The TGF-β superfamily is the largest family of secreted proteins in mammals, and members of the TGF-β family are involved in most developmental and physiological processes. Growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), oocyte-secreted paralogs of the TGF-β superfamily, have been shown genetically to control ovarian physiology. Although previous studies found that GDF9 and BMP15 homodimers can modulate ovarian pathways in vitro, the functional species-specific significance of GDF9:BMP15 heterodimers remained unresolved. Therefore, we engineered and produced purified recombinant mouse and human GDF9 and BMP15 homodimers and GDF9:BMP15 heterodimers to compare their molecular characteristics and physiological functions. In mouse granulosa cell and cumulus cell expansion assays, mouse GDF9 and human BMP15 homodimers can up-regulate cumulus expansion-related genes (Ptx3, Has2, and Ptgs2) and promote cumulus expansion in vitro, whereas mouse BMP15 and human GDF9 homodimers are essentially inactive. However, we discovered that mouse GDF9:BMP15 heterodimer is ∼10-to 30-fold more biopotent than mouse GDF9 homodimer, and human GDF9:BMP15 heterodimer is ∼1,000-to 3,000-fold more bioactive than human BMP15 homodimer. We also demonstrate that the heterodimers require the kinase activities of ALK4/5/7 and BMPR2 to activate SMAD2/3 but unexpectedly need ALK6 as a coreceptor in the signaling complex in granulosa cells. Our findings that GDF9:BMP15 heterodimers are the most bioactive ligands in mice and humans compared with homodimers explain many puzzling genetic and physiological data generated during the last two decades and have important implications for improving female fertility in mammals.igands of the TGF-β superfamily, the largest family of secreted proteins in mammals, are synthesized as dimers and bind transmembrane type 1 and type 2 serine-threonine kinase receptors to activate downstream signaling cascades (e.g., the SMADs) in many developmental, physiological, and pathophysiological processes (1, 2). Growth differentiation factor 9 (GDF9) and bone morphogenic protein 15 (BMP15) are key oocyte-secreted members of the TGF-β superfamily and can regulate female fertility in several mammals (2, 3). Although GDF9 and BMP15 are closely related paralogs, they have been shown in vitro to signal through divergent SMAD2/3 and SMAD1/5/8 pathways, respectively (4-6).By studying gene knockouts and mutant models, putative roles of GDF9 and BMP15 in female reproduction have been described in mice, sheep, and humans. Our group previously discovered that Gdf9-null female mice are sterile (7), and Gdf9 +/− Bmp15 −/− double-mutant mice had more severe fertility defects than subfertile Bmp15 −/− mice (8, 9). BMP15 or GDF9 heterozygous mutant sheep have increased litter size, whereas homozygous mutants are sterile and phenocopy Gdf9 −/− mice (10, 11). In humans, mutations in GDF9 and BMP15 have been associated with premature ovarian failure and dizygotic twinning (12)(13)(14). These data suggest syner...

Summary Actively dividing cells perform robust and accurate DNA replication during fluctuating nutrient availability, yet factors that prevent disruption of replication remain largely unknown. Here we report that DksA, a nutrient-responsive transcription factor, ensures replication completion in Escherichia coli by removing transcription roadblocks. In the absence of DksA, replication is rapidly arrested upon amino acid starvation. This arrest requires active transcription, and is alleviated by RNA polymerase mutants that compensate for DksA activity. This replication arrest occurs independently of exogenous DNA damage, yet it induces the DNA damage response and recruits the main recombination protein RecA. This novel function of DksA is independent of its transcription initiation activity, but requires its less studied transcription elongation activity. Finally, GreA/B elongation factors also prevent replication arrest during nutrient stress. We conclude that transcription elongation factors alleviate fundamental conflicts between replication and transcription, thereby protecting replication fork progression and DNA integrity.

