Regulatory mechanism of ABCB1 transcriptional repression by HDAC5 in rat hepatocytes under hypoxic environment

Ziqin Wei ¹ Hongfang Mu ² Fangfang Qiu ¹ Minghui Zhao ¹ Xiaojing Zhang ² Wenbin LI ² Rong Wang ^2*

• ¹ Lanzhou University, Lanzhou, China
• ² Key Laboratory of the Plateau of Environmental Damage Control, Lanzhou Army General Hospital, Lanzhou, Gansu Province, China

Previous research has demonstrated that the hypoxic environment at high altitudes significantly alters the pharmacokinetics of many drugs, reducing their efficacy and increasing adverse effects. A key factor in this altered drug metabolism is the inhibition of ATP-binding cassette subfamily B member 1 (ABCB1), an efflux transporter protein, in the liver tissues of plateau rats. Rat ABCB1, encoded by the ABCB1A and ABCB1B genes, has two isoforms functionally analogous to human ABCB1. Histone acetylation, an epigenetic mechanism, may regulate ABCB1 transcription in hypoxic conditions by modifying chromatin structure and interacting with signaling pathways. However, its role in ABCB1 transcriptional regulation under hypoxia remains unclear. In this study, we investigated histone acetylation's involvement in ABCB1 expression in BRL cells under hypoxic conditions using histone deacetylase (HDAC) inhibitors. Among those tested, tricostatin A (TSA), bufexamac, and vorinostat (SAHA) significantly upregulated ABCB1 expression. These HDAC inhibitors enhanced ABCB1 transcription by inhibiting histone deacetylase 5 (HDAC5), elevating histone 3 lysine 9 acetylation (H3K9ac) levels at the ABCB1B promoter. Notably, specific protein 1 (SP1) was upregulated during SAHA-induced ABCB1 expression, and SP1 knockdown blocked SAHA-mediated ABCB1 activation at both mRNA and protein levels. Coimmunoprecipitation and fluorescence colocalization confirmed interactions among ABCB1, HDAC5, and SP1. In BRL cells, HDAC5 may be recruited by SP1 to form a complex, reducing free HDAC5, increasing H3K9ac at the ABCB1B promoter, and activating ABCB1 transcription. In the BRL hypoxia model, disruption of the SP1-HDAC5 complex increased free HDAC5, lowered H3K9ac at the ABCB1B promoter, and suppressed ABCB1 transcription.Fluorescent dye accumulation assays using the ABCB1 substrate rhodamine 123 revealed that HDAC inhibitors restored ABCB1 efflux function, reducing intracellular rhodamine 123 accumulation in the hypoxia model, consistent with the transcriptional findings. These results suggest that HDAC inhibitors enhance ABCB1 expression in hypoxic environments, indicating that combining HDAC inhibitors with therapeutic agents could mitigate reduced drug efficacy and adverse effects caused by ABCB1 suppression.