Autophagy is a pivotal regulatory and catabolic process, induced under various stressful conditions, including hypoxia. However, little is known about alternative splicing of autophagy genes in the hypoxic landscape in breast cancer. Our research unravels the hitherto unreported alternative splicing of BNIP3L, a crucial hypoxia-induced autophagic gene. We showed that BNIP3L, under hypoxic condition, forms two isoforms, a full-length isoform (BNIP3L-F) and a shorter isoform lacking exon 1 (BNIP3L-Δ1). The hypoxia-induced BNIP3L-F promotes autophagy, while under normoxia, the BNIP3L-Δ1 inhibits autophagy. We discovered a novel dimension of hypoxia-mediated epigenetic modification that regulates the alternative splicing of BNIP3L. Here, we showed differential DNA methylation of BNIP3L intron 1, causing reciprocal binding of epigenetic factor CCCTC-binding factor (CTCF) and its paralog BORIS. Additionally, we highlighted the role of CTCF and BORIS impacting autophagy in breast cancer. The differential binding of CTCF and BORIS results in alternative splicing of BNIP3L forming BNIP3L-F and BNIP3L-Δ1, respectively. The binding of CTCF on unmethylated BNIP3L intron 1 under hypoxia results in RNA Pol-II pause and inclusion of exon 1, promoting BNIP3L-F and autophagy. Interestingly, the binding of BORIS on methylated BNIP3L intron 1 under normoxia also results in RNA Pol-II pause but leads to the exclusion of exon 1 from BNIP3L mRNA. Finally, we reported the critical role of BORIS-mediated RNA Pol-II pause, which subsequently recruits SRSF6, redirecting the proximal splice-site selection, promoting BNIP3L-Δ1, and inhibiting autophagy. Our study provides novel insights into the potential avenues for breast cancer therapy by targeting autophagy regulation, specifically under hypoxic condition.

中文翻译：

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CTCF 和 BORIS 在乳腺癌中通过 BNIP3L 选择性剪接介导的自噬调节


自噬是一个关键的调节和分解代谢过程，在各种压力条件下（包括缺氧）诱导。然而，人们对乳腺癌缺氧环境中自噬基因的选择性剪接知之甚少。我们的研究揭示了迄今为止尚未报道的 BNIP3L（缺氧诱导的重要自噬基因）的选择性剪接。我们发现 BNIP3L 在缺氧条件下形成两种亚型，一种全长亚型 (BNIP3L-F) 和一种缺少外显子 1 的较短亚型 (BNIP3L-Δ1)。缺氧诱导的 BNIP3L-F 促进自噬，而在常氧条件下，BNIP3L-Δ1 抑制自噬。我们发现了缺氧介导的表观遗传修饰的一个新维度，可以调节 BNIP3L 的选择性剪接。在这里，我们展示了 BNIP3L 内含子 1 的差异 DNA 甲基化，导致表观遗传因子 CCCTC 结合因子 (CTCF) 及其旁系同源物 BORIS 相互结合。此外，我们强调了 CTCF 和 BORIS 对乳腺癌自噬的影响。 CTCF 和 BORIS 的差异结合导致 BNIP3L 选择性剪接，分别形成 BNIP3L-F 和 BNIP3L-Δ1。在缺氧条件下，CTCF 与未甲基化的 BNIP3L 内含子 1 结合，导致 RNA Pol-II 暂停并包含外显子 1，从而促进 BNIP3L-F 和自噬。有趣的是，常氧下 BORIS 与甲基化 BNIP3L 内含子 1 的结合也会导致 RNA Pol-II 暂停，但导致外显子 1 从 BNIP3L mRNA 中排除。最后，我们报道了 BORIS 介导的 RNA Pol-II 暂停的关键作用，该暂停随后招募 SRSF6，重定向近端剪接位点选择，促进 BNIP3L-Δ1 并抑制自噬。 我们的研究通过针对自噬调节，特别是在缺氧条件下，为乳腺癌治疗的潜在途径提供了新的见解。