Immunohistochemical and immunoblotting identification of protein phosphatase 1γ1 in rat salivary glands

Shirakawa, Satoko; Mochizuki, Hajime; Kobayashi, Shigeru; Takehara, Tadamichi; Shima, Hiroshi; Nagao, Minako; Haneji, Tatsuji

doi:10.1016/0014-5793(96)00821-6

Cited by 9 publications

(7 citation statements)

References 20 publications

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“…These findings suggest that protein phosphatases may be significant factors in the function and regulation of salivary glands, including normal growth and repair processes, pathogenesis of SS, or apoptosis. We also have reported that the ␥ isotype of PP-1 catalytic subunit is expressed in rat salivary glands (Shirakawa et al, 1996). Although detailed future analysis is needed, our present results suggest that hyper-phosphorylation induces apoptosis in HSG salivary gland cells.…”

Section: Discussionsupporting

confidence: 58%

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The protein phosphatase inhibitors, okadaic acid and calyculin A, induce apoptosis in human submandibular gland ductal cell line HSG cells

Morimoto

Kobayashi

et al. 1999

Oral Diseases

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OBJECTIVE: To investigate a possible relationship between protein phosphorylation or dephosphorylation status and apoptosis in salivary gland cells, we examined the effects of okadaic acid and calyculin A, the protein phosphatase inhibitors, on cultured human submandibular gland ductal cell line, HSG cells. METHODS: HSG cells at subconfluent stages were exposed to varying concentrations of okadaic acid or calyculin A.Apoptoses were analysed in HSG cells by phase‐contrast microscopy, WST‐1 cytotoxicity assay, Hoechst 33342 staining, and DNA ladder formation. RESULT: Both okadaic acid and calyculin A induced cell death in HSG cells in a dose‐dependent fashion. Marked nuclear condensation and fragmentation of chromatin was observed in HSG cellS. DNA ladder formation was also detected in HSG cells by treatment with okadaic acid or calyculin A. The induced DNA ladder formation was dose‐dependent with maximal effect at concentrations of 50 nM okadaic acid and 2 nM calyculin A, respectively, and were time‐dependent from 14 h to 48 h. To further determine if new gene transcription and protein synthesis regulate okadaic acid‐induced apoptosis in HSG cells, the cells were treated with cycloheximide or actinomycin D in the presence of 20 nM okadaic acid. Neither inhibitor protected the cells against okadaic acid‐induced apoptosis. CONCLUSION: Based on the known selectivity of oka‐daic acid and calyculin A, our results indicate that the pathway of the apoptosis in the cultured salivary gland cells is regulated by protein phosphatase type 1 or type 2A.Our results also suggest that new protein synthesis and/or mRNA expression are not involved in okadaic acid‐induced apoptosis in HSG cells.

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Section: Discussionsupporting

confidence: 58%

“…In a previous study, we demonstrated that the ␥1 isotype of PP-1 catalytic subunit is distributed in rat salivary glands (Shirakawa et al, 1996). It has also been reported that OA inhibited amylase exocytosis in rat salivary acini (Takuma et al, 1993).…”

Section: Introductionmentioning

confidence: 66%

The protein phosphatase inhibitors, okadaic acid and calyculin A, induce apoptosis in human submandibular gland ductal cell line HSG cells

Morimoto

Kobayashi

et al. 1999

Oral Diseases

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“…The expression of PP1s was investigated at the mRNA and protein levels in various tissues and organs [21,[56][57][58]. We examined the subcellular localization of PP1 isotypes in mouse osteoblastic MC3T3-E1 cells [20].…”

Section: Subcellular Localization Of Pp1 Isozymesmentioning

confidence: 99%

Association of Protein Phosphatase 1 Delta with Nucleolin in Osteoblastic Cells and Cleavage of Nucleolin in Apoptosis-induced Osteoblastic Cells

Haneji

2005

Acta Histochem. Cytochem

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Protein phosphorylation and dephosphorylation has been recognized as a key mechanism in cell function. Okadaic acid is a potent inhibitor of PP1 and PP 2A and induces apoptosis in human or mouse cells including osteoblasts. Nucleolin is an abundantly expressed nucleolar phosphoprotein and is located mainly in the nucleolus. The staining pattern of nucleolin in cultured osteoblastic cells is similar to that of PP1d. Nucleolin was demonstrated to bind to PP1d in nucleolus in human osteoblastic cells by using immunocytochemical and immunoprecipitation methods. AgNORs and nucleolin, visible as dots in the nuclei of the control osteoblastic cells, disappeared from the nuclei of the osteoblastic cells treated with okadaic acid. A major band, 110 kDa, was detected in the proteins obtained from osteoblastic cells. The level of the 110 kDa protein decreased in the apoptotic osteoblastic cells, whereas an additional band, 80 kDa, appeared and the level of this protein increased in the proteins prepared from okadaic acid-induced apoptotic osteoblastic cells. Our results indicate that PP1d directly binds to nucleolin in the nucleolus of osteoblastic cells and that nucleolin is cleaved during apoptosis in osteoblastic cells.

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“…Immunocytochemistry was also carried out in rat cerebellum and salivary glands for PP1γ1 [8, 27] and in testis for PP1γ2 [26]. In previous studies, we reported the subcellular localization of PP1 isoforms in mouse osteoblastic MC3T3-E1 cells and human osteoblastic MG63 cells [6, 17].…”

Section: Introductionmentioning

confidence: 99%

Interaction of Protein Phosphatase 1δ with Nucleophosmin in Human Osteoblastic Cells

Haneji

Teramachi

Hirashima

et al. 2012

Acta Histochem. Cytochem

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Protein phosphorylation and dephosphorylation has been recognized as an essential mechanism in the regulation of cellular metabolism and function in various tissues. Serine and threonine protein phosphatases (PP) are divided into four categories: PP1, PP2A, PP2B, and PP2C. At least four isoforms of PP1 catalytic subunit in rat, PP1α, PP1γ1, PP1γ2, and PP1δ, were isolated. In the present study, we examined the localization and expression of PP1δ in human osteoblastic Saos-2 cells. Anti-PP1δ antibody recognized a protein present in the nucleolar regions in Saos-2 cells. Cellular fractionation revealed that PP1δ is a 37 kDa protein localized in the nucleolus. Nucleophosmin is a nucleolar phosphoprotein and located mainly in the nucleolus. Staining pattern of nucleophosmin in Saos-2 cells was similar to that of PP1δ. PP1δ and nucleophosmin were specifically stained as dots in the nucleus. Dual fluorescence images revealed that PP1δ and nucleophosmin were localized in the same regions in the nucleolus. Similar distribution patterns of PP1δ and nucleophosmin were observed in osteoblastic MG63 cells. The interaction of PP1δ and nucleophosmin was also shown by immunoprecipitation and Western analysis. These results indicated that PP1δ associate with nucleophosmin directly in the nucleolus and suggested that nucleophosmin is one of the candidate substrate for PP1δ.

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Immunohistochemical and immunoblotting identification of protein phosphatase 1γ1 in rat salivary glands

Cited by 9 publications

References 20 publications

The protein phosphatase inhibitors, okadaic acid and calyculin A, induce apoptosis in human submandibular gland ductal cell line HSG cells

The protein phosphatase inhibitors, okadaic acid and calyculin A, induce apoptosis in human submandibular gland ductal cell line HSG cells

Association of Protein Phosphatase 1 Delta with Nucleolin in Osteoblastic Cells and Cleavage of Nucleolin in Apoptosis-induced Osteoblastic Cells

Interaction of Protein Phosphatase 1δ with Nucleophosmin in Human Osteoblastic Cells

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