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PTBP1 is required for embryonic development before gastrulation.

Suckale J, Wendling O, Masjkur J, Jäger M, Münster C, Anastassiadis K, Stewart AF, Solimena M - PLoS ONE (2011)

Bottom Line: Embryos lacking PTBP1 did not display differentiation into the 3 germ layers and cavitation of the epiblast, which are hallmarks of gastrulation.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 animals with a phenotype of dramatically reduced size and aberration in embryonic and extra-embryonic structures.

View Article: PubMed Central - PubMed

Affiliation: Molecular Diabetology, Paul Langerhans Institute Dresden, School of Medicine and University Clinic Carl Gustav Carus, Dresden University of Technology, Dresden, Germany.

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 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 animals with a phenotype of dramatically reduced size and aberration in embryonic and extra-embryonic structures.

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PTBP1  embryos show defects in yolk sac & placenta development.This figures compares yolk sac and placental structures in knockout and control embryos. A) Hematoxylin and eosin-stained paraffin sections of an E5.5 wild-type embryo, chosen for its comparable size to the  mutants, and an E7.5 yolk sac (left) juxtaposed with mutant E7.5 embryos and mutant yolk sacs, respectively (right). Mutants lack the typical structure of a normal embryo at this stage. If discernable at all they lack a typical yolk sac with a thin Reichert's membrane in close proximity to trophoblast giant cells and the decidua. B) Immunofluorescence for collagen 4 on a section of a control E7.5 embryo is shown next to a similar image from a homozygous embryo. The control embryo shows collagen 4 signal from several basement membranes including the Reichert's membrane of the yolk sac. The  mutant, on the other hand, shows an unusually extended area high in collagen 4, not resembling a membrane. Separate channels for the collagen immunomicroscopy can be seen in Figure S4.
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pone-0016992-g005: PTBP1 embryos show defects in yolk sac & placenta development.This figures compares yolk sac and placental structures in knockout and control embryos. A) Hematoxylin and eosin-stained paraffin sections of an E5.5 wild-type embryo, chosen for its comparable size to the mutants, and an E7.5 yolk sac (left) juxtaposed with mutant E7.5 embryos and mutant yolk sacs, respectively (right). Mutants lack the typical structure of a normal embryo at this stage. If discernable at all they lack a typical yolk sac with a thin Reichert's membrane in close proximity to trophoblast giant cells and the decidua. B) Immunofluorescence for collagen 4 on a section of a control E7.5 embryo is shown next to a similar image from a homozygous embryo. The control embryo shows collagen 4 signal from several basement membranes including the Reichert's membrane of the yolk sac. The mutant, on the other hand, shows an unusually extended area high in collagen 4, not resembling a membrane. Separate channels for the collagen immunomicroscopy can be seen in Figure S4.

Mentions: A wild-type embryo at E7.5 has formed an ectoplacental cone invaded by maternal blood [3] and has surrounded itself with a yolk sac (Figure 5a). The latter structure is cooperatively build by parietal endoderm forming a thick basement membrane (the Reichert's membrane) and trophoblast giant cells on the embryo side, as well as maternal blood and decidual cells on the outside [37]. In many homozygous mutant embryos the yolk sac was not clearly separated from the epiblast. Also, the Reichert's membrane was not properly formed and contained unusual quantities of eosinophilic material (Figure 5a). Similarly, the ectoplacental cone did not properly form, often lacking the characteristic connection of decidual cells, giant cell, the Reichert's membrane and parietal cells. A typical embryo is enveloped in visceral endodermal cells, which are subdivided into cuboidal cells with characteristic apical vacuoles and squamous cells that differentiate later. Even in the most advanced homozygous mutant at E7.5, the squamous endodermal cells were not visible showing a delay in the formation of the visceral yolk sac.


PTBP1 is required for embryonic development before gastrulation.

Suckale J, Wendling O, Masjkur J, Jäger M, Münster C, Anastassiadis K, Stewart AF, Solimena M - PLoS ONE (2011)

PTBP1  embryos show defects in yolk sac & placenta development.This figures compares yolk sac and placental structures in knockout and control embryos. A) Hematoxylin and eosin-stained paraffin sections of an E5.5 wild-type embryo, chosen for its comparable size to the  mutants, and an E7.5 yolk sac (left) juxtaposed with mutant E7.5 embryos and mutant yolk sacs, respectively (right). Mutants lack the typical structure of a normal embryo at this stage. If discernable at all they lack a typical yolk sac with a thin Reichert's membrane in close proximity to trophoblast giant cells and the decidua. B) Immunofluorescence for collagen 4 on a section of a control E7.5 embryo is shown next to a similar image from a homozygous embryo. The control embryo shows collagen 4 signal from several basement membranes including the Reichert's membrane of the yolk sac. The  mutant, on the other hand, shows an unusually extended area high in collagen 4, not resembling a membrane. Separate channels for the collagen immunomicroscopy can be seen in Figure S4.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3040740&req=5

pone-0016992-g005: PTBP1 embryos show defects in yolk sac & placenta development.This figures compares yolk sac and placental structures in knockout and control embryos. A) Hematoxylin and eosin-stained paraffin sections of an E5.5 wild-type embryo, chosen for its comparable size to the mutants, and an E7.5 yolk sac (left) juxtaposed with mutant E7.5 embryos and mutant yolk sacs, respectively (right). Mutants lack the typical structure of a normal embryo at this stage. If discernable at all they lack a typical yolk sac with a thin Reichert's membrane in close proximity to trophoblast giant cells and the decidua. B) Immunofluorescence for collagen 4 on a section of a control E7.5 embryo is shown next to a similar image from a homozygous embryo. The control embryo shows collagen 4 signal from several basement membranes including the Reichert's membrane of the yolk sac. The mutant, on the other hand, shows an unusually extended area high in collagen 4, not resembling a membrane. Separate channels for the collagen immunomicroscopy can be seen in Figure S4.
Mentions: A wild-type embryo at E7.5 has formed an ectoplacental cone invaded by maternal blood [3] and has surrounded itself with a yolk sac (Figure 5a). The latter structure is cooperatively build by parietal endoderm forming a thick basement membrane (the Reichert's membrane) and trophoblast giant cells on the embryo side, as well as maternal blood and decidual cells on the outside [37]. In many homozygous mutant embryos the yolk sac was not clearly separated from the epiblast. Also, the Reichert's membrane was not properly formed and contained unusual quantities of eosinophilic material (Figure 5a). Similarly, the ectoplacental cone did not properly form, often lacking the characteristic connection of decidual cells, giant cell, the Reichert's membrane and parietal cells. A typical embryo is enveloped in visceral endodermal cells, which are subdivided into cuboidal cells with characteristic apical vacuoles and squamous cells that differentiate later. Even in the most advanced homozygous mutant at E7.5, the squamous endodermal cells were not visible showing a delay in the formation of the visceral yolk sac.

Bottom Line: Embryos lacking PTBP1 did not display differentiation into the 3 germ layers and cavitation of the epiblast, which are hallmarks of gastrulation.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 animals with a phenotype of dramatically reduced size and aberration in embryonic and extra-embryonic structures.

View Article: PubMed Central - PubMed

Affiliation: Molecular Diabetology, Paul Langerhans Institute Dresden, School of Medicine and University Clinic Carl Gustav Carus, Dresden University of Technology, Dresden, Germany.

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 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 animals with a phenotype of dramatically reduced size and aberration in embryonic and extra-embryonic structures.

Show MeSH
Related in: MedlinePlus