Biomedical subjects

R R Behringer

Publications and source records attributed to R R Behringer.

At least 73 records · Page 4Linked to original sources

PubMed1996

Targeted deletion in astrocyte intermediate filament (Gfap) alters neuronal physiology.

Glial fibrillary acidic protein (GFAP) is a member of the family of intermediate filament structural proteins and is found predominantly in astrocytes of the central nervous system (CNS). To assess the function of GFAP, we created GFAP-null mice using gene targeting in embryonic stem cells. The GFAP-null mice have normal development and fertility, and show no gross alterations in behavior or CNS morphology. Astrocytes are present in the CNS of the mutant mice, but contain a severely reduced number of intermediate filaments. Since astrocyte processes contact synapses and may modulate synaptic function, we examined whether the GFAP-null mice were altered in long-term potentiation in the CA1 region of the hippocampus. The GFAP-null mice displayed enhanced long-term potentiation of both population spike amplitude and excitatory post-synaptic potential slope compared to control mice. These data suggest that GFAP is important for astrocyte-neuronal interactions, and that astrocyte processes play a vital role in modulating synaptic efficacy in the CNS. These mice therefore represent a direct demonstration that a primary defect in astrocytes influences neuronal physiology.

Animals↗

PubMed1996

Analysis of the role of Amh and Fra1 in the Sry regulatory pathway.

Sry is the Y-chromosomal gene responsible for initiating the pathway of male development in mammals, presumably by regulating downstream target genes. As a basis for examining the role played by Sry and other candidate sex-determining genes, we have used reverse-transcriptase PCR to produce a precise temporal profile of expression of Sry in the developing gonad. Sry expression first occurs at 10.5 days postcoitum (dpc) and is upregulated to reach a maximum level from 11.25 to 12 dpc. Expression is then steadily downregulated from 12.25 to 13.25 dpc and is completely extinguished by 13.5 dpc. Two genes implicated in sexual development are Amh, encoding anti-müllerian hormone (AMH), and Fra1, encoding Fos-related antigen-1. Expression profiles of Amh and Sry in fetal gonads indicate a possible role for AMH in the downregulation of Sry expression. We show, however, that the timing of Sry downregulation is unaffected in Amh-deficient mice, excluding a role for AMH as a negative regulator of Sry. Further, we have examined the possibility that SRY may activate Fra1 during testis determination by analysing the expression of Fra1 in mouse fetal gonads. Fra1 is not expressed at any stage at or around the critical period of sex determination when Sry transcripts are present, thus excluding a role for Fra1 in sex determination and differentiation.

Animals↗

PubMed1996

Prenatal folic acid treatment suppresses acrania and meroanencephaly in mice mutant for the Cart1 homeobox gene.

The paired-class homeobox-containing gene, Cart1, is expressed in forebrain mesenchyme, branchial arches, limb buds and cartilages during embryogenesis. Here, we show that Cart1-homozygous mutant mice are born alive with acrania and meroanencephaly but die soon after birth-a phenotype that strikingly resembles a corresponding human syndrome caused by a neural tube closure defect. Developmental studies suggest that Cart1 is required for forebrain mesenchyme survival and that its absence disrupts cranial neural tube morphogenesis by blocking the initiation of closure in the midbrain region that ultimately leads to the generation of lethal craniofacial defects. Prenatal treatment of Cart1 homozygous mutants with folic acid suppresses the development of the acrania/meroanencephaly phenotype.

Anencephaly↗

PubMed1996

Transgenic models to study the roles of inhibins and activins in reproduction, oncogenesis, and development.

With the advent of gene targeting in pluripotent mouse embryonic stem cells, it is now possible to modify the mammalian genome to generate mutant strains of mice with precise genetic mutations. The major goal of my laboratory is to generate transgenic mice to use as physiologic models to study mammalian reproduction and development. The initial focus of our research has been to generate mice deficient in inhibins, activins, activin binding proteins (i.e., follistatin), and activin receptors (i.e., activin receptor type II) to understand their interactions and roles in the hypothalamic-pituitary-gonadal axis and mammalian development. Inhibins and activins, dimeric members of the TGF-beta superfamily, were discovered due to their role in pituitary follicle stimulating hormone homeostasis. However, these proteins have later been shown to have diverse endocrine, paracrine, and autocrine functions. Activins have been shown to mediate their signals through type I and type II serine/threonine kinase receptors. The high interspecies conservation of activins, inhibins, and activin receptors and the universal presence of activins in mammals, birds, amphibians, and fish suggest an evolutionarily conserved role of these proteins in animal development. Our initial studies have demonstrated a tumor suppressor role of inhibin in the gonads and adrenals and have also suggested a role of activins in cancer cachexia-like syndrome. To further study the gonadal tumor development and the cancer cachexia-like syndrome in these mice, we have begun to generate mice with multiple genetic alterations (e.g., mice deficient in both inhibin and Mullerian inhibiting substance). We have also generated mice deficient in other components of this complex system (e.g., activin beta A, activin receptor type II, follistatin). Analysis of these transgenic mutant models has aided our overall understanding of the critical roles these proteins play in the development of the reproductive system, in the modulation of the endocrine milieu that regulates reproductive function, and in mammalian development.

