BSU11271 q

-f-tqXqav •ftsqXqav


•-rrppaqaaXagp •-hspptqaaXagp •-qpqXtrratih ••qpqXtqragXq

••tlvpiXayagtiaahenXrXpgawterasaar aaqitqvabeupcaXfdpafd ll»gvqllpf«pptgli»l»rkd SL----+ taiXa

TJAj Receptor Amino Acid Sequences

Alignment of lbeTXA2 receptor amino acid sequences of mouse (MUSTXA2R). rat (RATTA2R), bovine (BTU53485 & BTU53484), human pbcental (HUMHTAR), human endothelium (HSU11271). The amino acid sequences are available from the Genbaok file server under accession numbers: D10849(mouse lung), D21158 (rat kidney), US34&5 (bovine liver), US3484 (bovine heart), U11271 (human endothelium) and D38081 (human placenta). Dots indicate identity will the consensus sequence, asterisks represent gaps, dashes represent the lack of a consensus and blank areas indicate sequence has not been completed nor the acute-phase response stimulators, IL-1, IL-6, lipopolysacharide, C-reactive protein or TNF, induced expression of TP receptor protein (66). These results indicated that TP receptor gene transcription in HEL cells was controlled primarily by promoter region I (66). The TP receptor gene promoter region was characterized by D'Angelo, et al (64). They determined that the PMA/protein kinase C (PKC) responsiveness of the TP receptor gene was attributable to AP-2-like binding to sites between 1.85 and 1.95 kb 5' of the major transcription initiation site (64). This site was fiither defined as a 14-nucleotide C-rich sequence, flanked by an octanucleotide inverted repeat, located -1.938 to -1.925 kb 5' of the transcription start site (67). The PMA-responsive enhancer factor that bound to the C-rich sequence was identified as the transcription factor, Spl (67). However, Spl binding alone was not sufficient to facilitate phoibol enhancement of TP gene expression. Additional, unidentified down-stream elements, located between -1.91 and -1.84 kb, were also required (67).


Only one TP receptor protein was found to be coded by the TP receptor gene identified by Nusing et al. (61). Although these investigators found no evidence for alternative splicing of transcripts when RT-PCR of poly(A)+RNA obtained from human placenta, MEG-01 cells ormesangial cells was performed using primers flanking intron 2, a proposed second form of the human TP receptor (TP-P), considered to be a product of alternative splicing, was later cloned from human endothelial cells by Raychowdhury, et al (56,68). The nucleotide sequence of the full length TP-P_receptor cDNA obtained from endothelial cells was originally reported to be 1504 nucleotides with a single open reading frame, and a predicted sequence of369 amino acids (56). However, the accurate nucleotide sequence, published as a correction of the original work (68), predicted 407 amino acids. The sequence of the first 328 amino acids was exactly the same as that of the TP receptor cDNA derived from placenta (TP-a) (50), but the sequences diverged substantially thereafter. The final 15 amino acids of the carboxyl terminus of the TP-a_receptor were found to be replaced by a 79 amino acid segment in the TP*p_receptor (Figure 1, TABLE). The spliced segment was located entirely in the cytoplasmic tail distal to the seventh transmembrane domain. The nucleotides that encoded the divergent portion of the cytoplasmic tail, plus the 3' untranslated region of TP-p, were found to match a sequence contained entirely within the 3' untranslated region of TP-a_(56). Although Raychowdhury, et al found only TP-P mRNA in endothelial cells, and both TP-a_and TP-P_mRNA in placenta, they were unable to demonstrate either TP-a_or TP-P mRNA in platelets (56). In contrast, Hirata, et al (69) identified mRNAs for both TP-a and TP-P variants of the human TP receptor in human platelets.


Despite the description of several examples of impaired functional and biochemical responses to TP receptor agonists in human and animal platelets (70-76), only one TP receptor mutation has been described (77). A single amino acid substitution (Rw-L) was found in human TXR-a_cDNA obtained by RT-PCR from platelets of patients with a dominantly inherited bleeding disorder (See V. ALTERED TP RECEPTOR FUNCTION, A. TP RECEPTOR MUTATION below).

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