1. Conservation of key sequence features and endocytic localisation in TbCALM.
Conservation of key sequence features and endocytic localisation in TbCALM. (A) Overview of CALM and AP180 proteins from human (Hs), yeast (Saccharomyces cerevisiae, Sc) and T. brucei. The extent of the ANTH domain is indicated (black box) along with the distribution of putative binding sites for clathrin and adaptors (black and grey bars). Clathrin and adaptor sites are well conserved along with general domain architecture. (B) Multiple sequence alignment of CALM proteins from across eukaryotes. Residues highlighted in dark grey are conserved in over 80% of the included sequences, light grey indicates greater than 60% conservation. Taxa are indicated to the left and horizontal bars above the sequence indicate the positions of secondary structural features (Ford et al., 2001). Residues identified as important for PtdIns(4,5)P2 binding are denoted with asterisks below the sequence. PtdIns(4,5)P2-binding residues are well conserved. (C) Immunofluorescence localisation of endogenous-locus-tagged TbCALM–GFP in bloodstream form T. brucei. Incubation with ConA at 4°C specifically labels the flagellar pocket (green); anti-GFP staining (red) shows TbCALM–GFP colocalised with the flagellar pocket, consistent with an endocytic function. DNA is stained with DAPI to show the nucleus and kinetoplast (blue).
Paul T. Manna et al. J Cell Sci 2015;128:
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