Figure 1

Expression and processing of CB. A. Chromosomal location (adapted from NCBI/NIH) and exon-intron organization of the hCB gene (top panel, modified from [19]), alternative splicing variants of CB primary transcripts (centre panel, modified from [56]), and domain organization of the entire translation product (bottom panel). Lightened sectors: non-coding exons (5'-UTR and 3'-UTR); darkened sectors: translated CB regions. The transcript population CB(-2) encodes the entire CB sequence, the alternative transcript population CB(-2, 3) leads to the truncated translation product Δ⁵¹CB which lacks the complete signal sequence (pre) as well as parts of the proregion (N'pro). In vivo, the single chain form of the native CB is mainly split into the light and the heavy chain while the C'pro-region is cut off during secretion. B. Processing of intrinsic CB (C, control) and of transiently overexpressed Δ⁷²CB-EGFP (1) and CB(SC)-EGFP (2) in LCLC-103H cells (on the left: α-CB-Ab; on the right: α-GFP-Ab). The immunoblot reveals distinct native CB fragments at similar expression levels of ~31 kDa (single chain form), ~26 kDa (heavy chain), and several isoforms thereof indicated by weaker adjacent bands. Unspecific low molecular mass protein bands below signify degradation products. Additional expression products of ~62 kDa (α-CB, α-GFP) in the samples from the transfected cells represent entire unprocessed sequences. The Δ⁷²CB-EGFP band runs slightly slower than the CB(SC)-EGFP band. Weak bands at ~22 kDa (α-GFP) indicate that a small amount of the EGFP tag is removed from the fusion proteins.