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Expression and distribution of PPP2R5C gene in leukemia.

Zheng H, Chen Y, Chen S, Niu Y, Yang L, Li B, Lu Y, Geng S, Du X, Li Y - J Hematol Oncol (2011)

Bottom Line: Significantly higher expression of PPP2R5C was found in AML, CML, T-ALL, and B-CLL groups in comparison with healthy controls.High expression of PPP2R5C was detected in the B-ALL group; however, no significant difference was found compared with the healthy group.The expression level of PPP2R5C in the CML-CR group decreased significantly compared with that in the de novo CML group and was not significantly different from the level in the healthy group.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Hematology, Medical College, Jinan University, Guangzhou, People's Republic of China.

ABSTRACT

Background: Recently, we clarified at the molecular level novel chromosomal translocation t(14;14)(q11;q32) in a case of Sézary syndrome, which caused a rearrangement from TRAJ7 to the PPP2R5C gene. PPP2R5C is one of the regulatory B subunits of protein phosphatase 2A (PP2A). It plays a crucial role in cell proliferation, differentiation, and transformation. To characterize the expression and distribution of five different transcript variants of the PPP2R5C gene in leukemia, we analyzed the expression level of PPP2R5C in peripheral blood mononuclear cells from 77 patients with de novo leukemia, 26 patients with leukemia in complete remission (CR), and 20 healthy individuals by real-time PCR and identified the different variants of PPP2R5C by RT-PCR.

Findings: Significantly higher expression of PPP2R5C was found in AML, CML, T-ALL, and B-CLL groups in comparison with healthy controls. High expression of PPP2R5C was detected in the B-ALL group; however, no significant difference was found compared with the healthy group. The expression level of PPP2R5C in the CML-CR group decreased significantly compared with that in the de novo CML group and was not significantly different from the level in the healthy group. By using different primer pairs that covered different exons, five transcript variants of PPP2R5C could be identified. All variants could be detected in healthy samples as well as in all the leukemia samples, and similar frequencies and distributions of PPP2R5C were indicated.

Conclusions: Overexpression of PPP2R5C in T-cell malignancy as well as in myeloid leukemia cells might relate to its proliferation and differentiation. Investigation of the effect of target inhibition of this gene might be beneficial to further characterization of molecular mechanisms and targeted therapy in leukemia.

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Results of PCR amplification for PPP2R5C gene using different primer pairs. Lane M: 100-bp DNA ladder; 1 and 2: amplicom using PPP2R5C1f-/PPP2R5C1b primers; small products were 277 bp (12 + 14 exons) and large products were 394 bp (12 + 13 + 14 exons); 3 and 4: amplicom using PPP2R5C3f/PPP2R5C3b primers, the product was 304 bp (10 + 11 + 12 + 12a exons); 5 and 6: amplicom using PPP2R5C5f/PPP2R5C5b primers, the product was 242 bp (III + 2 exons); 7 and 8: ampilcom using PPP2R5C6f/PPP2R5C6b primers, the product was 213 bp (IV + 2 exons); 9: negative control.
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Figure 4: Results of PCR amplification for PPP2R5C gene using different primer pairs. Lane M: 100-bp DNA ladder; 1 and 2: amplicom using PPP2R5C1f-/PPP2R5C1b primers; small products were 277 bp (12 + 14 exons) and large products were 394 bp (12 + 13 + 14 exons); 3 and 4: amplicom using PPP2R5C3f/PPP2R5C3b primers, the product was 304 bp (10 + 11 + 12 + 12a exons); 5 and 6: amplicom using PPP2R5C5f/PPP2R5C5b primers, the product was 242 bp (III + 2 exons); 7 and 8: ampilcom using PPP2R5C6f/PPP2R5C6b primers, the product was 213 bp (IV + 2 exons); 9: negative control.

