Human macrophage cathepsin B-mediated C-terminal cleavage of apolipoprotein A-I at Ser(228) severely impairs antiatherogenic capacity

Donna Lee M. Dinnes, Melanie Y. White, Stuart J. Cordwell, Leonard Kritharides, Maaike Kockx, Mathew Traini, Victar Hsieh, Mi-Jurng Kim, Liming Hou, Wendy Jessup, Kerry-Anne Rye, Morten Thaysen-Andersen

Research output: Contribution to journalArticlepeer-review

13 Citations (Scopus)

Abstract

Apolipoprotein A-I (apoA-I) is the major component of HDL and central to the ability of HDL to stimulate ATP-binding cassette transporter A1 (ABCA1)-dependent, antiatherogenic export of cholesterol from macrophage foam cells, a key player in the pathology of atherosclerosis. Cell-mediated modifications of apoA-I, such as chlorination, nitration, oxidation, and proteolysis, can impair its antiatherogenic function, although it is unknown whether macrophages themselves contribute to such modifications. To investigate this, human monocyte-derived macrophages (HMDMs) were incubated with human apoA-I under conditions used to induce cholesterol export. Two-dimensional gel electrophoresis and Western blot analysis identified that apoA-I is cleaved (∼20-80%) by HMDMs in a time-dependent manner, generating apoA-I of lower MW and isoelectric point. Mass spectrometry analysis identified a novel C-terminal cleavage site of apoA-I between Ser²²⁸-Phe²²⁹. Recombinant apoA-I truncated at Ser²²⁸ demonstrated profound loss of capacity to solubilize lipid and to promote ABCA1-dependent cholesterol efflux. Protease inhibitors, small interfering RNA knockdown in HMDMs, mass spectrometry analysis, and cathepsin B activity assays identified secreted cathepsin B as responsible for apoA-I cleavage at Ser²²⁸. Importantly, C-terminal cleavage of apoA-I was also detected in human carotid plaque. Cleavage at Ser²²⁸ is a novel, functionally important post-translational modification of apoA-Imediated byHMDMsthat limits the antiatherogenic properties of apoA-I.
Original languageEnglish
Pages (from-to)4239-4255
Number of pages17
JournalFASEB Journal
Volume30
Issue number12
DOIs
Publication statusPublished - Dec 2016

Keywords

  • atherosclerosis
  • coronary artery disease
  • high-density lipoprotein
  • proteolysis

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