Activins↗

PubMed1995

Bmpr encodes a type I bone morphogenetic protein receptor that is essential for gastrulation during mouse embryogenesis.

Bone morphogenetic proteins (BMPs) are secreted proteins that interact with cell-surface receptors and are believed to play a variety of important roles during vertebrate embryogenesis. Bmpr, also known as ALK-3 and Brk-1, encodes a type I transforming growth factor-beta (TGF-beta) family receptor for BMP-2 and BMP-4. Bmpr is expressed ubiquitously during early mouse embryogenesis and in most adult mouse tissues. To study the function of Bmpr during mammalian development, we generated Bmpr-mutant mice. After embryonic day 9.5 (E9.5), no homozygous mutants were recovered from heterozygote matings. Homozygous mutants with morphological defects were first detected at E7.0 and were smaller than normal. Morphological and molecular examination demonstrated that no mesoderm had formed in the mutant embryos. The growth characteristics of homozygous mutant blastocysts cultured in vitro were indistinguishable from those of controls; however, embryonic ectoderm (epiblast) cell proliferation was reduced in all homozygous mutants at E6.5 before morphological abnormalities had become prominent. Teratomas arising from E7.0 mutant embryos contained derivatives from all three germ layers but were smaller and gave rise to fewer mesodermal cell types, such as muscle and cartilage, than controls. These results suggest that signaling through this type I BMP-2/4 receptor is not necessary for preimplantation or for initial postimplantation development but may be essential for the inductive events that lead to the formation of mesoderm during gastrulation and later for the differentiation of a subset of mesodermal cell types.

Alleles↗

PubMed1995

The müllerian inhibitor and mammalian sexual development.

The elegant embryological experiments of Jost demonstrated the existence of a foetal testicular factor that is required to cause the regression of the müllerian duct system, the anlagen of the uterus, oviducts and upper portion of the vagina, during male sexual development. The müllerian inhibitor currently known as müllerian-inhibiting substance (MIS) or anti-müllerian hormone (AMH), is a member of the transforming growth factor-beta (TGF-beta) family of growth and differentiation factors. The genetic manipulation of the mouse germline has lead to the generation of animal models for MIS function. Female transgenic mice that chronically express MIS during embryogenesis are born without a uterus or oviducts and their ovaries lose germ cells and degenerate, recapitulating the phenotype of the bovine freemartin. Some male transgenic mice from very high MIS-expressing lines are feminized, suggesting alterations in androgen biosynthesis. Male mice homozygous for a targeted mutation of the MIS gene develop as male pseudohermaphrodites with both male (testes and Wolffian duct-derived) and female (müllerian duct-derived) reproductive organs. Most are infertile because the development of two reproductive systems physically blocks the exit of sperm from these males. In addition, Leydig cell hyperplasia is detected in a proportion of these males and in one case a Leydig cell tumour was found. Recently, a gene encoding a TGF-beta family type II Ser/Thr kinase membrane-bound receptor has been isolated that is expressed in both male and female gonads and in the mesenchyme surrounding the müllerian ducts during embryogenesis. These findings suggest that MIS-mediated müllerian duct regression occurs indirectly through mesenchymal tissue. A targeted mutation of this receptor has been established in the mouse germline. Mice homozygous for this receptor mutation should be useful in understanding the MIS signalling pathway for müllerian duct regression and gonadal function.

Animals↗

PubMed1995

Compound mutants for the paralogous hoxa-4, hoxb-4, and hoxd-4 genes show more complete homeotic transformations and a dose-dependent increase in the number of vertebrae transformed.