Mentions: It has been reported that five transcript variants of PPP2R5C might contribute to the specificity of PP2A [4]. However, little is known about the distribution of different variants in different organs, tissues, and cells, as well as in leukemia cells. Based on the structure of the PPP2R5C gene reported from Genbank and previous studies [4], we drew a schematic diagram of the genomic organization of PPP2R5C with five transcript variants (Figure 1), designed four primer pairs to amplify different exons, and tried to identify different variants in the same sample. By using PPP2R5C-1f/PPP2R5C-1b, which covered exons 12-14, two expected PCR products were detected. The small one comprised 277 bp containing exon 12 and 14 segments (corresponding to B56γ2 or B56γ6), and the large one comprised 394 bp containing exon 12, 13, and 14 segments (corresponding to B56γ3 or B56γ5). PPP2R5C-3f/PPP2R5C-3b, PPP2R5C-5f/PPP2R5C-5b, and PPP2R5C-6f/PPP2R5C-6b primer pairs covered exons 10 to 12a, exon III to 2, and exon IV to 2, respectively, and the expected PCR products were 304, 242, and 213 bp (corresponding to transcript B56γ1, B56γ5, and B56γ6 respectively) (Table 2 Figure 4). According the structure of transcript variants of PPP2R5C gene and the size of the amplicons, we confirmed the B56γ1, B56γ5, and B56γ6 variants. However, using the designed primer pairs, we were unable to distinguish between transcripts 1 and 5 or between transcripts 2 and 6 because PCR products of the same size were amplified by using the PPP2R5C-1f/PPP2R5C-1b, and it could not distinguish these variants using different primer pair combinations, except for whole gene sequencing. B56γ3 (variant 1) and B56γ2 (variant 2) are the frequency variants; therefore, it is thought that they might be expressed when positive PCR products are found. More importantly, using the present methods, we confirmed the expression of B56γ5 and B56γ6, which have been newly identified. Therefore, it could be concluded that all variants can be detected in healthy as well as leukemia samples with a similar frequency and distribution of PPP2R5C.


Expression and distribution of PPP2R5C gene in leukemia.

Zheng H, Chen Y, Chen S, Niu Y, Yang L, Li B, Lu Y, Geng S, Du X, Li Y - J Hematol Oncol (2011)

Results of PCR amplification for PPP2R5C gene using different primer pairs. Lane M: 100-bp DNA ladder; 1 and 2: amplicom using PPP2R5C1f-/PPP2R5C1b primers; small products were 277 bp (12 + 14 exons) and large products were 394 bp (12 + 13 + 14 exons); 3 and 4: amplicom using PPP2R5C3f/PPP2R5C3b primers, the product was 304 bp (10 + 11 + 12 + 12a exons); 5 and 6: amplicom using PPP2R5C5f/PPP2R5C5b primers, the product was 242 bp (III + 2 exons); 7 and 8: ampilcom using PPP2R5C6f/PPP2R5C6b primers, the product was 213 bp (IV + 2 exons); 9: negative control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3117819&req=5

