Human and mouse essentiality screens as a resource for disease gene discovery.

The identification of causal variants in sequencing studies remains a considerable challenge that can be partially addressed by new gene-specific knowledge. Here, we integrate measures of how essential a gene is to supporting life, as inferred from viability and phenotyping screens performed on knockout mice by the International Mouse Phenotyping Consortium and essentiality screens carried out on human cell lines. We propose a cross-species gene classification across the Full Spectrum of Intolerance to Loss-of-function (FUSIL) and demonstrate that genes in five mutually exclusive FUSIL categories have differing biological properties. Most notably, Mendelian disease genes, particularly those associated with developmental disorders, are highly overrepresented among genes non-essential for cell survival but required for organism development. After screening developmental disorder cases from three independent disease sequencing consortia, we identify potentially pathogenic variants in genes not previously associated with rare diseases. We therefore propose FUSIL as an efficient approach for disease gene discovery

Mitochondria, redox signaling and axis specification in metazoan embryos.

Mitochondria are not only the major energy generators of the eukaryotic cell but they are also sources of signals that control gene expression and cell fate. While mitochondria are often asymmetrically distributed in early embryos, little is known about how they contribute to axial patterning. Here we review studies of mitochondrial distribution in metazoan eggs and embryos and the mechanisms of redox signaling, and speculate on the role that mitochondrial anisotropies might play in the developmental specification of cell fate during embryogenesis of sea urchins and other animals

Oocyte-dependent activation of mitogen-activated protein kinase (ERK1/2) in cumulus cells is required for the maturation of the mouse oocyte-cumulus cell complex.

Luteinizing hormone (LH) induces maturational processes in oocyte-cumulus cell complexes (OCC) of preovulatory follicles that include both resumption of meiosis in the oocyte and expansion (mucification) of the cumulus oophorus. Both processes require activation of mitogen-activated protein kinase (MAPK) in granulosa cells. Here, it is reported that inhibition of MAPK activation prevented gonadotropin-stimulated resumption of meiosis as well as the rise in expression of two genes whose products are necessary for normal cumulus expansion, Has2 and Ptgs2. However, inhibition of MAPK did not block gonadotropin-induced elevation of granulosa cell cAMP, indicating that the activation of MAPK required for inducing GVB and cumulus expansion is downstream of cAMP. Moreover, activation of MAPK in cumulus cells requires one or more paracrine factors from the oocyte to induce GVB and cumulus expansion; MAPK activation alone is not sufficient to initiate these maturational processes. This study demonstrates a remarkable interaction between the oocyte and cumulus cells that is essential for gonadotropin-induced maturational processes in OCC. By enabling gonadotropin-dependent MAPK activation in granulosa cells, oocytes promote the generation of a return signal from these cells that induces the resumption of meiosis. It also appears that an oocyte-dependent pathway downstream from oocyte-enabled activation of MAPK, and distinct from that promoting the resumption of meiosis, governs cumulus expansion

Synergistic roles of BMP15 and GDF9 in the development and function of the oocyte-cumulus cell complex in mice: genetic evidence for an oocyte-granulosa cell regulatory loop.

Bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9) are oocyte-specific growth factors that appear to play key roles in granulosa cell development and fertility in most mammalian species. We have evaluated the role(s) of these paracrine factors in the development and function of both the cumulus cells and oocytes by assessing cumulus expansion, oocyte maturation, fertilization, and preimplantation embryogenesis in Gdf9+/-Bmp15-/- [hereafter, double mutant (DM)] mice. We found that cumulus expansion, as well as the expression of hyaluronon synthase 2 (Has2) mRNA was impaired in DM oocyte-cumulus cell complexes. This aberrant cumulus expansion was not remedied by coculture with normal wild-type (WT) oocytes, indicating that the development and/or differentiation of cumulus cells in the DM, up to the stage of the preovulatory luteinizing hormone (LH) surge, is impaired. In addition, DM oocytes failed to enable FSH to induce cumulus expansion in WT oocytectomized (OOX) cumulus. Moreover, LH-induced oocyte meiotic resumption was significantly delayed in vivo, and this delayed resumption of meiosis was correlated with the reduced activation of mitogen-activated protein kinase (MAPK) in the cumulus cells, thus suggesting that GDF9 and BMP15 also regulate the function of cumulus cells after the preovulatory LH surge. Although spontaneous in vitro oocyte maturation occurred normally, oocyte fertilization and preimplantation embryogenesis were significantly altered in the DM, suggesting that the full complement of both GDF9 and BMP15 are essential for the development and function of oocytes. Because receptors for GDF9 and BMP15 have not yet been identified in mouse oocytes, the effects of the mutations in the Bmp15 and Gdf9 genes on oocyte development and functions must be produced indirectly by first affecting the granulosa cells and then the oocyte. Therefore, this study provides further evidence for the existence and functioning of an oocyte-granulosa cell regulatory loop