The Hox gene products are transcription factors involved in specifying regional identity along the anteroposterior body axis. In the mouse, several single mutants for Hox genes show variably penetrant, partial homeotic transformations of vertebrae at their anterior limits of expression, suggesting that compound Hox mutants might show more complete transformations with greater penetrance than the single Hox mutants. Compound mutants for the paralogous group 3 genes, hoxa-3 and hoxd-3, show deletion of a cervical vertebrae, which is not readily interpretable in terms of an alteration in regional identity. Here, we report the skeletal phenotypes of compound mutants in the group 4 Hox genes, hoxa-4, hoxb-4, and hoxd-4. Mice mutant for each of these genes were intercrossed to generate the three possible double mutant combinations and the triple mutant. In contrast to the hoxa-3, hoxd-3 double mutants, group 4 Hox compound mutants displayed clear alterations in regional identity, including a nearly complete transformation of the second cervical vertebrae toward the morphology of the first cervical vertebra in one double mutant combination. In comparing the types of homeotic transformations observed, different double mutant combinations showed different degrees of synergism. These results suggest a certain degree of functional redundancy among paralogous genes in specifying regional identity. Furthermore, there was a remarkable dose-dependent increase in the number of vertebrae transformed to a first cervical vertebra identity, including the second through the fifth cervical vertebrae in the triple mutant. Thus, these genes are required in a larger anteroposterior domain than is revealed by the single mutant phenotypes alone, such that multiple mutations in these genes result in transformations of vertebrae that are not at their anterior limit of expression.

Animals↗

PubMed1995

Inactivation of the myogenic bHLH gene MRF4 results in up-regulation of myogenin and rib anomalies.

The myogenic basic helix-loop-helix (bHLH) proteins MyoD, myf5, myogenin, and MRF4 can initiate myogenesis when expressed in nonmuscle cells. During embryogenesis, each of the myogenic bHLH genes is expressed in a unique temporospatial pattern within the skeletal muscle lineage, suggesting that they play distinct roles in muscle development. Gene targeting has shown that MyoD and myf5 play partially redundant roles in the genesis of myoblasts, whereas myogenin is required for terminal differentiation. MRF4 is expressed transiently in the somite myotome during embryogenesis and then becomes up-regulated during late fetal development to eventually become the predominant myogenic bHLH factor expressed in adult skeletal muscle. On the basis of its expression pattern, it has been proposed that MRF4 may regulate skeletal muscle maturation and aspects of adult myogenesis. To determine the function of MRF4, we generated mice carrying a homozygous germ-line mutation in the MRF4 gene. These mice showed only a subtle reduction in expression of a subset of muscle-specific genes but showed a dramatic increase in expression of myogenin, suggesting that it may compensate for the absence of MRF4 and demonstrating that MRF4 is required for the down-regulation of myogenin expression that normally occurs in postnatal skeletal muscle. Paradoxically, MRF4-null mice exhibited multiple rib anomalies, including extensive bifurcations, fusions, and supernumerary processes. These results demonstrate an unanticipated regulatory relationship between myogenin and MRF4 and suggest that MRF4 influences rib outgrowth through an indirect mechanism.

Animals↗

PubMed1995

Requirement for Lim1 in head-organizer function.

Lim1 is a homeobox gene expressed in the organizer region of mouse embryos. To investigate the role of Lim1 during embryogenesis, a targeted deletion of the Lim1 gene was generated in embryonic stem cells. Embryos homozygous for the null allele lacked anterior head structures but the remaining body axis developed normally. A partial secondary axis developed anteriorly in some mutant embryos. Lim1 is thus an essential regulator of the vertebrate head organizer.

Animals↗

PubMed1995

twist is required in head mesenchyme for cranial neural tube morphogenesis.

To understand the role of twist during mammalian development, we generated twist-null mice. twist-null embryos died at embryonic day 11.5. Their most prominent phenotype was a failure of the cranial neural folds to fuse. Mutant embryos also had defects in head mesenchyme, somites, and limb buds. Chimera analysis suggested that head mesenchyme was required for cranial neural tube closure and that twist acted in a cell-autonomous manner in this tissue. In addition, in the head mesenchyme region of chimeras, twist-null cells were segregated from wild-type cells, and in the forebrain they lacked mesenchymal characteristics. These results suggest that twist regulates the cellular phenotype and behavior of head mesenchyme cells that are essential for the subsequent formation of the cranial neural tube.

Animals↗

PubMed1995

Genomic organization and chromosomal location of the mouse type I BMP-2/4 receptor.

