The Frizzled Family

The Frizzled family of receptors [for recent reviews, see References 9, 10] includes 10 mammalian isoforms. Along with the related receptor Smoothened (SMO) [11] . Frizzleds were recently classified by the International Union of Basic and Clinical Pharmacology (IUPHAR) as a novel family of GPCRs [12] and taken up into the IUPHAR's database of GPCRs and list of 7TM receptors (see also www.iuphar.org). The recommended nomenclature for the mammalian Frizzleds 1-10 is FZD1-10.

Based on homology studies, FZDs were originally described as secretin receptor-like proteins [13] but more recently have been grouped together with bitter taste2 receptors ] 14] . The latter analysis indicated that human FZDs share from 20% to 40% identity, with closer identity in four different clusters, notably FZD1,2,7 (75% identity), FZD5,8 (70% identity), FZD4,9,10, and FZD3, 6 (50% identity). One of the most conserved regions between all the FZDs is the N-terminal cysteine-rich domain (CRD) linked to the receptor core by a highly divergent linker ranging from about 40-100 aa. Further, the receptors contain 7 hydrophobic stretches of 20-25 aa resembling the transmembrane helices I-VII as well as a short C-terminus in the range of 25-200 aa [8]. The C-terminus contains two domains important for binding of the PDZ (PSD-95/disc large/ZO-1 homologous) ligand (see Fig. 5.1): (1) a highly conserved KTxxxW sequence, which is necessary for FZD signal transduction, serves a docking site for the PDZ domain of the central phosphoprotein Dishevelled (DVL) and (2) a rather well-conserved terminal PDZ ligand domain that can interact with many intracellular proteins, for example, post-synaptic density proteins [9] .

Other structural features of classical GPCRs, which are implicated in receptor function and G protein coupling are the presence of charged amino acids, such as arginine residues both on the C- and N- t erminal side of the third intracellular loop are present in FZDs. Further, several FZDs (FZD3, 4, 5, 6, 8, 9, 10) contain cysteine residues, putative targets for palmytoylation, in the C-terminus, which are required for membrane anchorage and formation human FZD1: Intracellular loop 1

fzd, - 344-dmrrfsyperp-354 Intracellular loop 2

fzd, - 424-sltwflaagmkwgheaieansq-445 Intracellular loop 3

fzd, -511-vslfrirtimkhdgtktekleklmvr-536 C-terminus fzd, - 623-sgKJLNSWrkfytrltnskqge -647

human FZD2: Intracellular loop 1

fzd2 - 269-dmqrfryperp-279 Intracellular loop 2

fzd2 - 349-sltwflaagmkwgheaieansq-370 Intracellular loop 3

fzd2 - 436-vslfrirtimkhdgtkteklerlmvr-461 C-terminus fzd2-541-sgKTLHSWrkfytrlinsrhge -565

human FZD3: Intracellular loop 1

fzd3 - 227-dvtrfryperp-237 Intracellular loop 2

fzd3 - 310-twflaavpkwgseaiekka-328 Intracellular loop 3

fzd3 - 396-slnrvrieiplekenqdklvkfmir-420 C-terminus fzd3-499-gskKTCFEWasffhgrrkkeivnesrqvl qepdfaqsllrdpntpiirksrgtstqgtsthasstq lamvddqrskagsihskvssyhgslhrsrdgrytpc syrgmeerlphgsmsrltdhsrhssshrlneqsr hssirdlsnnpmthiihgtsmnrvieedgtsa-666

human FZD4: Intracellular loop 1

fzd4 - 244-dssrfsyperp-254 Intracellular loop 2

fzd4 - 324-tltwflaaglkwgheaiemhs-344 Intracellular loop 3

fzd4 - 411-valfkirsnlqkdgikidklerlmvk-436 C-terminus fzd4-499-KTLHTWqkcsnrlvnsgkvkrekrgngw vkpgkgse -537

