Janet Heasman

Overexpression of cadherins and underexpression of β-catenin inhibit dorsal mesoderm induction in early Xenopus embryos

The cadherin-catenin complex has an important role in cell-cell adhesion and may also function in signaling pathways. We report that overexpression of three cadherin types in Xenopus embryos causes them to develop with reduced dorsal axial structures. The same phenotype is produced in embryos that have been depleted of maternal beta-catenin protein by an antisense oligodeoxynucleotide complementary to beta-catenin mRNA. They show an inhibition in the expression of dorsal mesodermal markers MyoD and goosecoid, but not of ventral and general mesodermal markers. They lack notochords, somites, and neural tubes and are defective in dorsal mesodermal signaling in Nieuwkoop assays. The phenotype can be rescued by the injection of beta-catenin mRNA and not by the injection of Xwnt-8 mRNA. These results show that beta-catenin has an important role in dorsal mesoderm induction. They directly demonstrate the activity of a maternal mRNA in axis specification.

The Role of Maternal VegT in Establishing the Primary Germ Layers in Xenopus Embryos

VegT is a T-box transcription factor whose mRNA is synthesized during oogenesis and localized in the vegetal hemisphere of the egg and early embryo. We show that maternally expressed VegT controls the pattern of primary germ layer specification in Xenopus embryos. Reduction of the maternal store completely alters the fates of different regions of the blastula so that animal cell fate is changed from epidermis and nervous system to epidermis only, equatorial cell fate is changed from mesoderm to ectoderm, and vegetal cell fate is changed from endoderm to mesoderm and ectoderm. Vegetal cells lose their capacity both to form endoderm and to release mesoderm-inducing signals. These results show that a single maternally expressed gene controls the patterning of the Xenopus blastula.

The onset of germ cell migration in the mouse embryo.

Mouse primordial germ cells (PGCs) are specified between embryonic day 6.5 (E6.5) and E7.5, when they have been visualized as an alkaline phosphatase-positive (AP+) cell population in the developing allantois. By E8.5, they are embedded in the hind-gut epithelium. Previous experiments have suggested different sites for PGCs origin, and it is unclear how they reach the gut epithelium. We have used transgenic mice expressing GFP under a truncated Oct4 promoter to visualize living PGCs. We find GFP+/AP+ cells in the posterior end of the primitive streak as a dispersed population of cells actively migrating into the allantois, and directly into the adjacent embryonic endoderm. Time-lapse analysis shows these cells to be actively migratory from the time they exit the primitive streak.

Xklp15 a chromosomal xenopus kinesin-like protein essential for spindle organization and chromosome positioning

Xklp1 is a novel Xenopus kinesin-like protein with a motor domain at the amino terminus, nuclear localization sequences in the stalk, and a putative zinc finger-like sequence in the tail. It is nuclear during interphase and chromosomal during mitosis. During late anaphase, a fraction of the protein relocalizes to the spindle interzone and accumulates in the midbody during telophase. Depletion of Xklp1 protein by antisense oligo knockout in oocytes leads to defective mitosis during the first cell cycles following fertilization. The bipolarity of spindles assembled in vitro in the presence of anti-Xklp1 antibodies is unstable, and the chromosomes fail to congress on the metaphase plate.

The role of cadherins during primordial germ cell migration and early gonad formation in the mouse

Primordial germ cells (PGCs) are the founder cells of the gametes. In mammals, PGCs migrate from the hindgut to the genital ridges, where they coalesce with each other and with somatic cells to form the primary sex cords. We show here that, in both sexes, PGCs express P- and E-cadherins during and after migration, and N-cadherin at post-migratory stages. E-Cadherin is not expressed by PGCs whilst in the hindgut, but is upregulated as they leave. Blocking antibodies against E-, but not P-cadherin cause defective PGC-PGC coalescence, and in some cases, ectopic PGCs.

A Kinesin-like Protein Is Required for Germ Plasm Aggregation in Xenopus

In embryos of X. laevis, and many other species, early development requires targeted movements of molecules and molecular aggregates within the oocyte or egg cytoplasm. One well-known example in Xenopus is the aggregation of germ plasm, a group of cytoplasmic islands that become distributed during the first few cell cycles to cells that will give rise to the germ line. Nothing is known about the cytoskeletal motor proteins that may drive these movements. We show here that a recently identified Xenopus kinesin-like protein, Xklp1, is required for the aggregation of germ plasm in early Xenopus embryos, thus assigning this protein a role in a developmentally important cytoplasmic localization.

Early events in the mammalian germ line.

Germ cells represent the genetic and cellular link between generations, as well as the transmitters of inherited diseases. Despite their central importance, not much is known about the molecular mechanisms whereby a germ cell lineage becomes set aside during development, or how the germ cells, once formed, migrate to the gonads and combine with somatic cells to make a gonad. This article provides a brief review of current knowledge on these issues, with particular focus on the mammalian germ line.

Early stages in male germ cell differentiation in the mouse

Primordial germ cells arise during gastrulation and migrate from the hindgut into the gonad primordium during early organogenesis. In this article, we discuss factors that control migration, proliferation and targeting of the PGCs. In particular we discuss how changes in adhesiveness control germ cell positioning in the gonad, and the molecules involved.

What my mother told me: Examining the roles of maternal gene products in a vertebrate

In Xenopus, mRNAs synthesized during oocyte differentiation are inherited by the egg and direct all protein synthesis until the late-blastula stage. This provides an opportunity to study the roles of maternally expressed genes in embryonic development of a vertebrate. Oocytes can be depleted of specific mRNAs by the injection of antisense deoxyoligonucleotides and then fertilized to assay for developmental abnormalities. The ease of experimental manipulation of early Xenopus embryos in culture gives considerable opportunity for the analysis of the abnormalities seen.

A rapid and quantitative assay of adhesiveness of mouse primordial germ cells that can be applied to many types of early embryonic cell

Cell migration is an essential component of embryogenesis. In order to move from one location to another, cells must quantitatively alter their adhesiveness to the different substrata they encounter. Little is known about these changes in adhesive strengths, nor how they are controlled. Here we describe a combination of simple, quantitative, and reproducible methods to partially purify primordial germ cells from different stages of early mouse embryos, and compare their strengths of adhesion to different extracellular matrix glycoproteins. Primordial germ cells arise very early in the mouse embryo, and migrate through the tissues of the embryo to reach the site of gonad formation. Here they assemble into sex cords and start to differentiate into male or female gametes. Although the focus here is germ line cells, in principle this method can be used on any population of cells in the early embryo.