Aberrant smad3 phosphoisoforms in cyst-lining epithelial cells in the CPK mouse, a model of autosomal recessive polycystic kidney disease

Taketsugu Hama, Koichi Nakanishi, Masashi Sato, Hironobu Mukaiyama, Hiroko Togawa, Yuko Shima, Masayasu Miyajima, Kandai Nozu, Shizuko Nagao, Hisahide Takahashi, Mayumi Sako, Kazumoto Iijima, Norishige Yoshikawa, Hiroyuki Suzuki

Research output: Contribution to journalArticle

Abstract

Cystic epithelia acquire mesenchymal-like features in polycystic kidney disease (PKD). In this phenotypic alteration, it is well known that transforming growth factor (TGF)-β/Smad3 signaling is involved; however, there is emerging new data on Smad3 phosphoisoforms: Smad3 phosphorylated at linker regions (pSmad3L), COOH-terminal regions (pSmad3C), and both (pSmad3L/C). pSmad3L/C has a pathological role in colorectal cancer. Mesenchymal phenotype-specific cell responses in the TGF-β/Smad3 pathway are implicated in carcinomas. In this study, we confirmed mesenchymal features and examined Smad3 phosphoisoforms in the cpk mouse, a model of autosomal recessive PKD. Kidney sections were stained with antibodies against mesenchymal markers and domain-specific phospho-Smad3. TGF-β, pSmad3L, pSmad3C, JNK, cyclin-dependent kinase (CDK) 4, and c-Myc were evaluated by Western blotting. Cophosphorylation of pSmad3L/C was assessed by immunoprecipitation. α-Smooth muscle actin, which indicates mesenchymal features, was expressed higher in cpk mice. pSmad3L expression was increased in cpk mice and was predominantly localized in the nuclei of tubular epithelial cells in cysts; however, pSmad3C was equally expressed in both cpk and control mice. Levels of pSmad3L, JNK, CDK4, and c-Myc protein in nuclei were significantly higher in cpk mice than in controls. Immunoprecipitation showed that Smad3 was cophosphorylated (pSmad3L/C) in cpk mice. Smad3 knockout/cpk double-mutant mice revealed amelioration of cpk abnormalities. These findings suggest that upregulating c-Myc through the JNK/CDK4-dependent pSmad3L pathway may be key to the pathophysiology in cpk mice. In conclusion, a qualitative rather than a quantitative abnormality of the TGF-β/Smad3 pathway is involved in PKD and may be a target for disease-specific intervention.

Original languageEnglish
Pages (from-to)F1223-F1231
JournalAmerican Journal of Physiology - Renal Physiology
Volume313
Issue number6
DOIs
Publication statusPublished - 01-12-2017

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Autosomal Recessive Polycystic Kidney
Cysts
Epithelial Cells
Transforming Growth Factors
Polycystic Kidney Diseases
Immunoprecipitation
Cyclin-Dependent Kinase 4
Proto-Oncogene Proteins c-myc
Smooth Muscle
Actins
Colorectal Neoplasms
Epithelium
Western Blotting
Carcinoma
Phenotype
Kidney

All Science Journal Classification (ASJC) codes

  • Physiology
  • Urology

Cite this

Hama, Taketsugu ; Nakanishi, Koichi ; Sato, Masashi ; Mukaiyama, Hironobu ; Togawa, Hiroko ; Shima, Yuko ; Miyajima, Masayasu ; Nozu, Kandai ; Nagao, Shizuko ; Takahashi, Hisahide ; Sako, Mayumi ; Iijima, Kazumoto ; Yoshikawa, Norishige ; Suzuki, Hiroyuki. / Aberrant smad3 phosphoisoforms in cyst-lining epithelial cells in the CPK mouse, a model of autosomal recessive polycystic kidney disease. In: American Journal of Physiology - Renal Physiology. 2017 ; Vol. 313, No. 6. pp. F1223-F1231.
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Hama, T, Nakanishi, K, Sato, M, Mukaiyama, H, Togawa, H, Shima, Y, Miyajima, M, Nozu, K, Nagao, S, Takahashi, H, Sako, M, Iijima, K, Yoshikawa, N & Suzuki, H 2017, 'Aberrant smad3 phosphoisoforms in cyst-lining epithelial cells in the CPK mouse, a model of autosomal recessive polycystic kidney disease', American Journal of Physiology - Renal Physiology, vol. 313, no. 6, pp. F1223-F1231. https://doi.org/10.1152/ajprenal.00697.2016

Aberrant smad3 phosphoisoforms in cyst-lining epithelial cells in the CPK mouse, a model of autosomal recessive polycystic kidney disease. / Hama, Taketsugu; Nakanishi, Koichi; Sato, Masashi; Mukaiyama, Hironobu; Togawa, Hiroko; Shima, Yuko; Miyajima, Masayasu; Nozu, Kandai; Nagao, Shizuko; Takahashi, Hisahide; Sako, Mayumi; Iijima, Kazumoto; Yoshikawa, Norishige; Suzuki, Hiroyuki.

