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Identification of novel genes selectively expressed in the follicle-associated epithelium from the meta-analysis of transcriptomics data from multiple mouse cell and tissue populations.

Kobayashi A, Donaldson DS, Kanaya T, Fukuda S, Baillie JK, Freeman TC, Ohno H, Williams IR, Mabbott NA - DNA Res. (2012)

Bottom Line: The follicle-associated epithelium (FAE) overlying the Peyer's patches and the microfold cells (M cells) within it are important sites of antigen transcytosis across the intestinal epithelium.This study provides new insight into the FAE transcriptome.Further characterization of the candidate genes identified here will aid the identification of novel regulators of cell function in the FAE.

View Article: PubMed Central - PubMed

Affiliation: The Roslin Institute and Royal (Dick) School of Veterinary Sciences, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK.

ABSTRACT
The follicle-associated epithelium (FAE) overlying the Peyer's patches and the microfold cells (M cells) within it are important sites of antigen transcytosis across the intestinal epithelium. Using a meta-analysis approach, we identified a transcriptional signature that distinguished the FAE from a large collection of mouse cells and tissues. A co-expressed cluster of 21 FAE-specific genes was identified, and the analysis of the transcription factor binding site motifs in their promoter regions indicated that these genes shared an underlying transcriptional programme. This cluster contained known FAE- (Anxa10, Ccl20, Psg18 and Ubd) and M-cell-specific (Gp2) genes, suggesting that the others were novel FAE-specific genes. Some of these novel candidate genes were expressed highly by the FAE and M cells (Calcb, Ces3b, Clca2 and Gjb2), and others only by the FAE (Ascl2, Cftr, Fgf15, Gpr133, Kcna1, Kcnj15, Mycl1, Pgap1 and Rps6kl). We also identified a subset of novel FAE-related genes that were induced in the intestinal epithelium after receptor activator of nuclear factor (NF)-κB ligand stimulation. These included Mfge8 which was specific to FAE enterocytes. This study provides new insight into the FAE transcriptome. Further characterization of the candidate genes identified here will aid the identification of novel regulators of cell function in the FAE.

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KCNJ15 is expressed throughout the FAE, but not by M cells or the villous epithelium. (A) The Affymetrix probe sets registered for Kcnj15 which were co-expressed within cluster 65 were surveyed with the RefDIC probe-mapping tool to confirm that they recognized exons within this gene (red bars). NM_019664 and NM_001039056 accession numbers of reference sequences. (B) Heat map showing the expression of profile throughout the intestine of probe sets encoding KCNJ potassium channels (a representative probe set is shown when multiple probe sets for a gene were present on the array). Each column represents the mean probe set intensity (log2) for all samples from each source (n = 2–4). Kcnj15 was the only gene which was expressed significantly within the FAE at levels >2.0-fold when compared with the villous epithelium. The red-boxed area indicates the mAb NKM 16-2-4-enriched Peyer's patch M cell data sets; the blue-boxed area indicates the FAE data sets. (C) IHC analysis of mouse Peyer's patches shows the cytoplasmic expression of KCNJ15 (red) throughout cells within the FAE, but not in M cells (GP2+ cells, green, arrows) or the villous epithelium. SED, subepithelial dome. Dotted line indicates the border of the FAE.
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DSS022F5: KCNJ15 is expressed throughout the FAE, but not by M cells or the villous epithelium. (A) The Affymetrix probe sets registered for Kcnj15 which were co-expressed within cluster 65 were surveyed with the RefDIC probe-mapping tool to confirm that they recognized exons within this gene (red bars). NM_019664 and NM_001039056 accession numbers of reference sequences. (B) Heat map showing the expression of profile throughout the intestine of probe sets encoding KCNJ potassium channels (a representative probe set is shown when multiple probe sets for a gene were present on the array). Each column represents the mean probe set intensity (log2) for all samples from each source (n = 2–4). Kcnj15 was the only gene which was expressed significantly within the FAE at levels >2.0-fold when compared with the villous epithelium. The red-boxed area indicates the mAb NKM 16-2-4-enriched Peyer's patch M cell data sets; the blue-boxed area indicates the FAE data sets. (C) IHC analysis of mouse Peyer's patches shows the cytoplasmic expression of KCNJ15 (red) throughout cells within the FAE, but not in M cells (GP2+ cells, green, arrows) or the villous epithelium. SED, subepithelial dome. Dotted line indicates the border of the FAE.

Mentions: Among the genes in cluster 65 that had not previously been reported to be expressed within the FAE was Kcnj15 which encodes for potassium inwardly rectifying channel, subfamily J, member 15 (KCNJ15). The specificity of the Kcnj15 probe sets was confirmed using the RefDIC probe mapping tool (http://refdic.rcai.riken.jp/tools/id_on_map.cgi) which predicts whether or not each probe set recognizes coding regions of target genes17 (Fig. 5A). A large diversity of potassium channels has been described. However, in the intestine, Kcnj15 was the only subfamily J potassium channel-encoding gene represented on the microarray that was significantly and specifically expressed only within the FAE when compared with the villous epithelium samples (Fig. 5B). Immunohistochemistry (IHC) analysis demonstrated that KCNJ15 was distributed within the cytoplasm of most cells throughout the FAE, but appeared to be absent in GP2+ M cells (Fig. 5C). Consistent with the microarray data, the villous epithelium lacked the expression of KCNJ15. These data support the conclusion that the genes within cluster 65 are expressed highly by cells in the FAE.Figure 5.


