Extended Data Fig. 6: Somatic C-terminal truncation variants of SERPINA1 do not rescue A1AT-secretion but have a short protein half-life.

(a) Western blot analysis of A1AT secretion by immunoblotting of media conditioned for 24 h by CHO cells transfected with the indicated A1AT-variant constructs. Immunoblotting was performed using a monoclonal antibody raised against an N-terminal fragment of human α1-antitrypsin (MA5-15521). Images representative of n = 3 biologically independent experiments. (b) Micrographs of the ER marker protein mEmerald-KDEL expressed in COS7 cells (left images) with accompanying single-particle tracks of HaloTagged variants of A1AT labelled with PAJF646, colour coded by mean velocity (right images). Frequency distribution histograms show mean velocity of tracks analysed. A dashed line denotes the median velocity of M-A1AT. For each condition over 10,000 particle tracks were analysed across a minimum of 17 cells from 3 repeats. (c) CHO cells conditionally-expressing untagged Tetracycline-inducible (Tet-ON) Z-A1AT were transfected with expression vectors encoding HaloTagged variants of A1AT, 2 h prior to 1 µg/mL doxycycline treatment. Input and Halo-affinity purified samples were then separated by SDS-PAGE prior to immunoblotting with antibody recognising total (MA5-15521) A1AT. Images representative of n = 3 biologically independent experiments.