Figure 4: Results of PCR amplification for PPP2R5C gene using different primer pairs. Lane M: 100-bp DNA ladder; 1 and 2: amplicom using PPP2R5C1f-/PPP2R5C1b primers; small products were 277 bp (12 + 14 exons) and large products were 394 bp (12 + 13 + 14 exons); 3 and 4: amplicom using PPP2R5C3f/PPP2R5C3b primers, the product was 304 bp (10 + 11 + 12 + 12a exons); 5 and 6: amplicom using PPP2R5C5f/PPP2R5C5b primers, the product was 242 bp (III + 2 exons); 7 and 8: ampilcom using PPP2R5C6f/PPP2R5C6b primers, the product was 213 bp (IV + 2 exons); 9: negative control.
Mentions: It has been reported that five transcript variants of PPP2R5C might contribute to the specificity of PP2A [4]. However, little is known about the distribution of different variants in different organs, tissues, and cells, as well as in leukemia cells. Based on the structure of the PPP2R5C gene reported from Genbank and previous studies [4], we drew a schematic diagram of the genomic organization of PPP2R5C with five transcript variants (Figure 1), designed four primer pairs to amplify different exons, and tried to identify different variants in the same sample. By using PPP2R5C-1f/PPP2R5C-1b, which covered exons 12-14, two expected PCR products were detected. The small one comprised 277 bp containing exon 12 and 14 segments (corresponding to B56γ2 or B56γ6), and the large one comprised 394 bp containing exon 12, 13, and 14 segments (corresponding to B56γ3 or B56γ5). PPP2R5C-3f/PPP2R5C-3b, PPP2R5C-5f/PPP2R5C-5b, and PPP2R5C-6f/PPP2R5C-6b primer pairs covered exons 10 to 12a, exon III to 2, and exon IV to 2, respectively, and the expected PCR products were 304, 242, and 213 bp (corresponding to transcript B56γ1, B56γ5, and B56γ6 respectively) (Table 2 Figure 4). According the structure of transcript variants of PPP2R5C gene and the size of the amplicons, we confirmed the B56γ1, B56γ5, and B56γ6 variants. However, using the designed primer pairs, we were unable to distinguish between transcripts 1 and 5 or between transcripts 2 and 6 because PCR products of the same size were amplified by using the PPP2R5C-1f/PPP2R5C-1b, and it could not distinguish these variants using different primer pair combinations, except for whole gene sequencing. B56γ3 (variant 1) and B56γ2 (variant 2) are the frequency variants; therefore, it is thought that they might be expressed when positive PCR products are found. More importantly, using the present methods, we confirmed the expression of B56γ5 and B56γ6, which have been newly identified. Therefore, it could be concluded that all variants can be detected in healthy as well as leukemia samples with a similar frequency and distribution of PPP2R5C.

Bottom Line: Significantly higher expression of PPP2R5C was found in AML, CML, T-ALL, and B-CLL groups in comparison with healthy controls.High expression of PPP2R5C was detected in the B-ALL group; however, no significant difference was found compared with the healthy group.The expression level of PPP2R5C in the CML-CR group decreased significantly compared with that in the de novo CML group and was not significantly different from the level in the healthy group.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute of Hematology, Medical College, Jinan University, Guangzhou, People's Republic of China.

ABSTRACT

Background: Recently, we clarified at the molecular level novel chromosomal translocation t(14;14)(q11;q32) in a case of Sézary syndrome, which caused a rearrangement from TRAJ7 to the PPP2R5C gene. PPP2R5C is one of the regulatory B subunits of protein phosphatase 2A (PP2A). It plays a crucial role in cell proliferation, differentiation, and transformation. To characterize the expression and distribution of five different transcript variants of the PPP2R5C gene in leukemia, we analyzed the expression level of PPP2R5C in peripheral blood mononuclear cells from 77 patients with de novo leukemia, 26 patients with leukemia in complete remission (CR), and 20 healthy individuals by real-time PCR and identified the different variants of PPP2R5C by RT-PCR.

Findings: Significantly higher expression of PPP2R5C was found in AML, CML, T-ALL, and B-CLL groups in comparison with healthy controls. High expression of PPP2R5C was detected in the B-ALL group; however, no significant difference was found compared with the healthy group. The expression level of PPP2R5C in the CML-CR group decreased significantly compared with that in the de novo CML group and was not significantly different from the level in the healthy group. By using different primer pairs that covered different exons, five transcript variants of PPP2R5C could be identified. All variants could be detected in healthy samples as well as in all the leukemia samples, and similar frequencies and distributions of PPP2R5C were indicated.

Conclusions: Overexpression of PPP2R5C in T-cell malignancy as well as in myeloid leukemia cells might relate to its proliferation and differentiation. Investigation of the effect of target inhibition of this gene might be beneficial to further characterization of molecular mechanisms and targeted therapy in leukemia.

Show MeSH
Related in: MedlinePlus