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. 2020 Feb 27;11:1101. doi: 10.1038/s41467-020-14931-8

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 5 — a Chromatin immunoprecipitation (ChIP) assay of SP1 association with the Inpp5a promoter in HEK293 cells transfected with TBP-13Q or TBP-105Q. Cross-linked chromatin materials were immunoprecipitated (IP) by anti-SP1 and were subjected to polymerase chain reaction (PCR) with primers specific for the promoter region of Inpp5a and PCNA (proliferating cell nuclear antigen), which served as a control. Representative agarose gel electrophoresis images of PCR products (left). Quantification of the ratios of immunoprecipitated products to input is presented (right). T = 7.757, **P = 0.0015 (unpaired t test). b Schematic map of a luciferase construct for reporter assay. c Western blotting showing the expression of SP1 and TBP in transfected PC12 cells. d Luciferase activity analysis of stable PC12 cell lines expressing TBP-13Q or TBP-105Q with or without transfected SP1. Luminescence intensity was analyzed with one-way ANOVA followed with Tukey’s multiple comparisons test. F = 35.19, *P < 0.05, **P < 0.005, ***P < 0.0005. e Western blot analysis of HEK293 cells transfected with SP1 and TBP-13Q or 105Q. f Luciferase activity analysis of transfected HEK293 cells. Cells transfected only with Inpp5a promoter reporter were used as control. Luminescence intensity was analyzed with one-way ANOVA followed with Tukey’s multiple comparisons test. F = 16.39, *P < 0.05, **P < 0.005. g Co-immunoprecipitation of transfected SP1 with TBP-13Q or TBP-105Q in HEK293 cells showing an increased interaction of SP1 with soluble TBP-105Q compared to TBP-13Q. Right panel showing the ratio of precipitated TBP to input. T = 3.518, *P = 0.0245 (Student’s t test). h TBP antibody immunoprecipitation of mouse cerebellar lysates also showed that more SP1 was coprecipitated in KI mice than in WT mice. One-month-old mice were used. Right panel showing the ratio of precipitated SP1 to input. T = 4.389, *P = 0.0118 (Student’s t test). Data are represented as mean ± SEM. Source data and full blots are provided as a Source Data file.