Fatal embryonic bleeding events in mice lacking tissue factor, the cell-associated initiator of blood coagulation.

Tissue factor (TF) is the cellular receptor for coagulation factor VI/VIIa and is the membrane-bound glycoprotein that is generally viewed as the primary physiological initiator of blood coagulation. To define in greater detail the physiological role of TF in development and hemostasis, the TF gene was disrupted in mice. Mice heterozygous for the inactivated TF allele expressed approximately half the TF activity of wild-type mice but were phenotypically normal. However, homozygous TF-/- pups were never born in crosses between heterozygous mice. Analysis of mid-gestation embryos showed that TF-/- embryos die in utero between days 8.5 and 10.5. TF-/- embryos were morphologically distinct from their TF+/+ and TF+/- littermates after day 9.5 in that they were pale, edematous, and growth retarded. Histological studies showed that early organogenesis was normal. The initial failure in TF-/- embryos appeared to be hemorrhaging, leading to the leakage of embryonic red cells from both extraembryonic and embryonic vessels. These studies indicate that TF plays an indispensable role in establishing and/or maintaining vascular integrity in the developing embryo at a time when embryonic and extraembryonic vasculatures are fusing and blood circulation begins.

Absence of yolk sac hematopoiesis from mice with a targeted disruption of the scl gene.

The scl gene encodes a basic-helix-loop-helix transcription factor which was identified through its involvement in chromosomal translocations in T-cell leukemia. To elucidate its physiological role, scl was targeted in embryonic stem cells. Mice heterozygous for the scl null mutation were intercrossed and their offspring were genotyped. Homozygous mutant (scl-/-) pups were not detected in newborn litters, and analysis at earlier time points demonstrated that scl-/- embryos were dying around embryonic day 9.5. The scl-/- embryos were pale, edematous, and markedly growth retarded after embryonic day 8.75. Histological studies showed complete absence of recognizable hematopoiesis in the yolk sac of these embryos. Early organogenesis appeared to be otherwise normal. Culture of yolk sac cells of wild-type, heterozygous, and homozygous littermates confirmed the absence of hematopoietic cells in scl-/- yolk sacs. Reverse transcription PCR was used to examine the transcripts of several genes implicated in early hematopoiesis. Transcripts of GATA-1 and PU.1 transcription factors were absent from RNA from scl-/- yolk sacs and embryos. These results implicate scl as a crucial regulator of early hematopoiesis.

A rice homeobox gene, OSH1, is expressed before organ differentiation in a specific region during early embryogenesis.

Homeobox genes encode a large family of homeodomain proteins that play a key role in the pattern formation of animal embryos. By analogy, homeobox genes in plants are thought to mediate important processes in their embryogenesis, but there is very little evidence to support this notion. Here we described the temporal and spatial expression patterns of a rice homeobox gene, OSH1, during rice embryogenesis. In situ hybridization analysis revealed that in the wild-type embryo, OSH1 was first expressed at the globular stage, much earlier than organogenesis started, in a ventral region where shoot apical meristem and epiblast would later develop. This localized expression of OSH1 indicates that the cellular differentiation has already occurred at this stage. At later stages after organogenesis had initiated, OSH1 expression was observed in shoot apical meristem [except in the L1 (tunica) layer], epiblast, radicle, and their intervening tissues in descending strength of expression level with embryonic maturation. We also performed in situ hybridization analysis with a rice organless embryo mutant, orl1, that develops no embryonic organs. In the orl1 embryo, the expression pattern of OSH1 was the same as that in the wild-type embryo in spite of the lack of embryonic organs. This shows that OSH1 is not directly associated with organ differentiation, but may be related to a regulatory process before or independent of the organ determination. The results described here strongly suggest that, like animal homeobox genes, OSH1 plays an important role in regionalization of cell identity during early embryogenesis.