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Functional differences between wild-type and mutant-type BRCA1-associated protein 1 tumor suppressor against malignant mesothelioma cells.

Hakiri S, Osada H, Ishiguro F, Murakami H, Murakami-Tonami Y, Yokoi K, Sekido Y - Cancer Sci. (2015)

Bottom Line: Transduction of the WT BAP1 vector into MM cells with homozygous deletion at the BAP1 3' side resulted in both inhibition of cell proliferation and anchorage-independent cell growth, whereas BAP1 mutants of a missense or C-terminal truncated form showed impaired growth inhibitory effects.Furthermore, using the MM cells with BAP1 deletion, we found that WT BAP1, and even a missense mutant, conferred a higher survival rate after IR compared to the control vector.Our results suggested that, whereas WT BAP1 suppresses MM cell proliferation and restores cell survival after IR damage, some mutant BAP1 may also moderately retain these functions.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Oncology, Aichi Cancer Center Research Institute, Nagoya, Japan.

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BRCA1-associated protein 1 (BAP1) gene mutations in malignant mesothelioma (MM). (a) Schematic diagram of BAP1 mutations in MM cell lines. (b) Schematic diagram of an insertion mutation due to G to T change at the acceptor site of exon 8 in Y-MESO-61. (c) Genomic PCR analysis of exons of BAP1 detected homozygous deletion (HD) in Y-MESO-25 cells (arrowhead). (d) Western blot analysis of BAP1. Expression of β-actin was used as the control. (e) Summary of five gene mutation statuses in MM cell lines. Red boxes indicate inactivating mutation or HD. The mutation statuses of neurofibromatosis type 2 (NF2), large tumor suppressor homolog 2 (LATS2), Salvador homolog 1 (SAV1), and cyclin-dependent kinase inhibitor 2A (CDKN2A) were previously reported.11 NLS, nuclear localization signal; UCH, ubiquitin COOH-terminal hydrolase.
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fig01: BRCA1-associated protein 1 (BAP1) gene mutations in malignant mesothelioma (MM). (a) Schematic diagram of BAP1 mutations in MM cell lines. (b) Schematic diagram of an insertion mutation due to G to T change at the acceptor site of exon 8 in Y-MESO-61. (c) Genomic PCR analysis of exons of BAP1 detected homozygous deletion (HD) in Y-MESO-25 cells (arrowhead). (d) Western blot analysis of BAP1. Expression of β-actin was used as the control. (e) Summary of five gene mutation statuses in MM cell lines. Red boxes indicate inactivating mutation or HD. The mutation statuses of neurofibromatosis type 2 (NF2), large tumor suppressor homolog 2 (LATS2), Salvador homolog 1 (SAV1), and cyclin-dependent kinase inhibitor 2A (CDKN2A) were previously reported.11 NLS, nuclear localization signal; UCH, ubiquitin COOH-terminal hydrolase.

Mentions: We carried out mutational analyses of BAP1 using 25 MM cell lines. Among 19 cell lines established from Japanese patients, four non-synonymous or insertion/deletion mutations were found; a nonsense mutation in ACC-MESO-4, two truncating mutations in Y-MESO-9 and Y-MESO-14, and a 40-bp insertion mutation at the intron 7–exon 8 junction in Y-MESO-61 (Fig.1a,b). We also carried out sequence analysis of cDNA synthesized from these cell line RNAs and confirmed the expression of the mutant RNAs (data not shown). As our previous study11 using array comparative genomic hybridization analysis detected a possible homozygous deletion (HD) in Y-MESO-25 (Fig. S1a), we carried out a genomic PCR analysis and confirmed that this cell line has HD of BAP1 at exons 13–17 (Fig.1c). In total, we found BAP1 mutations in 26% (5/19) of the Japanese MM cell lines (Table1). In addition, of six cell lines, the mutation status of which has been previously reported by another group,14 we confirmed the same mutations in two cell lines (NCI-H28 and NCI-H2452) and in the WT among four cell lines (Table1).


