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. 2021 Mar 25;16:19. doi: 10.1186/s13024-021-00443-6

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© The Author(s) 2021

Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 6 — The gain-of-function role for hCD33m is signaling-dependent but not stem from loss of glycan-binding in hCD33M. a,b Cartoon diagram of the structure of (a) hCD33M and (b) hCD33m in the plasma membrane. The cytoplasmic tail of the proteins contains the ITIM and ITIM-like residues. Point mutants at each residue in each isoform were generated to test the importance of these residues. c,d Expression of (c) hCD33M (red) and (c) hCD33m (blue) and their associated mutants, transduced into CD33^−/− U937 cells. For hCD33M (c), cell surface expression (non-permeabilized) was assessed. For hCD33m (d), intracellular expression (permeabilized) was assessed. Isotype-stained cells are shown in grey. e,f Phagocytosis of aggregated fluorescent Aβ_1–42 in CD33^−/− U937 (clone 4) cells virally-transduced to express (e) hCD33M or (f) hCD33m, along with their associated ITIM and ITIM-like mutants. % Phagocytosis represents the cytochalasin-D subtracted values referenced to the average of the empty vector (EV) cells set to 100%. (N = 4–5). g Cell surface expression of hCD33M and its R119A glycan-binding deficient mutant. Isotype-stained cells are shown in grey. h Phagocytosis of aggregated fluorescent Aβ_1–42 in CD33^−/− U937 cells (clone 4) transduced to express WT or R119A hCD33M. % Phagocytosis represents the cytochalasin-D subtracted values referenced to the average of the empty vector (EV) cells set to 100%. (N = 5)