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Identification of KRAP-expressing cells and the functional relevance of KRAP to the subcellular localization of IP3R in the stomach and kidney.

Fujimoto T, Shirasawa S - Int. J. Mol. Med. (2012)

Bottom Line: We have recently found that KRAP is a molecule associated with inositol 1,4,5-trisphosphate receptor (IP(3)R) and is critical for the proper subcellular localization of IP(3)R in the liver and the pancreas.Finally, co-immunoprecipitation study in the stomachs and the kidneys validated the physical association of KRAP with IP(3)Rs.These findings demonstrate that KRAP physically associates with IP(3)Rs and regulates the proper localization of IP(3)Rs in the mucous cells and the chief cells of the stomach and in the proximal tubular cells of the kidneys.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka 814-0180, Japan.

ABSTRACT
KRAS-induced actin-interacting protein (KRAP), originally identified as one of the deregulated genes expressed in colorectal cancer, participates under physiological conditions in the regulation of systemic energy homeostasis and of the exocrine system. We have recently found that KRAP is a molecule associated with inositol 1,4,5-trisphosphate receptor (IP(3)R) and is critical for the proper subcellular localization of IP(3)R in the liver and the pancreas. However, the expression of KRAP and its precise function in other tissues remain elusive. In this study, we aimed to identify the KRAP-expressing cells in mouse stomach and kidneys and to examine the relevance of KRAP expression in the regulation of IP(3)R localization in these tissues. In the stomach, double immunohistochemical staining for KRAP and IP(3)R demonstrated that KRAP was expressed along with the apical regions in the mucous cells and the chief cells, and IP(3)R3 was dominantly co-localized with KRAP in these cells. Furthermore, IP(3)R2 was also co-localized with IP(3)R3 in the chief cells. It is of note that the proper localization of IP(3)R3 and IP(3)R2 in the chief cells and of IP(3)R3 in the mucous cells were significantly abrogated in KRAP-deficient mice. In the kidneys, KRAP was expressed in both the apical and the basal regions of the proximal tubular cells. Intriguingly, KRAP deficiency abrogated the localization of IP(3)R1 in the proximal tubular cells. Finally, co-immunoprecipitation study in the stomachs and the kidneys validated the physical association of KRAP with IP(3)Rs. These findings demonstrate that KRAP physically associates with IP(3)Rs and regulates the proper localization of IP(3)Rs in the mucous cells and the chief cells of the stomach and in the proximal tubular cells of the kidneys.

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Impaired localization of IP3Rs in the KRAP-deficient chiefcells and the mucous cells. (A and B) Fluorescent confocal images of the base region ofgastric glands for IP3R3 (red), F-actin with phalloidin (green), and themerged photo from wild-type (WT) (A) or KRAP-deficient (KO) (B) mice.Asterisks and arrows indicate the parietal cells and the apical membranes of the chiefcells, respectively. (C and D) Fluorescent confocal images of the pit region of gastricglands for IP3R3 (red), F-actin (green), and the merged photo from WT (C) orKO (D) mice. Arrows indicate the apical membranes of the mucous cells. (E and F)Fluorescent confocal images of the base region of gastric glands for IP3R2(red), F-actin (green), and the merged photo from WT (E) or KO (F) mice. Asterisks andarrows indicate the parietal cells and the apical membranes of the chief cells,respectively. Blue, 4′,6-diamidino-2-phenylindole (DAPI) staining; scale bar, 25μm.
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f3-ijmm-30-06-1287: Impaired localization of IP3Rs in the KRAP-deficient chiefcells and the mucous cells. (A and B) Fluorescent confocal images of the base region ofgastric glands for IP3R3 (red), F-actin with phalloidin (green), and themerged photo from wild-type (WT) (A) or KRAP-deficient (KO) (B) mice.Asterisks and arrows indicate the parietal cells and the apical membranes of the chiefcells, respectively. (C and D) Fluorescent confocal images of the pit region of gastricglands for IP3R3 (red), F-actin (green), and the merged photo from WT (C) orKO (D) mice. Arrows indicate the apical membranes of the mucous cells. (E and F)Fluorescent confocal images of the base region of gastric glands for IP3R2(red), F-actin (green), and the merged photo from WT (E) or KO (F) mice. Asterisks andarrows indicate the parietal cells and the apical membranes of the chief cells,respectively. Blue, 4′,6-diamidino-2-phenylindole (DAPI) staining; scale bar, 25μm.

Mentions: We addressed the functional relevance of KRAP to the proper localization ofIP3R by using KRAP-KO mice. IP3R3 was located inthe apical region of the chief cells (Fig.3A, arrow) and of the mucous cells (Fig. 3C, arrows) in the wild-type (WT) mouse stomach, whereas the restrictedlocalization of IP3R3 appeared to be diminished in the KRAP-KOstomach (Fig. 3B, arrow; 3D, arrows).Furthermore, IP3R2 was detected in both the chief cells (Fig. 3E, arrows) and the parietal cells(Fig. 3E, asterisks) in the WTstomach, whereas the localization of IP3R2 in the KRAP-KOstomach was impaired in the chief cells (Fig. 3F, arrows) but not in the parietal cells (Fig. 3F, asterisks). Thus, KRAP plays critical role in theregulation of the proper localization of IP3R2 and IP3R3 in thechief cells and of IP3R3 in the mucous cells.


