Abstract
Polypyrimidine-tract binding protein 1 (PTBP1) is an important cellular regulator of messenger RNAs influencing the alternative splicing profile of a cell as well as its mRNA stability, location and translation. In addition, it is diverted by some viruses to facilitate their replication. Here, we used a novel PTBP1 knockout mouse to analyse the tissue expression pattern of PTBP1 as well as the effect of its complete removal during development. We found evidence of strong PTBP1 expression in embryonic stem cells and throughout embryonic development, especially in the developing brain and spinal cord, the olfactory and auditory systems, the heart, the liver, the kidney, the brown fat and cartilage primordia. This widespread distribution points towards a role of PTBP1 during embryonic development. Homozygous offspring, identified by PCR and immunofluorescence, were able to implant but were arrested or retarded in growth. At day 7.5 of embryonic development (E7.5) the null mutants were about 5x smaller than the control littermates and the gap in body size widened with time. At mid-gestation, all homozygous embryos were resorbed/degraded. No homozygous mice were genotyped at E12 and the age of weaning. Embryos lacking PTBP1 did not display differentiation into the 3 germ layers and cavitation of the epiblast, which are hallmarks of gastrulation. In addition, homozygous mutants displayed malformed ectoplacental cones and yolk sacs, both early supportive structure of the embryo proper. We conclude that PTBP1 is not required for the earliest isovolumetric divisions and differentiation steps of the zygote up to the formation of the blastocyst. However, further post-implantation development requires PTBP1 and stalls in homozygous null animals with a phenotype of dramatically reduced size and aberration in embryonic and extra-embryonic structures.
Highlights
Mouse embryonic development lasts about 3 weeks in total
Broad expression of Polypyrimidine-tract binding protein 1 (PTBP1) during gestation In order to gauge the importance of PTBP1 in embryonic development, we performed X-gal staining of heterozygous embryonic stem cells (ESCs) and heterozygous whole embryos at equal time intervals between implantation at E4-4.5 and birth at E20-21 (Figure 2)
We conclude that PTBP1 is strongly and broadly expressed during the entire gestational period and in the cultured embryonic stem cells used to generate this knockout mouse model
Summary
During the first days after fertilization the embryo does not grow significantly in size, remaining at around 0.1 mm in diameter, and differentiates only into 2 tissue types, the outside trophoblast and the inner cell mass. Implantation at about day 4.5 of embryonic development (E4.5) [1] marks a change in pace. After this point, the volume of the embryo increases exponentially from ,0.2 mm in length at E4.5 to 0.7–0.8 at E7.5. Afterwards the 3 germ layers, ectoderm, mesoderm and definitive endoderm, are formed [2]. This marks the beginning of major differentiation events
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