regulatory force that drives tissue-specific gene expression programs. Hence, master regulatory factors such as p63 that play crucial roles in developmental and cell fate decisions are likely to be associated with an expansive repertoire of TFs. Presumably, these p63-linked TFs act in a concerted fashion to modulate the p63 biological output. One obvious corollary to this idea is the possibility that such co-regulators within the p63 network are likely to exhibit similar patterns of expression across different cell types. To this end, we examined expression datasets detailing the relative abundance of a large number of human TFs across 40 normal and cancer cell types. Given the distinct differences in expression patterns of ΔN and TA, we incorporated transcript-specific data information for p63 isoforms in our analysis of the expression dataset. The dataset was also filtered to remove both highly expressed ubiquitous TFs and those with less abundant transcripts (<5 FPKM in at least 1 cell-type). As ΔNp63γ and TAp63 (β, γ) isoforms are expressed at low levels in most of the human cells, they were not represented in this analysis. Unsupervised hierarchical clustering across both genes (867 differentially expressed TFs) and experiments (52 RNA-Seq data sets) revealed quite interesting expression patterns (Fig. 3a). Interestingly, cell-lines appeared to cluster according to the germ layer of origin and their physiological functions, irrespective of the karyotype (normal vs. cancer). Replicates (as represented by MCF7_rep1, MCF7_rep2) and cell-lines that were from identical sources (such as NHEK, DK0) usually clustered together, further ensuring that the method was unbiased to data originating from different sources (Additional file 9: Figure S7).Fig 3


A global analysis of the complex landscape of isoforms and regulatory networks of p63 in human cells and tissues.

Sethi I, Romano RA, Gluck C, Smalley K, Vojtesek B, Buck MJ, Sinha S - BMC Genomics (2015)

Transcription Factors predicted to coordinate with ΔNp63 and TAp63 based on expression profiles (a) Heatmap depicts hierarchical clustering of fold change in expression (over median) of 867 TFs across 40 cell-types (54 experiments). Trimmed dendrogram (correlation > = 0.6) highlights the transcriptional regulators with similar expression pattern to (b) ΔNp63 and (c) TAp63, across a subset of cell-lines (depicted by yellow dotted rectangle). The specific p63 isoforms are shown in red
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4528692&req=5

Fig3: Transcription Factors predicted to coordinate with ΔNp63 and TAp63 based on expression profiles (a) Heatmap depicts hierarchical clustering of fold change in expression (over median) of 867 TFs across 40 cell-types (54 experiments). Trimmed dendrogram (correlation > = 0.6) highlights the transcriptional regulators with similar expression pattern to (b) ΔNp63 and (c) TAp63, across a subset of cell-lines (depicted by yellow dotted rectangle). The specific p63 isoforms are shown in red
Mentions: Cooperative interactions between TFs constitute an important and indispensable regulatory force that drives tissue-specific gene expression programs. Hence, master regulatory factors such as p63 that play crucial roles in developmental and cell fate decisions are likely to be associated with an expansive repertoire of TFs. Presumably, these p63-linked TFs act in a concerted fashion to modulate the p63 biological output. One obvious corollary to this idea is the possibility that such co-regulators within the p63 network are likely to exhibit similar patterns of expression across different cell types. To this end, we examined expression datasets detailing the relative abundance of a large number of human TFs across 40 normal and cancer cell types. Given the distinct differences in expression patterns of ΔN and TA, we incorporated transcript-specific data information for p63 isoforms in our analysis of the expression dataset. The dataset was also filtered to remove both highly expressed ubiquitous TFs and those with less abundant transcripts (<5 FPKM in at least 1 cell-type). As ΔNp63γ and TAp63 (β, γ) isoforms are expressed at low levels in most of the human cells, they were not represented in this analysis. Unsupervised hierarchical clustering across both genes (867 differentially expressed TFs) and experiments (52 RNA-Seq data sets) revealed quite interesting expression patterns (Fig. 3a). Interestingly, cell-lines appeared to cluster according to the germ layer of origin and their physiological functions, irrespective of the karyotype (normal vs. cancer). Replicates (as represented by MCF7_rep1, MCF7_rep2) and cell-lines that were from identical sources (such as NHEK, DK0) usually clustered together, further ensuring that the method was unbiased to data originating from different sources (Additional file 9: Figure S7).Fig 3

Bottom Line: Furthermore using unsupervised clustering of human cell lines, tissues and organs, we show that ΔNp63 and TAp63 driven transcriptional networks involve very distinct sets of molecular players, which may underlie their different biological functions.We curate publicly available data generated in part by consortiums such as ENCODE, FANTOM and Human Protein Atlas to delineate the vastly different transcriptomic landscapes of ΔNp63 and TAp63.Our studies help not only in dispelling prevailing myths and controversies on p63 expression in commonly used human cell lines but also augur new isoform- and cell type-specific activities of p63.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Center of Excellence in Bioinformatics and Life Sciences, State University of New York, 701 Ellicott Street, Buffalo, NY, 14203, USA.

ABSTRACT

Background: The transcription factor p63 belongs to the p53/p63/p73 family and plays key functional roles during normal epithelial development and differentiation and in pathological states such as squamous cell carcinomas. The human TP63 gene, located on chromosome 3q28 is driven by two promoters that generate the full-length transactivating (TA) and N-terminal truncated (ΔN) isoforms. Furthermore alternative splicing at the C-terminus gives rise to additional α, β, γ and likely several other minor variants. Teasing out the expression and biological function of each p63 variant has been both the focus of, and a cause for contention in the p63 field.

Results: Here we have taken advantage of a burgeoning RNA-Seq based genomic data-sets to examine the global expression profiles of p63 isoforms across commonly utilized human cell-lines and major tissues and organs. Consistent with earlier studies, we find ΔNp63 transcripts, primarily that of the ΔNp63α isoforms, to be expressed in most cells of epithelial origin such as those of skin and oral tissues, mammary glands and squamous cell carcinomas. In contrast, TAp63 is not expressed in the majority of normal cell-types and tissues; rather it is selectively expressed at moderate to high levels in a subset of Burkitt's and diffuse large B-cell lymphoma cell lines. We verify this differential expression pattern of p63 isoforms by Western blot analysis, using newly developed ΔN and TA specific antibodies. Furthermore using unsupervised clustering of human cell lines, tissues and organs, we show that ΔNp63 and TAp63 driven transcriptional networks involve very distinct sets of molecular players, which may underlie their different biological functions.

Conclusions: In this study we report comprehensive and global expression profiles of p63 isoforms and their relationship to p53/p73 and other potential transcriptional co-regulators. We curate publicly available data generated in part by consortiums such as ENCODE, FANTOM and Human Protein Atlas to delineate the vastly different transcriptomic landscapes of ΔNp63 and TAp63. Our studies help not only in dispelling prevailing myths and controversies on p63 expression in commonly used human cell lines but also augur new isoform- and cell type-specific activities of p63.

No MeSH data available.


Related in: MedlinePlus