Biochemical and Biophysical Research Communications
The KiSS-1 receptor GPR54 is essential for the development of the murine reproductive system
Section snippets
Materials and methods
Generation of GPR54 knockout mice. Mutant mice with targeted disruption of the GPR54 gene (GPR54 −/−) were custom generated by Deltagen (Palo Alto, CA). A colony was then established at SPRI and maintained according to the guidelines and ethical standards of the SPRI ACUC.
A gene targeting vector was constructed in which a 52 bp fragment on exon 2, corresponding to position 269–320 of the ORF [5] of the GPR54 receptor gene (GenBank AF343726) was replaced with a IRES-LacZ Neo insert containing a
GPR54 −/− mouse generation and phenotypic analysis
In order to generate GPR54 −/− mice, a targeting vector was designed to disrupt a 52 bp fragment from exon 2 of the GPR54 receptor and to replace it with a IRES-LacZ Neo insert. Neo-resistant ES cells clones were then screened by Southern blot analysis of HindIII-digested genomic DNA hybridized with a radiolabeled DNA fragment that hybridizes outside of and adjacent to the construct arm (Fig. 1A). Heterozygous mice (GPR54 +/−) were interbred and homozygous mice were recovered at the expected
Discussion
It has been recently demonstrated that the decapeptide KiSS-10 (KiSS-1 residues 112–121) isolated from human placenta can activate GPR54 through the Gαq-signaling pathway [2], [4], [11]. To further our understanding of the physiological role of GPR54, we generated homozygous GPR54-deficient mice. Analysis of the phenotype of these mice revealed abnormalities in the development of both male and female homozygous mice. These abnormalities included external and internal reproductive organs which
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