S-acylation status of bile acid transporter hASBT regulates its function, metabolic stability, membrane expression, and phosphorylation state

Ebehiremen N. Ayewoh, Lindsay C. Czuba, Thao T. Nguyen, Peter W. Swaan

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

The human apical sodium-dependent bile acid transporter (hASBT, SLC10A2) is the rate-limiting step of intestinal bile acid absorption in the enterohepatic circulation system of bile acids. Therefore, the regulation and stability of hASBT is vital in maintaining bile acid and cholesterol homeostasis and may serve as a potential target for cholesterol-related disorders. We hypothesized that post-translational mechanisms that govern hASBT function and regulation will provide novel insight on intestinal bile acid transport and homeostasis. In this study, we confirm the S-acylation status of hASBT via acyl biotin exchange in COS-1 cells and its impact on hASBT expression, function, kinetics, and protein stability. Using the acylation inhibitor, 2-bromopalmitate, we show that S-acylation is an important modification which modulates the function, surface expression, and maximal transporter flux (Jmax) of hASBT. By means of proteasome inhibitors, S-acylated hASBT was found to be cleared via the proteasome whereas a reduction in the palmitoylation status of hASBT resulted in rapid proteolytic degradation compared to the unmodified transporter. Screening of cysteine mutants in and or near transmembrane domains, some of which are exposed to the cytosol, confirmed Cys314 to be the predominate S-acylated residue. Lastly, we show that S-acylation was reduced in a mutant form of hASBT devoid of cytosolic facing tyrosine residues, suggestive of crosstalk between acylation and phosphorylation post-translational modification mechanisms.

Original languageEnglish
Article number183510
JournalBiochimica et Biophysica Acta - Biomembranes
Volume1863
Issue number2
DOIs
StatePublished - Feb 1 2021

Bibliographical note

Publisher Copyright:
© 2020

Keywords

  • ASBT
  • Lipids
  • Palmitoylation
  • Post-translational modifications
  • Transporter

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Cell Biology

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