CRB1 is essential for external limiting membrane integrity and photoreceptor morphogenesis in the mammalian retina.

Mutations within the CRB1 gene have been shown to cause human retinal diseases including retinitis pigmentosa and Leber congenital amaurosis. We have recently identified a mouse model, retinal degeneration 8 (rd8) with a single base deletion in the Crb1 gene. This mutation is predicted to cause a frame shift and premature stop codon which truncates the transmembrane and cytoplasmic domain of CRB1. Like in Drosophila crumbs (crb) mutants, staining for adherens junction proteins known to localize to the external limiting membrane, the equivalent of the zonula adherens in the mammalian retina, is discontinuous and fragmented. Shortened photoreceptor inner and outer segments are observed as early as 2 weeks after birth, suggesting a developmental defect in these structures rather than a degenerative process. Photoreceptor degeneration is observed only within regions of retinal spotting, which is seen predominantly in the inferior nasal quadrant of the eye, and is caused by retinal folds and pseudorosettes. Photoreceptor dysplasia and degeneration in Crb1 mutants strongly vary with genetic background, suggesting that the variability in phenotypes of human patients that carry mutations in CRB1 may be due to interactions with background modifiers in addition to allelic variations. The Crb1rd8 mouse model will facilitate the analysis of Crb1 function in the neural retina and the identification of interacting factors as candidate retinal disease genes

A Gene Implicated in Activation of Retinoic Acid Receptor Targets Is a Novel Renal Agenesis Gene in Humans.

Renal agenesis (RA) is one of the more extreme examples of congenital anomalies of the kidney and urinary tract (CAKUT). Bilateral renal agenesis is almost invariably fatal at birth, and unilateral renal agenesis can lead to future health issues including end-stage renal disease. Genetic investigations have identified several gene variants that cause RA, including EYA1, LHX1, and WT1 However, whereas compound null mutations of genes encoding α and γ retinoic acid receptors (RARs) cause RA in mice, to date there have been no reports of variants in RAR genes causing RA in humans. In this study, we carried out whole exome sequence analysis of two families showing inheritance of an RA phenotype, and in both identified a single candidate gene, GREB1L Analysis of a zebrafish greb1l loss-of-function mutant revealed defects in the pronephric kidney just prior to death, and F0 CRISPR/Cas9 mutagenesis of Greb1l in the mouse revealed kidney agenesis phenotypes, implicating Greb1l in this disorder. GREB1L resides in a chromatin complex with RAR members, and our data implicate GREB1L as a coactivator for RARs. This study is the first to associate a component of the RAR pathway with renal agenesis in humans. Genetics 2017 Sep; 207(1):215-228

Point mutations in murine Nkx2-5 phenocopy human congenital heart disease and induce pathogenic Wnt signaling

Mutations in the Nkx2-5 gene are a main cause of congenital heart disease. Several studies have addressed the phenotypic consequences of disrupting the Nkx2-5 gene locus, although animal models to date failed to recapitulate the full spectrum of the human disease. Here, we describe a new Nkx2-5 point mutation murine model, akin to its human counterpart disease–generating mutation. Our model fully reproduces the morphological and physiological clinical presentations of the disease and reveals an understudied aspect of Nkx2-5–driven pathology, a primary right ventricular dysfunction. We further describe the molecular consequences of disrupting the transcriptional network regulated by Nkx2-5 in the heart and show that Nkx2-5–dependent perturbation of the Wnt signaling pathway promotes heart dysfunction through alteration of cardiomyocyte metabolism. Our data provide mechanistic insights on how Nkx2-5 regulates heart function and metabolism, a link in the study of congenital heart disease, and confirms that our models are the first murine genetic models to our knowledge to present all spectra of clinically relevant adult congenital heart disease phenotypes generated by NKX2-5 mutations in patients