Properties of chimeric prostacyclin/prostaglandin D2 receptors: site-directed mutagenesis reveals the significance of the isoleucine residue at position 323.
Mouse prostacyclin (mIP) receptors transiently expressed in Chinese hamster ovary (CHO) cells activated both adenylyl cyclase and phospholipase C, with a 33-fold preference for signaling through Gs. The prostacyclin (IP) receptor agonists cicaprost, iloprost, carbacyclin, and prostaglandin E1 showed a similar order of potency for activation of both signaling pathways in cells transiently transfected with the mIP and the chimeric prostacyclin/prostaglandin D2 (IPN-VII/DPC and IPN-V/DPVI-C) receptors. Substitution of the carboxyl-terminal tail of the prostacyclin receptor with the corresponding region of the mDP receptor (IPN-VII/DPC) produced a receptor with increased coupling to both Gs and Gq. However, this increased G-protein coupling was lost in the IPN-V/DPVI-C receptor. The observation that both these chimeric receptors can activate phospholipase C indicates that the carboxyl-terminal tail of the IP receptor is not entirely responsible for its ability to couple to Gq. Site-directed mutagenesis studies suggest that isoleucine at position 323 in the IPN-VII/DPC receptor plays an important role in mediating the increased potency of this chimeric receptor.
Wise
Helen
H
Department of Pharmacology, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China. helenwise@cuhk.edu.hk
Chow
Kevin B S
KB
Wing
Kam Yiu
KY
Kobayashi
Takuya
T
Tse
Dicky L Y
DL
Cheng
Christopher H K
CH
D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004789:Enzyme Activation; D016285:Iloprost / Q000378:metabolism; D066298:In Vitro Techniques; D007532:Isoleucine / Q000737:chemistry; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D044006:Receptors, Epoprostenol / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011971:Receptors, Immunologic; D011982:Receptors, Prostaglandin / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism
D044006:Receptors, Epoprostenol; D011971:Receptors, Immunologic; D011982:Receptors, Prostaglandin; D011993:Recombinant Fusion Proteins; D007532:Isoleucine; D010738:Type C Phospholipases; D000262:Adenylyl Cyclases; D016285:Iloprost; C036293:prostaglandin D2 receptor