Mammalian fertilization is dependent upon a series of bicarbonate-induced, cAMP-dependent processes sperm undergo as they "capacitate," i.e., acquire the ability to fertilize eggs. Male mice lacking the bicarbonate- and calcium-responsive soluble adenylyl cyclase (sAC), the predominant source of cAMP in male germ cells, are infertile, as the sperm are immotile. Membrane-permeable cAMP analogs are reported to rescue the motility defect, but we now show that these "rescued" null sperm were not hyperactive, displayed flagellar angulation, and remained unable to fertilize eggs in vitro. These deficits uncover a requirement for sAC during spermatogenesis and/or epididymal maturation and reveal limitations inherent in studying sAC function using knockout mice. To circumvent this restriction, we identified a specific sAC inhibitor that allowed temporal control over sAC activity. This inhibitor revealed that capacitation is defined by separable events: induction of protein tyrosine phosphorylation and motility are sAC dependent while acrosomal exocytosis is not dependent on sAC.

Although it is well established that the circadian clock regulates mammalian reproductive physiology, the molecular mechanisms by which this regulation occurs are not clear. The authors investigated the reproductive capacity of mice lacking Bmal1 (Arntl, Mop3), one of the central circadian clock genes. They found that both male and female Bmal1 knockout (KO) mice are infertile. Gross and microscopic inspection of the reproductive anatomy of both sexes suggested deficiencies in steroidogenesis. Male Bmal1 KO mice had low testosterone and high luteinizing hormone serum concentrations, suggesting a defect in testicular Leydig cells. Importantly, Leydig cells rhythmically express BMAL1 protein, suggesting peripheral control of testosterone production by this clock protein. Expression of steroidogenic genes was reduced in testes and other steroidogenic tissues of Bmal1 KO mice. In particular, expression of the steroidogenic acute regulatory protein (StAR) gene and protein, which regulates the rate-limiting step of steroidogenesis, was decreased in testes from Bmal1 KO mice. A direct effect of BMAL1 on StAR expression in Leydig cells was indicated by in vitro experiments showing enhancement of StAR transcription by BMAL1. Other hormonal defects in male Bmal1 KO mice suggest that BMAL1 also has functions in reproductive physiology outside of the testis. These results enhance understanding of how the circadian clock regulates reproduction.Keywords circadian rhythms; fertility; testosterone; testes; sperm; StAR; mice Disruption of circadian rhythms results in a variety of pathophysiologic states (Hastings et al., 2003). Reproductive physiology, in particular, is profoundly influenced by circadian rhythms (Boden and Kennaway, 2006). In various insect species, the circadian clock is necessary for proper ovulation, sperm production, and fertility (Giebultowicz et al., 1989;Beaver et al., 2002;Beaver et al., 2003;Beaver and Giebultowicz, 2004 (Lucas and Eleftheriou, 1980;Clair et al., 1985;Chappell et al., 2003;Miller et al., 2004). For example, the surge of luteinizing hormone (LH) necessary for ovulation in rodents, which occurs at the same time of day during each estrous cycle, requires a functional circadian clock (Barbacka-Surowiak et al., 2003). In addition, at the onset of puberty, a clear diurnal rhythm of gonadotropin serum levels is established in both mice and humans (Andrews and Ojeda, 1981;Jean-Faucher et al., 1986;Dunkel et al., 1992;Apter et al., 1993). It is unclear whether this diurnal rhythm continues into adulthood, but testosterone serum concentration shows daily oscillations in adult male mice and humans (Lucas and Eleftheriou, 1980;Clair et al., 1985). Although the association between circadian rhythms and testosterone is a long-established phenomenon, the molecular mechanisms by which the circadian clock regulates testosterone production are unknown.The circadian clock is based on a transcription translation feedback loop that results in the cyclic expression of genes and proteins over a 24-h p...

Many procedures used in assisted reproductive technologies (ART) to treat human infertility entail culture of preimplantation embryos. Moreover, there is an increasing trend to culture embryos for longer periods of time before uterine transfer to identify the ''best'' embryos for transfer and to minimize multiple pregnancies. Embryo culture, however, can perturb embryo metabolism and gene expression, and the long-term consequences of culture are unknown. We have explored the behavioral consequences of embryo culture by using a 129S6͞SvEvTac͞C57BL͞6J F 1 mouse model and find that adults derived from cultured embryos exhibit specific behavioral alterations in the elevated zero maze and Morris water maze tasks.

Summary The master transcription factors play integral roles in the pluripotency transcription circuitry of embryonic stem cells (ESCs). How they selectively activate expression of the pluripotency network while simultaneously repressing genes involved in differentiation is not fully understood. Here we define a requirement for the INO80 complex, a SWI/SNF family chromatin remodeler, in ESC self-renewal, somatic cell reprogramming, and blastocyst development. We show that Ino80, the chromatin remodeling ATPase, co-occupies pluripotency gene promoters with the master transcription factors, and its occupancy is dependent on Oct4 and Wdr5. At the pluripotency genes, Ino80 maintains open chromatin architecture and licenses recruitment of Mediator and RNA Polymerase II for gene activation. Our data reveal an essential role for INO80 in the expression of the pluripotency network, and illustrate the coordination among chromatin remodeler, transcription factor, and histone modifying enzyme in the regulation of the pluripotent state.

Osteoporosis is characterized by low bone density, and osteopenia is responsible for 1.5 million fractures in the United States annually. 1 In order to identify regions of the genome which are likely to contain genes predisposing to osteopenia, we genotyped 149 members of seven large pedigrees having recurrence of low bone mineral density (BMD) with 330 DNA markers spread throughout the autosomal genome. Linkage analysis for this quantitative trait was carried out using spine and hip BMD values by the classical lod-score method using a genetic model with parameters estimated from the seven families. In addition, non-parametric analysis was performed using the traditional Haseman-Elston approach in 74 independent sib pairs from the same pedigrees. The maximum lod score obtained by parametric analysis in all families combined was + 2.08 (θ = 0.05) for the marker CD3D on chromosome 11q. All other combined lod scores from the parametric analysis were less than + 1.90, the threshold for suggestive linkage. Non-parametric analysis suggested linkage of low BMD to chromosomes 1p36 (Z max = + 3.51 for D1S450) and 2p23-24 (Z max = + 2.07 for D2S149). Maximum multi-point lod scores for these regions were + 2.29 and + 2.25, respectively. A third region with associated lod scores above the threshold of suggestive linkage in both singlepoint and multi-point non-parametric analysis was on chromosome 4qter (Z max = + 2.95 for D4S1539 and Z max = + 2.48 for D4S1554). Our data suggest the existence of multiple genes involved in controlling spine and hip BMD, and indicate several candidate regions for further screening in this and other independent samples.