human FZD5: Intracellular loop 1

fzd5 - 260-dmerfryperp-270 Intracellular loop 2

fzd5 - 337-sltwflaagmkwgneaiagyaq-358 Intracellular loop 3

fzd5 - 424-vslfrirsvikqggikidkleklmir-449 C-terminus fzd5-522-wsg KTVESWrrftsrcccrprrghksg gamaagdypeasaaligrtgppgpaatyhkqvs lshv -585

human FZD6: Intracellular loop 1

fzd6 - 223-dvrrfryperp-233 Intracellular loop 2

fzd6 - 306-twflaagrkwsceaieqka-324 Intracellular loop 3

fzd6 - 392-slnhvrqviqhdgrnqeklkkfmir-416 C-terminus fzd6-465-gskKICIEWagffkrnrkrdpisesrrvlq escefflkhnskvkhkkkhykpsshklkvisksmgist gatanhgtsavaitshdylgqetlteiqispetsmrev kadgastprlreqdcgepaspaasisrlsgeqvdgkg qagsvsesarsegrispksdiidtglaqsnnlqvpsss epsslkgsisllvhpvsgvrkeqgggchsdt-706

human FZD7: Intracellular loop 1

fzd7 - 278-dmrrfsyperp-288 Intracellular loop 2

fzd7 - 358-sltwflaagmkwgheaieansq-379 Intracellular loop 3

fzd7 - 445-vslfririimkhdgikiekleklmvr-470 C-terminus fzd7-550-sgKTLQSWrrfyhrlshsskge -574

human FZD8: Intracellular loop 1

fzd8 - 297-stflidmerfkyperp-312 Intracellular loop 2

fzd8 - 418-sltwflaagmkwgneaiagysqy-439 Intracellular loop 3

fzd8 - 505-vslfrirsvikqqdgptkihkleklmir-532 C-terminus fzd8-606-sgKTLESWrslcirccwaskgaavgggag ataagggggpgggggggpgggggpgggggslysdv stgltwrsgtassvsypkqmpl -694

human FZD9: Intracellular loop 1

fzd9 - 251-ltfllephrfqyperp-366 Intracellular loop 2

fzd9 - 337-twflaagkkwgheaieahg-355 Intracellular loop 3

fzd9 -422-valfhirkimktgginiekleklmvk-447 C-terminus fzd9-530-ssKTFQTWqslcyrkiaagrarakacrapg sygrgihchykapivvlhmiktdpslenpthl -591

human FZD10: Intracellular loop 1

fzd10 - 247-ltflidparfryperp-262 Intracellular loop 2

fzd10- 333-twflaagkkwgheaieans-351 Intracellular loop 3

fzd10 - 415-sgfvalfhirrvmkiggenidkleklmvr-443 C-terminus fzd10-524-tsKTLQSWqqvcsrrlkkksrrkpasviisgg iykkaqhpqkthhgkyeipaqsp -581

Figure 5.1 Amino acid sequence of the intracellular domains of human FZDi_i0. SwissProt accession numbers: FZDi: Q9UP38; FZD2: Q14332; FZD3; Q9NPG1; FZD4: Q9ULV1; FZD5: Q13467; FZD6: 060353; FZD7: 075084; FZD8: Q9H461;FZD9:000144; FZD10: Q9ULW2. Putative phosphorylation sites are marked as underlined letters in the amino acid sequences (determined with the MiniMotif Miner software). The internal KTxxxW-DVL binding sequence is highlighted in bold, and the terminal PDZ ligand domains are marked in light gray.

of a putative helix 8 between the helix VII and the cysteine [15]. Palmitoylation of GPCR at their C terminus was shown to be important for G protein coupling, receptor phosphorylation, and agonist-i nduced desensitization and downregulation [16].

In summary, the general architecture of the proteins indeed suggests that the FZDs belong to the superfamily of GPCRs [9, 17] even though other well-conserved GPCR features are lacking or are less well conserved such as the DRY motif at the C-terminus of the third intracellular loop.

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