Patrick P.L. Tam

Mouse gastrulation: The formation of a mammalian body plan

The process of gastrulation is a pivotal step in the formation of the vertebrate body plan. The primary function of gastrulation is the correct placement of precursor tissues for subsequent morphogenesis. There is now mounting evidence that the body plan is established through inductive interactions between germ layer tissues and by the global patterning activity emanating from embryonic organizers. An increasing number of mouse mutants have been described that have gastrulation defects, providing important insights into the molecular mechanisms that regulate this complex process. In this review, we explore the mouse embryo before and during gastrulation, highlighting its similarities with other vertebrate embryos and its unique characteristics.

Blastocyst lineage formation, early embryonic asymmetries and axis patterning in the mouse

The investigation into lineage allocation and early asymmetries in the pre- and peri-implantation mouse embryo is gaining momentum. As we review here, new insights have been gained into the cellular and molecular events that lead to the establishment of the three lineages of the blastocyst, to the determination of the origin and the fates of the visceral endoderm in the peri-implantation mouse embryo, and to the generation of cellular and molecular activities that accompany the emergence of asymmetries in the pre-gastrulation embryo. We also discuss the continuing debate that surrounds the relative impacts of early lineage bias versus the stochastic allocation of cells with respect to the events that pattern the blastocyst and initiate its later asymmetries.

Gene function in mouse embryogenesis: get set for gastrulation

During early mouse embryogenesis, temporal and spatial regulation of gene expression and cell signalling influences lineage specification, embryonic polarity, the patterning of tissue progenitors and the morphogenetic movement of cells and tissues. Uniquely in mammals, the extraembryonic tissues are the source of signals for lineage specification and tissue patterning. Here we discuss recent discoveries about the lead up to gastrulation, including early manifestations of asymmetry, coordination of cell and tissue movements and the interactions of transcription factors and signalling activity for lineage allocation and germ-layer specification.

Mutations of CDKL5 Cause a Severe Neurodevelopmental Disorder with Infantile Spasms and Mental Retardation

Rett syndrome (RTT) is a severe neurodevelopmental disorder caused, in most classic cases, by mutations in the X-linked methyl-CpG-binding protein 2 gene (MECP2). A large degree of phenotypic variation has been observed in patients with RTT, both those with and without MECP2 mutations. We describe a family consisting of a proband with a phenotype that showed considerable overlap with that of RTT, her identical twin sister with autistic disorder and mild-to-moderate intellectual disability, and a brother with profound intellectual disability and seizures. No pathogenic MECP2 mutations were found in this family, and the Xq28 region that contains the MECP2 gene was not shared by the affected siblings. Three other candidate regions were identified by microsatellite mapping, including 10.3 Mb at Xp22.31-pter between Xpter and DXS1135, 19.7 Mb at Xp22.12-p22.11 between DXS1135 and DXS1214, and 16.4 Mb at Xq21.33 between DXS1196 and DXS1191. The ARX and CDKL5 genes, both of which are located within the Xp22 region, were sequenced in the affected family members, and a deletion of nucleotide 183 of the coding sequence (c.183delT) was identified in CDKL5 in the affected family members. In a screen of 44 RTT cases, a single splice-site mutation, IVS13-1G-->A, was identified in a girl with a severe phenotype overlapping RTT. In the mouse brain, Cdkl5 expression overlaps--but is not identical to--that of Mecp2, and its expression is unaffected by the loss of Mecp2. These findings confirm CDKL5 as another locus associated with epilepsy and X-linked mental retardation. These results also suggest that mutations in CDKL5 can lead to a clinical phenotype that overlaps RTT. However, it remains to be determined whether CDKL5 mutations are more prevalent in specific clinical subgroups of RTT or in other clinical presentations.

Lineage choice and differentiation in mouse embryos and embryonic stem cells.

The use of embryonic stem (ES) cells for generating healthy tissues has the potential to revolutionize therapies for human disease or injury, for which there are currently no effective treatments. Strategies for manipulating stem cell differentiation should be based on knowledge of the mechanisms by which lineage decisions are made during early embryogenesis. Here, we review current research into the factors influencing lineage differentiation in the mouse embryo and the application of this knowledge to in vitro differentiation of ES cells. In the mouse embryo, specification of tissue lineages requires cell-cell interactions that are influenced by coordinated cell migration and cellular neighborhood mediated by the key WNT, FGF, and TGFbeta signaling pathways. Mimicking the cellular interactions of the embryo by providing appropriate signaling molecules in culture has enabled the differentiation of ES cells to be directed predominately toward particular lineages. Multistep strategies incorporating the provision of soluble factors known to influence lineage choices in the embryo, coculture with other cells or tissues, genetic modification, and selection for desirable cell types have allowed the production of ES cell derivatives that produce beneficial effects in animal models. Increasing the efficiency of this process can only result from a better understanding of the molecular control of cell lineage determination in the embryo.

