B6129S-Lhcgr^tm1.1Pnara/J

Stock number: 029311
Product name: KiLHR^D582G
Research areas: Cancer Research, Developmental Biology Research, Endocrine Deficiency Research, Reproductive Biology Research, Research Tools

Description

The KiLHR^D582G knock-in allele has endogenous Lhcgr exon 11 replaced by a mutant encoding the constitutively-active D582G amino acid change analogous to the most common D578G mutation found in human boys with familial male-limited precocious puberty (FMPP). Heterozygous males exhibit several characteristics of the human disease (early puberty, Leydig cell hyperplasia and elevated testosterone). Heterozygotes females are also affected (early puberty, infertility, elevated steroid hormone levels, polycystic ovaries and granulosa cell tumors). The mice are useful for studying gonadotropin hormones and receptors in reproductive physiology and pathophysiology, as well as cancer.

Detailed description

The luteinizing hormone receptor (LHCGR), a member of the G protein-coupled receptor family, is essential for fertility in males and females, with implications for studying gonadotropin hormones and receptors in reproductive physiology and pathophysiology, as well as cancer.

The KiLHR^D582G knock-in allele has the constitutively-active D582G mutation in exon 11 of the mouse luteinizing hormone/choriogonadotropin receptor locus (Lhcgr); analogous to the most common D578G mutation found in human boys with familial male-limited precocious puberty (FMPP).

The phenotype described below is for mice maintained on a B6129SF1/J genetic background.
Similar to boys with FMPP, heterozygous (KiLHR^D582G/+) male mice exhibit precocious puberty (early puberty) by 15 days of age. Leydig cell hyperplasia and elevated testosterone (in the context of prepubertal [low] luteinizing hormone levels) is observed as early as 7 days of age and through adulthood (24 weeks old). In addition, the donating investigator reports that although heterozygous males have normal sperm count/motility, they show progressive infertility and fail to produce litters by ~5-6 months of age. Compared to wildtype males, KiLHR^D582G/+ males are more aggressive to other males, but KiLHR^D582G/+ males are not aggressive to breeder females.
While human females with the corresponding activating mutation exhibit no phenotypic abnormalities, female KiLHR^D582G/+ mice exhibit early puberty (~15 days old), infertility, elevated steroid hormone levels (testosterone, estradiol and progesterone), polycystic ovaries (as early as three weeks of age) and granulosa cell tumors (~50% by six months of age). The donating investigator reports no increased aggression in KiLHR^D582G/+ females.
Because of the phenotype, the donating investigator reports it can be difficult to maintain the colony. They use harem mating of a young heterozygous male (starting at seven weeks of age) to three units of B6129SF1/J female pairs; rotating the male to a new female pair every two weeks. It has not been attempted to make this strain homozygous to date (July 2016).

Development

The KiLHR^D582G knock-in allele was designed by Dr. Prema Narayan (Southern Illinois University). First, an exon 11 cDNA sequence from the mouse luteinizing hormone/choriogonadotropin receptor locus (Lhcgr) on chromosome 17 was altered via site-directed mutagenesis to change codon 582 from aspartic acid to glycine (D582G); this corresponds to the most common D578G mutation found in human boys with familial male-limited precocious puberty (FMPP). A targeting vector containing this mutant exon 11 and a downstream loxP::frt::PGK-neo::frt cassette was electroporated into (129X1/SvJ x 129S1/Sv)F1-Kitl⁺-derived R1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts. Chimeric males were bred with B6;SJL-Tg(ACTFLPe)9205Dym/J females (Stock No. 003800) for germline transmission and to excise the neomycin cassette. The resulting KiLHR^D582G colony was maintained by breeding B6129SF1/J females (from Stock No. 101043) with young heterozygous (KiLHR^D582G/+) males for ten generations. After that, B6129SF2/J females (equivalent to Stock No. 101045) were bred with young KiLHR^D582G/+ males for several generations. In 2016, heterozygous males (either black or agouti coat color) were sent to The Jackson Laboratory Repository. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize B6129SF1/J oocytes (Stock No. 101043).
Thereafter, our live colony will be maintained by breeding wildtype sibling females with heterozygous males (using males as early as seven weeks of age). Periodically, we will breed B6129SF1/J females (Stock No. 101043) with heterozygous males to maintain ~50/50% mix of C57BL/6 and 129 in our live colony.

Allele

Symbol: Lhcgr^tm1.1Pnara
Name: targeted mutation 1.1, Prema Narayan
MGI accession ID: MGI:5538310
Generation method: Targeted
Marker symbol: Lhcgr
Marker name: luteinizing hormone/choriogonadotropin receptor
Chromosome: 17
Strain of origin: (129X1/SvJ x 129S1/Sv)F1-Kitl⁺
Molecular note: Nucleotide substitutions (GAC to GGA) in exon 11 result in the amino acid substitution of glycine for aspartic acid at position 582 (D582G). Flp-mediated recombination removed the FRT-flanked neomycin resistance cassette.