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The immunoglobulin heavy chain VH6-1 promoter regulates Ig transcription in non-B cells.

Wu L, Liu Y, Zhu X, Zhang L, Chen J, Zhang H, Hao P, Zhang S, Huang J, Zheng J, Zhang Y, Zhang Y, Qiu X - Cancer Cell Int. (2014)

Bottom Line: Our data showed that the sequence 1200 bp upstream of VH6-1 exhibited promoter activity in both B and non-B cells.No new regulatory elements were identified within the region 1200 bp to 300 bp upstream of VH6-1.The regulatory mechanisms among different cell types controlling the production of IgM heavy chains are worth discussing.

View Article: PubMed Central - PubMed

Affiliation: Key laboratory of Carcinogenesis and Translational Research (Ministry of Education), Central Laboratory, Peking University Cancer Hospital & Institute, Beijing, 100142 China.

ABSTRACT

Background: Non-B cell immunoglobulins (Igs) are widely expressed in epithelial cancer cells. The past 20 years of research have demonstrated that non-B cell Igs are associated with cancer cell proliferation, the cellular cytoskeleton and cancer stem cells. In this study we explored the transcriptional mechanism of IgM production in non-B cells.

Methods: The promoter region of a V-segment of the heavy mu chain gene (VH6-1) was cloned from a colon cancer cell line HT-29. Next, the promoter activities in non-B cells and B-cells were detected using the dual-luciferase reporter assay. Then the transcription factor binding to the promoter regions was evaluated by electrophoretic mobility shift assays (EMSAs) and gel supershift experiments.

Results: Our data showed that the sequence 1200 bp upstream of VH6-1 exhibited promoter activity in both B and non-B cells. No new regulatory elements were identified within the region 1200 bp to 300 bp upstream of VH6-1. In addition, Oct-1 was found to bind to the octamer element of the Ig gene promoter in cancer cells, in contrast to B cells, which utilize the transcriptional factor Oct-2.

Conclusion: The regulatory mechanisms among different cell types controlling the production of IgM heavy chains are worth discussing.

No MeSH data available.


Related in: MedlinePlus

The octamer element is important but not essential for non-B cell-derived Ig gene transcription. (A) Schematic diagram of mutated 5′ deletion truncations of the 1.2-kb pGL3 construct with a 4-bp deletion in the octamer motif (AGGCAAAT). (B) The intact and mutated 5′ deletion truncations of the 1.2-kb pGL3 construct were transfected into HeLa, HeLa MR, HeLa S3, HT-29, MDA-MB-231, HEK 293, L02, HK2, and Raji cells. Luciferase activity was measured using a dual-luciferase reporter system. The results are representative of three independent experiments after normalization to renilla luciferase activity (internal controls). Each bar represents mean ± SD.
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Fig2: The octamer element is important but not essential for non-B cell-derived Ig gene transcription. (A) Schematic diagram of mutated 5′ deletion truncations of the 1.2-kb pGL3 construct with a 4-bp deletion in the octamer motif (AGGCAAAT). (B) The intact and mutated 5′ deletion truncations of the 1.2-kb pGL3 construct were transfected into HeLa, HeLa MR, HeLa S3, HT-29, MDA-MB-231, HEK 293, L02, HK2, and Raji cells. Luciferase activity was measured using a dual-luciferase reporter system. The results are representative of three independent experiments after normalization to renilla luciferase activity (internal controls). Each bar represents mean ± SD.

Mentions: There is an atypical octamer sequence in VH6-1 (AGGCAAAT) that deviates from the typical sequence (ATGCAAAT). To ascertain the function of the octamer elements located in the Ig VH6-1 promoter, a mutated plasmid (pGL3-1.2 kb-mt) was constructed by deleting the middle four base pairs of the octamer element (Figure 2A). Unexpectedly, the deletion construct still exhibited promoter activity, although the activity was lower than for the construct that included the complete promoter construct pGL3-1.2 kb in all five cancer cells, three immortalized diploid cells and Raji cells (Figure 2B). These results indicated that the octamer element of VH6-1 in non-B cells is important but not essential.Figure 2


The immunoglobulin heavy chain VH6-1 promoter regulates Ig transcription in non-B cells.