Functional differences between wild-type and mutant-type BRCA1-associated protein 1 tumor suppressor against malignant mesothelioma cells.

Hakiri S, Osada H, Ishiguro F, Murakami H, Murakami-Tonami Y, Yokoi K, Sekido Y - Cancer Sci. (2015)

BRCA1-associated protein 1 (BAP1) gene mutations in malignant mesothelioma (MM). (a) Schematic diagram of BAP1 mutations in MM cell lines. (b) Schematic diagram of an insertion mutation due to G to T change at the acceptor site of exon 8 in Y-MESO-61. (c) Genomic PCR analysis of exons of BAP1 detected homozygous deletion (HD) in Y-MESO-25 cells (arrowhead). (d) Western blot analysis of BAP1. Expression of β-actin was used as the control. (e) Summary of five gene mutation statuses in MM cell lines. Red boxes indicate inactivating mutation or HD. The mutation statuses of neurofibromatosis type 2 (NF2), large tumor suppressor homolog 2 (LATS2), Salvador homolog 1 (SAV1), and cyclin-dependent kinase inhibitor 2A (CDKN2A) were previously reported.11 NLS, nuclear localization signal; UCH, ubiquitin COOH-terminal hydrolase.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4556387&req=5

fig01: BRCA1-associated protein 1 (BAP1) gene mutations in malignant mesothelioma (MM). (a) Schematic diagram of BAP1 mutations in MM cell lines. (b) Schematic diagram of an insertion mutation due to G to T change at the acceptor site of exon 8 in Y-MESO-61. (c) Genomic PCR analysis of exons of BAP1 detected homozygous deletion (HD) in Y-MESO-25 cells (arrowhead). (d) Western blot analysis of BAP1. Expression of β-actin was used as the control. (e) Summary of five gene mutation statuses in MM cell lines. Red boxes indicate inactivating mutation or HD. The mutation statuses of neurofibromatosis type 2 (NF2), large tumor suppressor homolog 2 (LATS2), Salvador homolog 1 (SAV1), and cyclin-dependent kinase inhibitor 2A (CDKN2A) were previously reported.11 NLS, nuclear localization signal; UCH, ubiquitin COOH-terminal hydrolase.
Mentions: We carried out mutational analyses of BAP1 using 25 MM cell lines. Among 19 cell lines established from Japanese patients, four non-synonymous or insertion/deletion mutations were found; a nonsense mutation in ACC-MESO-4, two truncating mutations in Y-MESO-9 and Y-MESO-14, and a 40-bp insertion mutation at the intron 7–exon 8 junction in Y-MESO-61 (Fig.1a,b). We also carried out sequence analysis of cDNA synthesized from these cell line RNAs and confirmed the expression of the mutant RNAs (data not shown). As our previous study11 using array comparative genomic hybridization analysis detected a possible homozygous deletion (HD) in Y-MESO-25 (Fig. S1a), we carried out a genomic PCR analysis and confirmed that this cell line has HD of BAP1 at exons 13–17 (Fig.1c). In total, we found BAP1 mutations in 26% (5/19) of the Japanese MM cell lines (Table1). In addition, of six cell lines, the mutation status of which has been previously reported by another group,14 we confirmed the same mutations in two cell lines (NCI-H28 and NCI-H2452) and in the WT among four cell lines (Table1).

Bottom Line: Transduction of the WT BAP1 vector into MM cells with homozygous deletion at the BAP1 3' side resulted in both inhibition of cell proliferation and anchorage-independent cell growth, whereas BAP1 mutants of a missense or C-terminal truncated form showed impaired growth inhibitory effects.Furthermore, using the MM cells with BAP1 deletion, we found that WT BAP1, and even a missense mutant, conferred a higher survival rate after IR compared to the control vector.Our results suggested that, whereas WT BAP1 suppresses MM cell proliferation and restores cell survival after IR damage, some mutant BAP1 may also moderately retain these functions.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Oncology, Aichi Cancer Center Research Institute, Nagoya, Japan.

Show MeSH
Related in: MedlinePlus