A proposed bovine neuropeptide Y (NPY) receptor cDNA clone, or its human homologue, confers neither NPY binding sites nor NPY responsiveness on transfected cells

Elena E. Jazin, Heahyun Yoo, Anders G. Blomqvist, Frances Yee, Gezhi Weng, Mary W. Walker, John Salon, Dan Larhammar, Claes R Wahlestedt

Research output: Contribution to journalArticle

76 Citations (Scopus)

Abstract

Receptors with seven transmembrane domains (7TM) constitute a large family of structurally and functionally related proteins which respond to various types of ligands. We describe here the cloning and expression of a human 7TM receptor, denoted hFB22 (human Fetal Brain 22), which is the homologue (92% amino acid identity) of a bovine receptor (LCR1) reported by others to bind neuropeptide Y (NPY) with a pharmacological profile of the Y3 receptor subtype. However, upon expression in COS1 (confirmed by Northern analysis), COS7 or CHO-K1 cells, the hFB22 receptor did not confer specific 125I-Bolton-Hunter-NPY, 3H-propionyl-NPY or 125I-peptide YY (PYY) binding sites, in either intact cells or in membrane preparations. Similarly, cells transfected with the corresponding bovine clone (LCR1) did not show specific NPY/PYY binding exceeding that resulting from endogenous binding sites; mock-transfected COS7 cells, used frequently for heterologous expression of receptors, were found to have endogenous specific 125I-NPY binding sites (Bmax = 112 fmol/mg protein; Kd = 0.25 nM). Moreover, the hFB22 transfected cells, when compared to control transfected cells, did not display de novo NPY- or PYY-induced second messenger responses, i.e., (1) inhibition of forskolin-stimulated cAMP accumulation or (2) 45Ca2+ influx. The presence of hFB22 mRNA was detected in several human neuroblastoma cell lines, none of which was found to express Y3-like NPY binding sites. hFB22 displays 39% amino acid sequence identity (in the transmembrane regions) to the human interleukin-8 receptor, and 32-36% amino acid identity to the human receptors of angiotensin II, bradykinin, and n-formylpeptide, but only 23% amino acid identity to the previously described human NPY/PYY receptor of the Y1 receptor subtype. Our results show that hFB22 and LCR1 do not encode NPY receptors, and their true ligand(s) remains to be identified.

Original languageEnglish
Pages (from-to)247-258
Number of pages12
JournalRegulatory Peptides
Volume47
Issue number3
DOIs
StatePublished - Sep 22 1993
Externally publishedYes

Fingerprint

Neuropeptide Y Receptors
Neuropeptide Y
Complementary DNA
Clone Cells
Binding Sites
Brain
Peptide YY
Amino Acids
Interleukin-8 Receptors
Ligands
Angiotensin Receptors
Cloning
Second Messenger Systems
Bradykinin
Colforsin
CHO Cells
Proteins
Display devices
Cells
Neuroblastoma

Keywords

  • Binding site
  • Cloning
  • Gene expression
  • Human
  • Neuropeptide Y receptor
  • Responsiveness

ASJC Scopus subject areas

  • Biochemistry
  • Endocrinology
  • Physiology
  • Neuroscience(all)

Cite this

A proposed bovine neuropeptide Y (NPY) receptor cDNA clone, or its human homologue, confers neither NPY binding sites nor NPY responsiveness on transfected cells. / Jazin, Elena E.; Yoo, Heahyun; Blomqvist, Anders G.; Yee, Frances; Weng, Gezhi; Walker, Mary W.; Salon, John; Larhammar, Dan; Wahlestedt, Claes R.

In: Regulatory Peptides, Vol. 47, No. 3, 22.09.1993, p. 247-258.

Research output: Contribution to journalArticle

Jazin, Elena E. ; Yoo, Heahyun ; Blomqvist, Anders G. ; Yee, Frances ; Weng, Gezhi ; Walker, Mary W. ; Salon, John ; Larhammar, Dan ; Wahlestedt, Claes R. / A proposed bovine neuropeptide Y (NPY) receptor cDNA clone, or its human homologue, confers neither NPY binding sites nor NPY responsiveness on transfected cells. In: Regulatory Peptides. 1993 ; Vol. 47, No. 3. pp. 247-258.
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abstract = "Receptors with seven transmembrane domains (7TM) constitute a large family of structurally and functionally related proteins which respond to various types of ligands. We describe here the cloning and expression of a human 7TM receptor, denoted hFB22 (human Fetal Brain 22), which is the homologue (92{\%} amino acid identity) of a bovine receptor (LCR1) reported by others to bind neuropeptide Y (NPY) with a pharmacological profile of the Y3 receptor subtype. However, upon expression in COS1 (confirmed by Northern analysis), COS7 or CHO-K1 cells, the hFB22 receptor did not confer specific 125I-Bolton-Hunter-NPY, 3H-propionyl-NPY or 125I-peptide YY (PYY) binding sites, in either intact cells or in membrane preparations. Similarly, cells transfected with the corresponding bovine clone (LCR1) did not show specific NPY/PYY binding exceeding that resulting from endogenous binding sites; mock-transfected COS7 cells, used frequently for heterologous expression of receptors, were found to have endogenous specific 125I-NPY binding sites (Bmax = 112 fmol/mg protein; Kd = 0.25 nM). Moreover, the hFB22 transfected cells, when compared to control transfected cells, did not display de novo NPY- or PYY-induced second messenger responses, i.e., (1) inhibition of forskolin-stimulated cAMP accumulation or (2) 45Ca2+ influx. The presence of hFB22 mRNA was detected in several human neuroblastoma cell lines, none of which was found to express Y3-like NPY binding sites. hFB22 displays 39{\%} amino acid sequence identity (in the transmembrane regions) to the human interleukin-8 receptor, and 32-36{\%} amino acid identity to the human receptors of angiotensin II, bradykinin, and n-formylpeptide, but only 23{\%} amino acid identity to the previously described human NPY/PYY receptor of the Y1 receptor subtype. Our results show that hFB22 and LCR1 do not encode NPY receptors, and their true ligand(s) remains to be identified.",
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AU - Yoo, Heahyun