Extrinsic regulation of pluripotent stem cells

During early mammalian development, as the pluripotent cells that give rise to all of the tissues of the body proliferate and expand in number, they pass through transition states marked by a stepwise restriction in developmental potential and by changes in the expression of key regulatory genes. Recent findings show that cultured stem-cell lines derived from different stages of mouse development can mimic these transition states. They further reveal that there is a high degree of heterogeneity and plasticity in pluripotent populations in vitro and that these properties are modulated by extrinsic signalling. Understanding the extrinsic control of plasticity will guide efforts to use human pluripotent stem cells in research and therapy.

Regulation of Murine Telomere Length by Rtel: An Essential Gene Encoding a Helicase-like Protein

Little is known about the genes that regulate telomere length diversity between mammalian species. A candidate gene locus was previously mapped to a region on distal mouse Chr 2q. Within this region, we identified a gene similar to the dog-1 DNA helicase-like gene in C. elegans. We cloned this Regulator of telomere length (Rtel) gene and inactivated its expression in mice. Rtel(-/-) mice died between days 10 and 11.5 of gestation with defects in the nervous system, heart, vasculature, and extraembryonic tissues. Rtel(-/-) embryonic stem cells showed telomere loss and displayed many chromosome breaks and fusions upon differentiation in vitro. Crosses of Rtel(+/-) mice with Mus spretus showed that Rtel from the Mus musculus parent is required for telomere elongation of M. spretus chromosomes in F1 cells. We conclude that Rtel is an essential gene that regulates telomere length and prevents genetic instability.

Separate Progenitors for Radial and Tangential Cell Dispersion during Development of the Cerebral Neocortex

Cell lineage analyses suggest that cortical neuroblasts are capable of undertaking both radial and tangential modes of cell movement. However, it is unclear whether distinct progenitors are committed to generating neuroblasts that disperse exclusively in either radial or tangential directions. Using highly unbalanced mouse stem cell chimeras, we have identified certain progenitors that are committed to one mode of cell dispersion only. Radially dispersed neurons expressed glutamate, the neurochemical signature of excitatory pyramidal cells. In contrast, tangential progenitors gave rise to widely scattered neurons that are predominantly GABAergic. These results suggest lineage-based mechanisms for early specification of certain progenitors to distinct dispersion pathways and neuronal phenotypes.

Early endoderm development in vertebrates: lineage differentiation and morphogenetic function

Gastrulation of the vertebrate embryo culminates in the formation of three primary germ layers: ectoderm, mesoderm and endoderm. The endoderm contributes to the lining of the gut and the associated organs. New components of the molecular pathway for endoderm specification have been identified in the zebrafish and Xenopus. In the mouse, the activity of orthologous factors is involved with the allocation and differentiation of the definitive endoderm. Morphogenetic interactions between the endoderm and the other germ layer derivatives are critical for the morphogenesis of head structures and organogenesis of gut derivatives.

A specific requirement for PDGF-C in palate formation and PDGFR-alpha signaling

PDGF-C is a member of the platelet-derived growth factor (PDGF) family, which signals through PDGF receptor (PDGFR) αα and αβ dimers. Here we show that Pdgfc−/− mice die in the perinatal period owing to feeding and respiratory difficulties associated with a complete cleft of the secondary palate. This phenotype was less severe than that of Pdgfra−/− embryos. Pdgfc−/− Pdgfa−/− embryos developed a cleft face, subepidermal blistering, deficiency of renal cortex mesenchyme, spina bifida and skeletal and vascular defects. Complete loss of function of both ligands, therefore, phenocopied the loss of PDGFR-α function, suggesting that both PDGF-A and PDGF-C signal through PDGFR-α to regulate the development of craniofacial structures, the neural tube and mesodermal organs. Our results also show that PDGF-C signaling is a new pathway in palatogenesis, different from, and independent of, those previously implicated.