Fig. 17.2 Isoforms of the rat H3 receptor. The genomic organization of the receptor (top) shows four exons (boxes, E1-E4) and three introns (sizes given in parentheses). Sequences corresponding to translated TM regions are identified as roman numerals (I-VII). An alternatively spliced region (grey box) is depicted between the stop sequences. Six isoforms of the receptor (labelled on the left) have been shown to form as a result of alternative splicing.
fourth intron and alternative splicing of the third intron results in three additional isoforms. Preliminary data (Leurs et al. unpublished) indicate that these new proteins are not able to bind ligands. For the human H3-receptor, a similar complexity has been observed. Currently, at least nine different human H3 -receptor isoforms have been identified as a result of various alternative splicing events (Coge et al. 2001a; Wellendorf et al. 2000). Again, splicing of the first intron at the C-terminal end of TM2 occurs, resulting in an inframe deletion of 14 amino acids and a non-functional receptor. Splicing within the I3 region results in the generation of H3-receptor proteins with 30, 80, and 116 amino acid deletions within I3 or a 144 amino acid deletion within I3 and TM6 and TM7. Another splicing event results in the deletion of 119 amino acids within TM5 and I3, whereas splicing within TM4 results in a frameshift and the loss of TM4, TM5, and a part of I3. Finally, double splicing events create H3-receptor proteins with either the 80 or 144 amino acid deletion and a novel eight amino acid C-terminus. Currently, only binding data and functional data have been reported for the various 80 and the 144 amino acid deletion variants and the H3-receptor protein that is generated after the frame shift. Only the two 80 amino acid deletion variants act as functional receptor proteins, although some controversy exists regarding their signalling (Coge et al. 2001a; Wellendorf et al. 2000). As found for the rat H3a and H3b isoforms, one study reported that the 80 amino acid deletion variant signals more efficiently compared with the 445 amino acid receptor protein and the former also shows a higher affinity for H3 agonists in binding studies (Wellendorf et al. 2000).
Based on the amino acid alignment (Fig. 17.1) and an intial 3D homology model, a role for Asp114 has been suggested to be important in ligand binding to the H3 receptor (Ligneau et al. 2000). Very recently, mutagenesis of the H3 receptor has been used to explore agonist binding and receptor activation (Uveges et al. 2002). Mutation of Asp114 indeed abolishes radioligand binding and agonist action. Moreover, an alanine scan of TM5 has indicated that Glu206 plays a key role in the interaction of agonist with the H3 receptor (Uveges et al. 2002). With respect to antagonist binding to the receptor, Thr119 and Ala122 are essential (probably due to indirect interactions) (Ligneau et al. 2000), explaining the marked species differences for some H3 antagonists (e.g. thioperamide, ciproxyfan), but not for others (clobenpropit) (Ligneau et al. 2000).
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