AU - Blomqvist, Anders G.

AU - Yee, Frances

AU - Weng, Gezhi

AU - Walker, Mary W.

AU - Salon, John

AU - Larhammar, Dan

AU - Wahlestedt, Claes R

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N2 - Receptors with seven transmembrane domains (7TM) constitute a large family of structurally and functionally related proteins which respond to various types of ligands. We describe here the cloning and expression of a human 7TM receptor, denoted hFB22 (human Fetal Brain 22), which is the homologue (92% amino acid identity) of a bovine receptor (LCR1) reported by others to bind neuropeptide Y (NPY) with a pharmacological profile of the Y3 receptor subtype. However, upon expression in COS1 (confirmed by Northern analysis), COS7 or CHO-K1 cells, the hFB22 receptor did not confer specific 125I-Bolton-Hunter-NPY, 3H-propionyl-NPY or 125I-peptide YY (PYY) binding sites, in either intact cells or in membrane preparations. Similarly, cells transfected with the corresponding bovine clone (LCR1) did not show specific NPY/PYY binding exceeding that resulting from endogenous binding sites; mock-transfected COS7 cells, used frequently for heterologous expression of receptors, were found to have endogenous specific 125I-NPY binding sites (Bmax = 112 fmol/mg protein; Kd = 0.25 nM). Moreover, the hFB22 transfected cells, when compared to control transfected cells, did not display de novo NPY- or PYY-induced second messenger responses, i.e., (1) inhibition of forskolin-stimulated cAMP accumulation or (2) 45Ca2+ influx. The presence of hFB22 mRNA was detected in several human neuroblastoma cell lines, none of which was found to express Y3-like NPY binding sites. hFB22 displays 39% amino acid sequence identity (in the transmembrane regions) to the human interleukin-8 receptor, and 32-36% amino acid identity to the human receptors of angiotensin II, bradykinin, and n-formylpeptide, but only 23% amino acid identity to the previously described human NPY/PYY receptor of the Y1 receptor subtype. Our results show that hFB22 and LCR1 do not encode NPY receptors, and their true ligand(s) remains to be identified.

AB - Receptors with seven transmembrane domains (7TM) constitute a large family of structurally and functionally related proteins which respond to various types of ligands. We describe here the cloning and expression of a human 7TM receptor, denoted hFB22 (human Fetal Brain 22), which is the homologue (92% amino acid identity) of a bovine receptor (LCR1) reported by others to bind neuropeptide Y (NPY) with a pharmacological profile of the Y3 receptor subtype. However, upon expression in COS1 (confirmed by Northern analysis), COS7 or CHO-K1 cells, the hFB22 receptor did not confer specific 125I-Bolton-Hunter-NPY, 3H-propionyl-NPY or 125I-peptide YY (PYY) binding sites, in either intact cells or in membrane preparations. Similarly, cells transfected with the corresponding bovine clone (LCR1) did not show specific NPY/PYY binding exceeding that resulting from endogenous binding sites; mock-transfected COS7 cells, used frequently for heterologous expression of receptors, were found to have endogenous specific 125I-NPY binding sites (Bmax = 112 fmol/mg protein; Kd = 0.25 nM). Moreover, the hFB22 transfected cells, when compared to control transfected cells, did not display de novo NPY- or PYY-induced second messenger responses, i.e., (1) inhibition of forskolin-stimulated cAMP accumulation or (2) 45Ca2+ influx. The presence of hFB22 mRNA was detected in several human neuroblastoma cell lines, none of which was found to express Y3-like NPY binding sites. hFB22 displays 39% amino acid sequence identity (in the transmembrane regions) to the human interleukin-8 receptor, and 32-36% amino acid identity to the human receptors of angiotensin II, bradykinin, and n-formylpeptide, but only 23% amino acid identity to the previously described human NPY/PYY receptor of the Y1 receptor subtype. Our results show that hFB22 and LCR1 do not encode NPY receptors, and their true ligand(s) remains to be identified.

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