We have characterized the structure of a mouse bone morphogenetic protein (BMP) type I receptor gene that can bind both BMP-2 and BMP-4. The mouse BMP-2/4 receptor gene is encoded by 11 exons and spans approximately 38-kb. Most of the intron/exon boundaries are not conserved compared to the kinase domain of the related, activin type II receptor. In addition, whereas the activin type II receptor gene contains large introns (> 40 kb), the largest intron of the BMP-2/4 receptor gene is only 6.4-kb. The BMP-2/4 receptor gene (Bmpr) was mapped to mouse chromosome 14. Bmpr is closely linked to Rbp3 in the region containing pugnose, a mutation that alters bone development. Knowledge of the genomic structure of Bmpr provides important information to create Bmpr-deficient mice.

Amino Acid Sequence↗

PubMed1995

Mutations in paralogous Hox genes result in overlapping homeotic transformations of the axial skeleton: evidence for unique and redundant function.

Hoxd-4 (previously known as Hox-4.2 and -5.1) is a mouse homeobox-containing gene homologous to the Drosophila homeotic gene Deformed. During embryogenesis, Hoxd-4 is expressed in the presumptive hindbrain and spinal cord, prevertebrae, and other tissues. In the adult, Hoxd-4 transcripts are expressed predominantly in the testis and kidney, and to a lesser extent in intestine and heart. To understand the role of Hoxd-4 during mouse embryogenesis, we generated Hoxd-4 mutant mice. Mice heterozygous or homozygous for the Hoxd-4 mutation exhibit homeotic transformations of the second cervical vertebrae (C2) to the first cervical vertebrae (C1) and malformations of the neural arches of C1 to C3 and of the basioccipital bone. The phenotype was incompletely penetrant and showed variable expressivity on both an F2 hybrid and 129 inbred genetic background. The mutant phenotype was detected in the cartilaginous skeleton of 14.5-day (E14.5) mutant embryos but no apparent differences were detected in the somites of E9.5 mutant embryos, suggesting that the abnormalities develop after E9.5 perhaps during or after resegmentation of the somites to form the prevertebrae. These results suggest that Hoxd-4 plays a role in conferring position information along the anteroposterior axis in the skeleton. The phenotypic similarities and differences between Hoxd-4 and previously reported Hoxa-4 and Hoxb-4 mutant mice suggest that Hox gene paralogs have both redundant and unique functions.

Animals↗

PubMed1995

An insertional mutation in the BTF3 transcription factor gene leads to an early postimplantation lethality in mice.

The gene that encodes the general transcription factor known as BTF3 was disrupted in mouse embryonic stem cells in a random mutagenesis screen for developmental genes with the ROSA beta-geo (Friedrich and Soriano, 1991) retroviral gene trap vector. The BTF3 mutation was transmitted through the germline of chimaeric mice. While the endogenous BTF3 gene is ubiquitously expressed, the expression pattern of the beta-galactosidase reporter gene present in the gene trap vector in BTF3 heterozygotes was restricted. Mice homozygous for the mutant allele died soon after implantation, around embryonic day 6.5. Thus, BTF3 is essential for postimplantation development. The isolation of the BTF3 sequences in this ROSA beta-geo insertion was facilitated by a relatively simple single lacZ primer reverse transcription PCR strategy.

Animals↗

PubMed1995

Ontogeny of reproductive abnormalities induced by deregulation of anti-müllerian hormone expression in transgenic mice.

Anti-müllerian hormone, normally responsible for the regression of müllerian ducts in male fetuses, induces stunting, germ cell loss, and seminiferous tubule formation in ovaries of bovine freemartin fetuses and of transgenic mice, which express the human anti müllerian hormone gene under the control of the metallothionein promoter. Because the latter have been studied only after birth, we undertook a detailed chronological study of their reproductive organs. Müllerian ducts of transgenic female fetuses regressed at the same time as those of normal or transgenic males. Maximal reduction of germ cell number occurred between 16 days postcoitus and birth, when most transgenic oocytes were still in the leptotene stage of the meiotic prophase, whereas normal oocytes had already reached the pachytene phase. Interference with progression of the meiotic prophase and germ cell loss in the fetal ovary are probably responsible for subsequent ovarian regression and retardation of follicle growth. Seminiferous tubule formation was not detectable prior to birth and occurred only rarely in postnatal ovaries. Aromatase activity of fetal transgenic ovaries was decreased, as well as serum concentration of testosterone in adult transgenic males, suggesting that high levels of anti-müllerian hormone may impair Leydig cell steroidogenesis.