In: American Journal of Physiology - Renal Physiology, Vol. 313, No. 6, 01.12.2017, p. F1223-F1231.

Research output: Contribution to journalArticle

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T1 - Aberrant smad3 phosphoisoforms in cyst-lining epithelial cells in the CPK mouse, a model of autosomal recessive polycystic kidney disease

AU - Hama, Taketsugu

AU - Nakanishi, Koichi

AU - Sato, Masashi

AU - Mukaiyama, Hironobu

AU - Togawa, Hiroko

AU - Shima, Yuko

AU - Miyajima, Masayasu

AU - Nozu, Kandai

AU - Nagao, Shizuko

AU - Takahashi, Hisahide

AU - Sako, Mayumi

AU - Iijima, Kazumoto

AU - Yoshikawa, Norishige

AU - Suzuki, Hiroyuki

PY - 2017/12/1

Y1 - 2017/12/1

N2 - Cystic epithelia acquire mesenchymal-like features in polycystic kidney disease (PKD). In this phenotypic alteration, it is well known that transforming growth factor (TGF)-β/Smad3 signaling is involved; however, there is emerging new data on Smad3 phosphoisoforms: Smad3 phosphorylated at linker regions (pSmad3L), COOH-terminal regions (pSmad3C), and both (pSmad3L/C). pSmad3L/C has a pathological role in colorectal cancer. Mesenchymal phenotype-specific cell responses in the TGF-β/Smad3 pathway are implicated in carcinomas. In this study, we confirmed mesenchymal features and examined Smad3 phosphoisoforms in the cpk mouse, a model of autosomal recessive PKD. Kidney sections were stained with antibodies against mesenchymal markers and domain-specific phospho-Smad3. TGF-β, pSmad3L, pSmad3C, JNK, cyclin-dependent kinase (CDK) 4, and c-Myc were evaluated by Western blotting. Cophosphorylation of pSmad3L/C was assessed by immunoprecipitation. α-Smooth muscle actin, which indicates mesenchymal features, was expressed higher in cpk mice. pSmad3L expression was increased in cpk mice and was predominantly localized in the nuclei of tubular epithelial cells in cysts; however, pSmad3C was equally expressed in both cpk and control mice. Levels of pSmad3L, JNK, CDK4, and c-Myc protein in nuclei were significantly higher in cpk mice than in controls. Immunoprecipitation showed that Smad3 was cophosphorylated (pSmad3L/C) in cpk mice. Smad3 knockout/cpk double-mutant mice revealed amelioration of cpk abnormalities. These findings suggest that upregulating c-Myc through the JNK/CDK4-dependent pSmad3L pathway may be key to the pathophysiology in cpk mice. In conclusion, a qualitative rather than a quantitative abnormality of the TGF-β/Smad3 pathway is involved in PKD and may be a target for disease-specific intervention.

AB - Cystic epithelia acquire mesenchymal-like features in polycystic kidney disease (PKD). In this phenotypic alteration, it is well known that transforming growth factor (TGF)-β/Smad3 signaling is involved; however, there is emerging new data on Smad3 phosphoisoforms: Smad3 phosphorylated at linker regions (pSmad3L), COOH-terminal regions (pSmad3C), and both (pSmad3L/C). pSmad3L/C has a pathological role in colorectal cancer. Mesenchymal phenotype-specific cell responses in the TGF-β/Smad3 pathway are implicated in carcinomas. In this study, we confirmed mesenchymal features and examined Smad3 phosphoisoforms in the cpk mouse, a model of autosomal recessive PKD. Kidney sections were stained with antibodies against mesenchymal markers and domain-specific phospho-Smad3. TGF-β, pSmad3L, pSmad3C, JNK, cyclin-dependent kinase (CDK) 4, and c-Myc were evaluated by Western blotting. Cophosphorylation of pSmad3L/C was assessed by immunoprecipitation. α-Smooth muscle actin, which indicates mesenchymal features, was expressed higher in cpk mice. pSmad3L expression was increased in cpk mice and was predominantly localized in the nuclei of tubular epithelial cells in cysts; however, pSmad3C was equally expressed in both cpk and control mice. Levels of pSmad3L, JNK, CDK4, and c-Myc protein in nuclei were significantly higher in cpk mice than in controls. Immunoprecipitation showed that Smad3 was cophosphorylated (pSmad3L/C) in cpk mice. Smad3 knockout/cpk double-mutant mice revealed amelioration of cpk abnormalities. These findings suggest that upregulating c-Myc through the JNK/CDK4-dependent pSmad3L pathway may be key to the pathophysiology in cpk mice. In conclusion, a qualitative rather than a quantitative abnormality of the TGF-β/Smad3 pathway is involved in PKD and may be a target for disease-specific intervention.

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