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. 2021 Mar 5;6:108. doi: 10.1038/s41392-021-00495-6

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© The Author(s) 2021

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — SRSF1-mediated splicing of Bcl-xL inhibits autophagy through disruption of the formation of the autophagy initiation complex. 293 T cells were co-transfected with different combinations of Beclin1, PIK3C3, and Bcl-xL expression vectors in a–f. a 293 T cells were co-transfected with pEGFP-C1-Beclin1 and control vector; or pEGFP-C1-Beclin1 and Flag-PIK3C3 expression vector; or pEGFP-C1-Beclin1 and HA-Bcl-xL expression vector, respectively. Co-immunoprecipitation assay was carried out with anti-GFP antibody and the precipitated complexes were measured by western blot with anti-HA, anti-Flag, or anti-GFP antibodies. b 293 T cells were co-transfected with pEGFP-C1-PIK3C3 and control vector; or pEGFP-C1-PIK3C3 and Flag-Beclin1 expression vector; or pEGFP-C1-PIK3C3, and Flag-Beclin1 and HA-Bcl-xL expression vectors. c 293 T cells were co-transfected with pEGFP-C1-Beclin1 and control vector; or pEGFP-C1-Beclin1 and Flag-PIK3C3 expression vector; or pEGFP-C1-Beclin1, and Flag-PIK3C3 and HA-Bcl-xL expression vectors. d 293 T cells were co-transfected with pEGFP-C1-PIK3C3 and control vector; or pEGFP-C1-PIK3C3, and Flag-Beclin1 expression vector; or pEGFP-C1-PIK3C3, and Flag-Beclin1 vector, with increased amounts of HA-Bcl-xL expression vectors (100 ng and 500 ng) respectively. e 293 T cells were co-transfected with pEGFP-C1-Beclin1 and control vector; or pEGFP-C1-Beclin1, and Flag-PIK3C3 expression vector; or pEGFP-C1-Beclin1, and Flag-PIK3C3 expression vector, with increased amounts of HA-Bcl-xL expression vectors (100 ng and 500 ng) respectively. Three experiments were conducted respectively and the mean ± SD of the relative ratio of the precipitated Beclin1 or PIK3C3 were analyzed and plotted in b–e. f 293 T cells were co-transfected with pEGFP-C1-Beclin1 and control vector; or pEGFP-C1-Beclin1 and Flag-Bcl-xL expression vector; or pEGFP-C1-Beclin1 and Flag-Bcl-xS expression vector. Co-immunoprecipitation assay was carried out with anti-Flag M2 beads and the precipitated complexes were measured by Western blot with anti-HA, anti-Flag, or anti-GFP antibodies in b–f. g H1299 cells with overexpression of Bcl-xL were used for the immunoprecipitation experiments. Immunoprecipitation assay was performed with anti-PIK3C3 antibody and the endogenously precipitated complexes were measured by western blot assay with anti-Beclin1 antibody. The lysate was applied to examine the autophagy status