Abstract
Since the first genetic description of a rhomboid in Drosophila melanogaster, tremendous efforts have been geared towards elucidating the proteolytic mechanism of this particular class of intramembrane proteases. In particular, mammalian rhomboid proteases sparked our interest and we aimed to investigate the human homologue RHBDL4. In light of our recent finding of the amyloid precursor protein (APP) family as efficient substrates of RHBDL4, we were enticed to further study the specific proteolytic mechanism of this enzyme by comparing cleavage patterns of wild type APP and APP TMS chimeras. Here, we demonstrate that the introduction of positively charged amino acid residues in the TMS redirects the RHBDL4-mediated cleavage of APP from its ectodomain closer towards the TMS, possibly inducing an ER-associated degradation (ERAD) of the substrate. In addition, we concluded that the cytoplasmic tail and proposed palmitoylation sites in the ectodomain of APP are not essential for the RHBDL4-mediated APP processing. In summary, our previously identified APP ectodomain cleavages by RHBDL4 are a subsidiary mechanism to the proposed RHBDL4-mediated ERAD of substrates likely through a single cleavage near or within the TMS.
Funding source: Alzheimer Society
Award Identifier / Grant number: PT-58872
Funding statement: We thank Dr. Claus Pietrzik for kindly providing plasmid cDNAs. This research was supported by grants to L.M.M. by NSERC Discovery grant no. RGPIN-2015-04645, Canada Foundation of Innovation Leaders Opportunity Fund (CFI-LOF, 32565), Alzheimer Society of Canada Young Investigator award, Funder Id: 10.13039/501100000143, PT-58872 and Research Grant 17-02, Fonds d’innovation Pfizer-FRQS sur la maladie d’Alzheimer et les maladies apparentées, Funder Id: 10.13039/501100000156, no. 31288 and 36571, McGill Faculty of Medicine Incentive funding and an award from The Scottish Rite Charitable Foundation of Canada. S.J.R. received a NSERC USRA summer student stipend, Funder Id: 10.13039/501100000096, Grant Number: 16112.
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