Identification of KRAP-expressing cells and the functional relevance of KRAP to the subcellular localization of IP3R in the stomach and kidney.

Fujimoto T, Shirasawa S - Int. J. Mol. Med. (2012)

Impaired localization of IP3Rs in the KRAP-deficient chiefcells and the mucous cells. (A and B) Fluorescent confocal images of the base region ofgastric glands for IP3R3 (red), F-actin with phalloidin (green), and themerged photo from wild-type (WT) (A) or KRAP-deficient (KO) (B) mice.Asterisks and arrows indicate the parietal cells and the apical membranes of the chiefcells, respectively. (C and D) Fluorescent confocal images of the pit region of gastricglands for IP3R3 (red), F-actin (green), and the merged photo from WT (C) orKO (D) mice. Arrows indicate the apical membranes of the mucous cells. (E and F)Fluorescent confocal images of the base region of gastric glands for IP3R2(red), F-actin (green), and the merged photo from WT (E) or KO (F) mice. Asterisks andarrows indicate the parietal cells and the apical membranes of the chief cells,respectively. Blue, 4′,6-diamidino-2-phenylindole (DAPI) staining; scale bar, 25μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4042864&req=5

f3-ijmm-30-06-1287: Impaired localization of IP3Rs in the KRAP-deficient chiefcells and the mucous cells. (A and B) Fluorescent confocal images of the base region ofgastric glands for IP3R3 (red), F-actin with phalloidin (green), and themerged photo from wild-type (WT) (A) or KRAP-deficient (KO) (B) mice.Asterisks and arrows indicate the parietal cells and the apical membranes of the chiefcells, respectively. (C and D) Fluorescent confocal images of the pit region of gastricglands for IP3R3 (red), F-actin (green), and the merged photo from WT (C) orKO (D) mice. Arrows indicate the apical membranes of the mucous cells. (E and F)Fluorescent confocal images of the base region of gastric glands for IP3R2(red), F-actin (green), and the merged photo from WT (E) or KO (F) mice. Asterisks andarrows indicate the parietal cells and the apical membranes of the chief cells,respectively. Blue, 4′,6-diamidino-2-phenylindole (DAPI) staining; scale bar, 25μm.
Mentions: We addressed the functional relevance of KRAP to the proper localization ofIP3R by using KRAP-KO mice. IP3R3 was located inthe apical region of the chief cells (Fig.3A, arrow) and of the mucous cells (Fig. 3C, arrows) in the wild-type (WT) mouse stomach, whereas the restrictedlocalization of IP3R3 appeared to be diminished in the KRAP-KOstomach (Fig. 3B, arrow; 3D, arrows).Furthermore, IP3R2 was detected in both the chief cells (Fig. 3E, arrows) and the parietal cells(Fig. 3E, asterisks) in the WTstomach, whereas the localization of IP3R2 in the KRAP-KOstomach was impaired in the chief cells (Fig. 3F, arrows) but not in the parietal cells (Fig. 3F, asterisks). Thus, KRAP plays critical role in theregulation of the proper localization of IP3R2 and IP3R3 in thechief cells and of IP3R3 in the mucous cells.

Bottom Line: We have recently found that KRAP is a molecule associated with inositol 1,4,5-trisphosphate receptor (IP(3)R) and is critical for the proper subcellular localization of IP(3)R in the liver and the pancreas.Finally, co-immunoprecipitation study in the stomachs and the kidneys validated the physical association of KRAP with IP(3)Rs.These findings demonstrate that KRAP physically associates with IP(3)Rs and regulates the proper localization of IP(3)Rs in the mucous cells and the chief cells of the stomach and in the proximal tubular cells of the kidneys.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Faculty of Medicine, Fukuoka University, Fukuoka 814-0180, Japan.

ABSTRACT
KRAS-induced actin-interacting protein (KRAP), originally identified as one of the deregulated genes expressed in colorectal cancer, participates under physiological conditions in the regulation of systemic energy homeostasis and of the exocrine system. We have recently found that KRAP is a molecule associated with inositol 1,4,5-trisphosphate receptor (IP(3)R) and is critical for the proper subcellular localization of IP(3)R in the liver and the pancreas. However, the expression of KRAP and its precise function in other tissues remain elusive. In this study, we aimed to identify the KRAP-expressing cells in mouse stomach and kidneys and to examine the relevance of KRAP expression in the regulation of IP(3)R localization in these tissues. In the stomach, double immunohistochemical staining for KRAP and IP(3)R demonstrated that KRAP was expressed along with the apical regions in the mucous cells and the chief cells, and IP(3)R3 was dominantly co-localized with KRAP in these cells. Furthermore, IP(3)R2 was also co-localized with IP(3)R3 in the chief cells. It is of note that the proper localization of IP(3)R3 and IP(3)R2 in the chief cells and of IP(3)R3 in the mucous cells were significantly abrogated in KRAP-deficient mice. In the kidneys, KRAP was expressed in both the apical and the basal regions of the proximal tubular cells. Intriguingly, KRAP deficiency abrogated the localization of IP(3)R1 in the proximal tubular cells. Finally, co-immunoprecipitation study in the stomachs and the kidneys validated the physical association of KRAP with IP(3)Rs. These findings demonstrate that KRAP physically associates with IP(3)Rs and regulates the proper localization of IP(3)Rs in the mucous cells and the chief cells of the stomach and in the proximal tubular cells of the kidneys.

Show MeSH
Related in: MedlinePlus