Molecular characterization of two novel isoforms of the human calcitonin receptor.
Calcitonin inhibits bone resorption by acting on osteoclasts via a specific receptor. The calcitonin receptor (CTR) is also found in many other normal and malignant tissues and cell lines. It has been cloned and sequenced in several species including humans. It belongs to a subclass of seven-transmembrane G protein-coupled receptors. Four human CTR (H-CTR) isoforms generated by alternatively spliced mRNA have previously been described. Two H-CTR encoding DNAs containing an unidentified 50-bp insert are now reported from T47D cells. The 50-bp insert corresponds to a DNA region located between exon 9 and exon 10, and appears to originate from an alternative splicing process. The two H-CTR cDNAs encode 274 and 290 aa long isoforms. Both are deleted from the putative fourth transmembrane domain to C-tail. They differ by the presence (H-CTR5) or absence (H-CTR6) of a previously known 16-aa insert in the putative first intracellular loop. Cell- and tissue-distribution analysis using RT-PCR demonstrates that the shorter one, HCTR6, is more prevalent. The mRNA of both isoforms was detected in giant cell tumor, whereas only H-CTR6 mRNA was detected in TT cells and kidney tissue. Neither H-CTR5 nor H-CTR6 could be detected in peripheral blood mononuclear cells cultured in the presence of RANKL, in MCF7 cells, and in cortical brain and ovarian tissues. When H-CTR6 was transiently expressed in HEK293 cells, CT failed to induce production of cAMP or to bind to the receptor. These suggest either an intrinsic loss of ligand binding function, or an altered intracellular trafficking. Our findings therefore indicate the existence of two novel splice variants of the H-CTR and confirm that multiple splicing patterns could be involved in the post-transcriptional regulation of the gene.
Beaudreuil
J
J
INSERM Unité 606, Centre Viggo Petersen, Hôpital Lariboisière, 6 rue Guy Patin, 75475 Paris Cedex 10, France.
Balasubramanian
Sundaravadivel
S
Chenais
J
J
Taboulet
J
J
Frenkian
M
M
Orcel
P
P
Jullienne
A
A
Horne
W C
WC
de Vernejoul
M C
MC
Cressent
M
M
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D002116:Calcitonin / Q000378:metabolism; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers; D005091:Exons / Q000235:genetics; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D020033:Protein Isoforms / Q000235:genetics; D012323:RNA Processing, Post-Transcriptional; D018003:Receptors, Calcitonin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D017931:DNA Primers; D020033:Protein Isoforms; D018003:Receptors, Calcitonin; D011994:Recombinant Proteins; D002116:Calcitonin