Wu L, Liu Y, Zhu X, Zhang L, Chen J, Zhang H, Hao P, Zhang S, Huang J, Zheng J, Zhang Y, Zhang Y, Qiu X - Cancer Cell Int. (2014)

The octamer element is important but not essential for non-B cell-derived Ig gene transcription. (A) Schematic diagram of mutated 5′ deletion truncations of the 1.2-kb pGL3 construct with a 4-bp deletion in the octamer motif (AGGCAAAT). (B) The intact and mutated 5′ deletion truncations of the 1.2-kb pGL3 construct were transfected into HeLa, HeLa MR, HeLa S3, HT-29, MDA-MB-231, HEK 293, L02, HK2, and Raji cells. Luciferase activity was measured using a dual-luciferase reporter system. The results are representative of three independent experiments after normalization to renilla luciferase activity (internal controls). Each bar represents mean ± SD.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4260249&req=5

Fig2: The octamer element is important but not essential for non-B cell-derived Ig gene transcription. (A) Schematic diagram of mutated 5′ deletion truncations of the 1.2-kb pGL3 construct with a 4-bp deletion in the octamer motif (AGGCAAAT). (B) The intact and mutated 5′ deletion truncations of the 1.2-kb pGL3 construct were transfected into HeLa, HeLa MR, HeLa S3, HT-29, MDA-MB-231, HEK 293, L02, HK2, and Raji cells. Luciferase activity was measured using a dual-luciferase reporter system. The results are representative of three independent experiments after normalization to renilla luciferase activity (internal controls). Each bar represents mean ± SD.
Mentions: There is an atypical octamer sequence in VH6-1 (AGGCAAAT) that deviates from the typical sequence (ATGCAAAT). To ascertain the function of the octamer elements located in the Ig VH6-1 promoter, a mutated plasmid (pGL3-1.2 kb-mt) was constructed by deleting the middle four base pairs of the octamer element (Figure 2A). Unexpectedly, the deletion construct still exhibited promoter activity, although the activity was lower than for the construct that included the complete promoter construct pGL3-1.2 kb in all five cancer cells, three immortalized diploid cells and Raji cells (Figure 2B). These results indicated that the octamer element of VH6-1 in non-B cells is important but not essential.Figure 2

Bottom Line: Our data showed that the sequence 1200 bp upstream of VH6-1 exhibited promoter activity in both B and non-B cells.No new regulatory elements were identified within the region 1200 bp to 300 bp upstream of VH6-1.The regulatory mechanisms among different cell types controlling the production of IgM heavy chains are worth discussing.

View Article: PubMed Central - PubMed

Affiliation: Key laboratory of Carcinogenesis and Translational Research (Ministry of Education), Central Laboratory, Peking University Cancer Hospital & Institute, Beijing, 100142 China.

ABSTRACT

Background: Non-B cell immunoglobulins (Igs) are widely expressed in epithelial cancer cells. The past 20 years of research have demonstrated that non-B cell Igs are associated with cancer cell proliferation, the cellular cytoskeleton and cancer stem cells. In this study we explored the transcriptional mechanism of IgM production in non-B cells.

Methods: The promoter region of a V-segment of the heavy mu chain gene (VH6-1) was cloned from a colon cancer cell line HT-29. Next, the promoter activities in non-B cells and B-cells were detected using the dual-luciferase reporter assay. Then the transcription factor binding to the promoter regions was evaluated by electrophoretic mobility shift assays (EMSAs) and gel supershift experiments.

Results: Our data showed that the sequence 1200 bp upstream of VH6-1 exhibited promoter activity in both B and non-B cells. No new regulatory elements were identified within the region 1200 bp to 300 bp upstream of VH6-1. In addition, Oct-1 was found to bind to the octamer element of the Ig gene promoter in cancer cells, in contrast to B cells, which utilize the transcriptional factor Oct-2.

Conclusion: The regulatory mechanisms among different cell types controlling the production of IgM heavy chains are worth discussing.

No MeSH data available.


Related in: MedlinePlus