Genomic organization and characterization of splice variants of the human histamine H3 receptor.

In the present paper we report the genomic organization of the human histamine H 3 -receptor gene, which consists of four exons spanning 5.5kb on chromosome 20. Using PCR, six alternative splice variants of the H 3 receptor were cloned from human thalamus. These variants were found to be coexpressed in human brain, but their relative distribution varied in a region-specific manner. These isoforms displayed either a deletion in the putative second transmembrane domain (TM), H 3(∆TM2, 431aa) or a variable deletion in the third intracellular loop (i3), H 3(∆i3, 415aa) , H 3(∆i3, 365aa) , H 3(∆i3, 329aa) and H 3(∆TM5+∆i3, 326aa) . In order to determine the biological role of the H 3 receptor variants compared with the ‘original’ H 3(445aa) receptor, three isoforms, namely H 3(445aa) , H 3(∆TM2, 431aa) and H 3(∆i3, 365aa) , were expressed in CHO cells and their pharmacological properties were investigated. Binding studies showed that H 3(∆TM2, 431aa) transiently expressed in CHO cells was unable to bind [ 125 I]iodoproxyfan, whereas both the H 3(445aa) and H 3(∆i3, 365aa) receptors displayed a high affinity for [ 125 I]iodoproxyfan [ K d = 28±5pM ( n = 4) and 8±1pM ( n = 5) respectively]. In addition, H 3(∆i3, 365aa) possessed the same pharmacological profile as the H 3(445aa) receptor. However, in CHO cells expressing H 3(∆i3, 365aa) , H 3 agonists did not inhibit forskolin-induced cAMP production, stimulate [ 35 S]guanosine 5′-[γ-thio]triphosphate ([ 35 S]GTP[S]) binding or stimulate intracellular Ca 2+ mobilization. Therefore the 80-amino-acid sequence located at the C-terminal portion of i3 plays an essential role in H 3 agonist-mediated signal transduction. The existence of multiple H 3 isoforms with different signal transduction capabilities suggests that H 3 -mediated biological functions might be tightly regulated through alternative splicing mechanisms.