Organogenesis of the ovary is a highly orchestrated process involving multiple lineage determinations of ovarian surface epithelium, granulosa cells, and theca cells. While the sources of ovarian surface epithelium and granulosa cells are known, the origin(s) of theca progenitor cells have not been definitively identified. Here we show that theca cells derive from two sources: Wt1+ cells indigenous to the ovary and Gli1+ mesenchymal cells migrated from the mesonephros. These progenitors acquire theca lineage marker Gli1 in response to paracrine signals Desert hedgehog (Dhh) and Indian hedgehog (Ihh) from granulosa cells. Ovaries lacking Dhh/Ihh exhibit theca layer loss, blunted steroid production, arrested folliculogenesis, and failure to form corpora lutea. Production of Dhh/Ihh in granulosa cells requires Growth differentiation factor 9 (GDF9) from the oocyte. Our studies provide the first genetic evidence for the origins of theca cells and reveal a multicellular interaction critical for the formation of a functional theca.

The stem cell niche is a specialized environment that dictates stem cell function during development and homeostasis. We show that Dll1, a Notch pathway ligand, is enriched in mammary gland stem cells (MaSCs) and mediates critical interactions with stromal macrophages in the surrounding niche in mouse models. Conditional deletion of Dll1 reduced the number of MaSCs and impaired ductal morphogenesis in the mammary gland. Moreover, MaSC-expressed Dll1 activates Notch signaling in stromal macrophages, increasing their expression of Wnt family ligands such as Wnt3, Wnt10A, and Wnt16, thereby initiating a feedback loop that promotes the function of Dll1-expressing MaSCs. Together, these findings reveal functionally important cross-talk between MaSCs and their macrophageal niche through Dll1-mediated Notch signaling.

Coordinated regulation of oocyte and ovarian follicular development is essential for fertility. In particular, the progression of meiosis, a germ cell-specific cell division that reduces the number of chromosomes from diploid to haploid, must be arrested until just before ovulation. Follicular somatic cells are well-known to impose this arrest, which is essential for oocyte-follicle developmental synchrony. Follicular somatic cells sustain meiotic arrest via the natriuretic peptide C/natriuretic peptide receptor 2 (NPPC/ NPR2) system, and possibly also via high levels of the purine hypoxanthine in the follicular fluid. Upon activation by the ligand NPPC, NPR2, the predominant guanylyl cyclase in follicular somatic cells, produces cyclic guanosine monophosphate (cGMP), which maintains meiotic arrest after transfer to the oocyte via gap junctions. Here we report that both the NPPC/NPR2 system and hypoxanthine require the activity of inosine monophosphate dehydrogenase (IMPDH), the rate-limiting enzyme required for the production of guanylyl metabolites and cGMP. Furthermore, oocyte-derived paracrine factors, particularly the growth differentiation factor 9-bone morphogenetic protein 15 heterodimer, promote expression of Impdh and Npr2 and elevate cGMP levels in cumulus cells. Thus, although the somatic compartment of ovarian follicles plays an essential role in the maintenance of oocyte meiotic arrest, as has been known for many years, this function of the somatic cells is surprisingly regulated by signals from the oocyte itself. F ertility in mammals depends on the coordinated development of ovarian follicles and the oocytes contained within them. The ovulation of oocytes by Graafian follicles must coincide with oocyte meiotic progression. Meiosis is a germ cell-specific cell division that reduces the number of chromosomes from diploid to haploid. In mammals, oocytes are arrested at the diplotene stage (diploid) of meiosis until the surge of luteinizing hormone (LH) from the pituitary gland initiates the ovulatory process. The LH surge initiates the resumption of meiosis and its progression to the metaphase of the second meiotic division, and these haploid oocytes (eggs) are ovulated into the oviduct to await fertilization. Ever since the classic experiments of Pincus and Enzmann (1) in the 1930s, it has been widely accepted that follicular somatic cells of Graafian follicles maintain the fully grown oocytes in meiotic arrest until the LH surge: Removing them from follicles for culture in a supportive medium before the surge results in an LH-independent resumption of meiosis. Thus, isolated oocytes resume meiosis simply because they have been separated from the inhibitory effect of follicular somatic cells. The mechanisms by which these cells maintain oocyte meiotic arrest have been the subject of numerous studies over the last 75 y.Oocytes arrested at the diplotene stage are referred to as germinal vesicle (GV)-stage oocytes, and the dissolution of the oocyte nuclear envelope, often referred to as germinal vesic...