Animals↗

PubMed1995

Synergistic effects of inhibins and müllerian-inhibiting substance on testicular tumorigenesis.

Members of the transforming growth factor-beta (TGF-beta) superfamily regulate diverse physiological processes in multiple tissues. In particular, important roles for the inhibins and müllerian-inhibiting substance (MIS) have been demonstrated in the regulation of cell growth control both in vitro and in vivo. Inhibin-deficient male and female mice develop mixed granulosa/Sertoli cell tumors with nearly 100% penetrance. MIS-deficient male mice develop as pseudohermaphrodites with oviducts and uteri. In addition, MIS-deficient males have Leydig cell hyperplasia and, in one case, a Leydig cell tumor. To determine whether MIS could modify the development of the granulosa/Sertoli cell tumors in inhibin-deficient mice or whether inhibin could alter the development of the Leydig cell hyperplasia of MIS-deficient mice, animals deficient for both inhibins and MIS were generated. Adult inhibin/MIS-deficient male mice developed testicular tumors and large fluid-filled uteri. The accumulation of uterine fluid was due in part to an increase in estradiol secretion from the tumors and was blocked by a pure estrogen antagonist, ICI 182,780. The testes of the inhibin/MIS-deficient males developed granulosa/Sertoli cell tumors and Leydig cell neoplasia earlier, grew faster, were less hemorrhagic, and produced less estradiol than the testes of inhibin-deficient controls. These results demonstrate that inhibin and MIS synergize to influence testicular tumor development.

Animals↗

PubMed1995

Goosecoid is not an essential component of the mouse gastrula organizer but is required for craniofacial and rib development.

Goosecoid (gsc) is an evolutionarily conserved homeobox gene expressed in the gastrula organizer region of a variety of vertebrate embryos, including zebrafish, Xenopus, chicken and mouse. To understand the role of gsc during mouse embryogenesis, we generated gsc-null mice by gene targeting in embryonic stem cells. Surprisingly, gsc-null embryos gastrulated and formed the primary body axes; gsc-null mice were born alive but died soon after birth with numerous craniofacial defects. In addition, rib fusions and sternum abnormalities were detected that varied depending upon the genetic background. Transplantation experiments suggest that the ovary does not provide gsc function to rescue gastrulation defects. These results demonstrate that gsc is not essential for organizer activity in the mouse but is required later during embryogenesis for craniofacial and rib cage development.

Animals↗

PubMed1994

Homeotic transformation of cervical vertebrae in Hoxa-4 mutant mice.

Hoxa-4 (previously known as Hox-1.4) is a mouse homeobox-containing gene that is expressed in the presumptive hindbrain and spinal cord, prevertebrae, and other tissues during embryogenesis. To understand the role of Hoxa-4 during development, we generated Hoxa-4 mutant mice. Homozygous mutants were viable and fertile. Analysis of neonatal skeletons revealed the development of ribs on the seventh cervical vertebra at variable penetrance and expressivity. A low frequency of alterations in sternal morphogenesis was also observed. In addition, we analyzed the skeletons of transgenic mice that overexpress Hoxa-4 and found that the formation of the small rib anlagen that often develop on the seventh cervical vertebra was suppressed. Analysis of adult homozygous mutant skeletons revealed that the dorsal process normally associated with the second cervical vertebra was also found on the third cervical vertebra. These results demonstrate that Hoxa-4 plays a role in conferring positional information along the anteroposterior axis to specify the identity of the third and the seventh cervical vertebrae.

Animals↗

PubMed1994

Müllerian-inhibiting substance function during mammalian sexual development.

To investigate the role of Müllerian-inhibiting substance (MIS) in mammalian sexual development, we generated MIS-deficient mice. Although MIS-deficient males had testes that were fully descended and produced functional sperm, they also developed female reproductive organs, which interfered with sperm transfer into females, rendering most of these males infertile. Their testes had Leydig cell hyperplasia and, in one instance, neoplasia. The actions of the two primary hormones of male sexual differentiation were genetically eliminated using the testicular feminization (Tfm) mutation in combination with the MIS mutant allele. XY Tfm/MIS double mutants developed as females, with a uterus, coiled oviducts, and no male reproductive organs except undescended dysfunctional testes. These results suggest that eliminating the presumptive female reproductive tract in male fetuses facilitates fertility and that in testes MIS is a negative regulator of Leydig cell proliferation. Eliminating the presumptive male reproductive tract is necessary for proper oviductal morphogenesis during female mouse development.

Androgen-Insensitivity Syndrome↗