The mitochondrion is involved in energy generation, apoptosis regulation, and calcium homeostasis. Mutations in genes involved in mitochondrial processes often result in a severe phenotype or embryonic lethality, making the study of mitochondrial involvement in aging, neurodegeneration, or reproduction challenging. Using a transgenic insertional mutagenesis strategy, we generated a mouse mutant, Immp2lTg(Tyr)979Ove, with a mutation in the inner mitochondrial membrane peptidase 2-like (Immp2l) gene. The mutation affected the signal peptide sequence processing of mitochondrial proteins cytochrome c1 and glycerol phosphate dehydrogenase 2. The inefficient processing of mitochondrial membrane proteins perturbed mitochondrial function so that mitochondria from mutant mice manifested hyperpolarization, higher than normal superoxide ion generation, and higher levels of ATP. Homozygous Immp2lTg(Tyr)979Ove females were infertile due to defects in folliculogenesis and ovulation, whereas mutant males were severely subfertile due to erectile dysfunction. The data suggest that the high superoxide ion levels lead to a decrease in the bioavailability of nitric oxide and an increase in reactive oxygen species stress, which underlies these reproductive defects. The results provide a novel link between mitochondrial dysfunction and infertility and suggest that superoxide ion targeting agents may prove useful for treating infertility in a subpopulation of infertile patients.

Estrogen receptors (ERα and ERβ) are clearly multifaceted in terms of structure and function. Several relatively abundant ERβ isoforms have been identified, which can be differentially expressed in various tissues. In order to provide insight into the possible role of the ERβ family in breast tissue a study of the putative functions of the human (h) ERβ1, hERβ2 and hERβ5 isoforms was undertaken. Only hERβ1 was found to bind ligand, which induced conformational changes as determined by protease digestion assays. All ERβ isoforms could bind to and bend DNA although the relative efficiency with which they bound DNA differed with hERα>hERβ1>hERβ2> >hERβ5. All ERβ isoforms inhibited ERα transcriptional activity on an estrogen-response element (ERE)-reporter gene. The relative activities were hERβ1>hERβ2 >hERβ5; however, only hERβ1 had transcriptional activity of its own. Both LY117018-hERα and LY117018-hERβ1 complexes alone could activate transcription on a TGF-β3-CAT gene. Although hERβ2 and hERβ5 had no activity alone, they inhibited ERα but not hERβ1 transcriptional activity of transforming growth factor (TGF)-β3-CAT. In marked contrast to activity on an ERE-CAT reporter gene, hERβ1 did not modulate ERα transcriptional activity on a TGF-β3-CAT reporter gene. These data support promoter-specific differential activities of hERβ isoforms with respect to models of ERα regulated gene expression, and suggest that they may have a role in differentially modulating estrogen action.

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Using transposon-mediated gene-trap mutagenesis, we have generated a novel mouse mutant termed Blad (Bloated Bladder). Homozygous mutant mice die perinatally showing a greatly distended bladder, underdeveloped diaphragm and a reduction in total skeletal muscle mass. Wild type and heterozygote mice appear normal. Using PCR, we identified a transposon insertion site in the first intron of Nmnat2 (Nicotinamide mononucleotide adenyltransferase 2). Nmnat2 is expressed predominantly in the brain and nervous system and has been linked to the survival of axons. Expression of this gene is undetectable in Nmnat2blad/blad mutants. Examination of the brains of E18.5 Nmnat2blad/blad mutant embryos did not reveal any obvious morphological changes. In contrast, E18.5 Nmnat2blad/blad homozygotes showed an approximate 60% reduction of spinal motoneurons in the lumbar region and a more than 80% reduction in the sensory neurons of the dorsal root ganglion (DRG). In addition, facial motoneuron numbers were severely reduced, and there was virtually a complete absence of axons in the hind limb. Our observations suggest that during embryogenesis, Nmnat2 plays an important role in axonal growth or maintenance. It appears that in the absence of Nmnat2, major target organs and tissues (e.g., muscle) are not functionally innervated resulting in perinatal lethality. In addition, neither Nmnat1 nor 3 can compensate for the loss of Nmnat2. Whilst there have been recent suggestions that Nmnat2 may be an endogenous modulator of axon integrity, this work represents the first in vivo study demonstrating that Nmnat2 is involved in axon development or survival in a mammal.