Identification of novel genes selectively expressed in the follicle-associated epithelium from the meta-analysis of transcriptomics data from multiple mouse cell and tissue populations.

Kobayashi A, Donaldson DS, Kanaya T, Fukuda S, Baillie JK, Freeman TC, Ohno H, Williams IR, Mabbott NA - DNA Res. (2012)

KCNJ15 is expressed throughout the FAE, but not by M cells or the villous epithelium. (A) The Affymetrix probe sets registered for Kcnj15 which were co-expressed within cluster 65 were surveyed with the RefDIC probe-mapping tool to confirm that they recognized exons within this gene (red bars). NM_019664 and NM_001039056 accession numbers of reference sequences. (B) Heat map showing the expression of profile throughout the intestine of probe sets encoding KCNJ potassium channels (a representative probe set is shown when multiple probe sets for a gene were present on the array). Each column represents the mean probe set intensity (log2) for all samples from each source (n = 2–4). Kcnj15 was the only gene which was expressed significantly within the FAE at levels >2.0-fold when compared with the villous epithelium. The red-boxed area indicates the mAb NKM 16-2-4-enriched Peyer's patch M cell data sets; the blue-boxed area indicates the FAE data sets. (C) IHC analysis of mouse Peyer's patches shows the cytoplasmic expression of KCNJ15 (red) throughout cells within the FAE, but not in M cells (GP2+ cells, green, arrows) or the villous epithelium. SED, subepithelial dome. Dotted line indicates the border of the FAE.
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Related In: Results  -  Collection

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DSS022F5: KCNJ15 is expressed throughout the FAE, but not by M cells or the villous epithelium. (A) The Affymetrix probe sets registered for Kcnj15 which were co-expressed within cluster 65 were surveyed with the RefDIC probe-mapping tool to confirm that they recognized exons within this gene (red bars). NM_019664 and NM_001039056 accession numbers of reference sequences. (B) Heat map showing the expression of profile throughout the intestine of probe sets encoding KCNJ potassium channels (a representative probe set is shown when multiple probe sets for a gene were present on the array). Each column represents the mean probe set intensity (log2) for all samples from each source (n = 2–4). Kcnj15 was the only gene which was expressed significantly within the FAE at levels >2.0-fold when compared with the villous epithelium. The red-boxed area indicates the mAb NKM 16-2-4-enriched Peyer's patch M cell data sets; the blue-boxed area indicates the FAE data sets. (C) IHC analysis of mouse Peyer's patches shows the cytoplasmic expression of KCNJ15 (red) throughout cells within the FAE, but not in M cells (GP2+ cells, green, arrows) or the villous epithelium. SED, subepithelial dome. Dotted line indicates the border of the FAE.
Mentions: Among the genes in cluster 65 that had not previously been reported to be expressed within the FAE was Kcnj15 which encodes for potassium inwardly rectifying channel, subfamily J, member 15 (KCNJ15). The specificity of the Kcnj15 probe sets was confirmed using the RefDIC probe mapping tool (http://refdic.rcai.riken.jp/tools/id_on_map.cgi) which predicts whether or not each probe set recognizes coding regions of target genes17 (Fig. 5A). A large diversity of potassium channels has been described. However, in the intestine, Kcnj15 was the only subfamily J potassium channel-encoding gene represented on the microarray that was significantly and specifically expressed only within the FAE when compared with the villous epithelium samples (Fig. 5B). Immunohistochemistry (IHC) analysis demonstrated that KCNJ15 was distributed within the cytoplasm of most cells throughout the FAE, but appeared to be absent in GP2+ M cells (Fig. 5C). Consistent with the microarray data, the villous epithelium lacked the expression of KCNJ15. These data support the conclusion that the genes within cluster 65 are expressed highly by cells in the FAE.Figure 5.

Bottom Line: The follicle-associated epithelium (FAE) overlying the Peyer's patches and the microfold cells (M cells) within it are important sites of antigen transcytosis across the intestinal epithelium.This study provides new insight into the FAE transcriptome.Further characterization of the candidate genes identified here will aid the identification of novel regulators of cell function in the FAE.

View Article: PubMed Central - PubMed

Affiliation: The Roslin Institute and Royal (Dick) School of Veterinary Sciences, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK.

ABSTRACT
The follicle-associated epithelium (FAE) overlying the Peyer's patches and the microfold cells (M cells) within it are important sites of antigen transcytosis across the intestinal epithelium. Using a meta-analysis approach, we identified a transcriptional signature that distinguished the FAE from a large collection of mouse cells and tissues. A co-expressed cluster of 21 FAE-specific genes was identified, and the analysis of the transcription factor binding site motifs in their promoter regions indicated that these genes shared an underlying transcriptional programme. This cluster contained known FAE- (Anxa10, Ccl20, Psg18 and Ubd) and M-cell-specific (Gp2) genes, suggesting that the others were novel FAE-specific genes. Some of these novel candidate genes were expressed highly by the FAE and M cells (Calcb, Ces3b, Clca2 and Gjb2), and others only by the FAE (Ascl2, Cftr, Fgf15, Gpr133, Kcna1, Kcnj15, Mycl1, Pgap1 and Rps6kl). We also identified a subset of novel FAE-related genes that were induced in the intestinal epithelium after receptor activator of nuclear factor (NF)-κB ligand stimulation. These included Mfge8 which was specific to FAE enterocytes. This study provides new insight into the FAE transcriptome. Further characterization of the candidate genes identified here will aid the identification of novel regulators of cell function in the FAE.

Show MeSH
Related in: MedlinePlus