Cloning and expression of cynomolgus monkey (Macaca fascicularis) gonadotropins luteinizing hormone and follicle-stimulating hormone and identification of two polymorphic sites in the luteinizing hormone beta subunit.
The genes encoding the cynomolgus monkey gonadotropin subunits, alpha, follicle-stimulating hormone (FSH) beta and luteinizing hormone (LH) beta, were cloned by reverse transcriptase polymerase chain reaction (RT-PCR) using pituitary RNA. The predicted amino acid sequences displayed 82, 96 and 87% identity to human subunit sequences, respectively. Northern blot hybridization of monkey tissues revealed pituitary specific transcripts of 1.0 and 0.6 kb for the alpha and LHbeta subunit, respectively, and two bands of 1.8 and 0.65 kb for the FSHbeta. Upon sequencing LHbeta cDNAs from different monkeys, two polymorphic sites were detected, resulting in the amino acid transitions Ser32Thr and His60Arg. Restriction analysis revealed different homo- and heterozygous combinations of the polymorphic sites indicating linkage dysequilibrium. Transient co-expression of the alpha subunit together with the FSHbeta or LHbeta subunit in COS7 and CHO cells resulted in secretion of in vitro bioactive hormones. This work represents a further step towards production of recombinant monkey LH and FSH which can be used in a homologous experimental setting in the cynomolgus monkey.
Schmidt
A
A
Institute of Reproductive Medicine of the University, Münster, Germany.
Gromoll
J
J
Weinbauer
G F
GF
Galla
H J
HJ
Chappel
S
S
Simoni
M
M
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D003001:Cloning, Molecular; D018076:DNA, Complementary; D005640:Follicle Stimulating Hormone / Q000737:chemistry / Q000235:genetics*; D006579:Heterozygote; D006801:Humans; D015810:Linkage Disequilibrium; D007986:Luteinizing Hormone / Q000737:chemistry / Q000235:genetics*; D008252:Macaca fascicularis / Q000235:genetics*; D008253:Macaca mulatta; D008322:Mammals; D008969:Molecular Sequence Data; D011110:Polymorphism, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D018076:DNA, Complementary; D011994:Recombinant Proteins; D007986:Luteinizing Hormone; D005640:Follicle Stimulating Hormone