Generation of aminoterminally truncated, stable types of bioactive bovine and porcine fibroblast growth factor 4 in Escherichia coli.

Fibroblast growth factor 4 (FGF4) is a crucial growth factor for the development of mammalian embryos. We previously produced hexahistidine-tagged, bovine and porcine FGF4 (Pro(32) to Leu(206) ) proteins without a secretory signal peptide at the aminoterminus in Escherichia coli. Here, we found that these were unstable; site-specific cleavage between Ser(54) and Leu(55) in both FGF4 derivatives was identified.

Production of an aminoterminally truncated, stable type of bioactive mouse fibroblast growth factor 4 in Escherichia coli.

In mice, fibroblast growth factor 4 (Fgf4) is a crucial gene for the generation of trophectoderm, progenitor cells of the placenta. Therefore, exogenous FGF4 promotes the isolation and maintenance of trophoblast stem cells from preimplantation embryos. We previously produced a 6× histidine (His)-tagged, mouse FGF4 (Pro(31)-Leu(202)) without a secretory signal peptide at the amino-terminus, referred to as HismFGF4, in Escherichia coli.

Identification of site-specific degradation in bacterially expressed human fibroblast growth factor 4 and generation of an aminoterminally truncated, stable form.

Fibroblast growth factor 4 (FGF4) is considered as a crucial gene for tumorigenesis in humans and the development of mammalian embryos. The secreted, mature form of human FGF4 is thought to be comprised of 175 amino acid residues (proline(32) to leucine(206), Pro(32)-Leu(206)). Here, we found that bacterially expressed, 6× histidine (His)-tagged human FGF4 (Pro(32)-Leu(206)) protein, referred to as HishFGF4, was unstable such as in phosphate-buffered saline.

Production of bioactive bovine fibroblast growth factor 4 in E. coli based on the common nucleotide sequence of its structural gene in three breeds.

Fibroblast growth factor 4 (FGF4) is considered a crucial gene in the proper development of bovine embryos. We recently determined the FGF4 gene sequence in eight cattle derived from three breeds and revealed a common nucleotide sequence of the structural gene encoding FGF4, which leads to the deletion and mutation of amino acid sequences in the mature FGF4 (Pro(32) -Leu(206) ) compared with the sequence previously reported. In the present study, HisbFGF4, a 6× histidine-tagged bovine FGF4 (Pro(32) -Leu(206) ), was produced in Escherichia coli based on the validated nucleotide sequence and purified by heparin column chromatography.

Common amino acid sequences deduced from coding exons of the porcine FGF4 gene in two breeds and production of the encoded protein in Escherichia coli.

Fibroblast growth factor 4 (FGF4) is considered a crucial gene in the development of mammalian embryos. Here we identified common amino acid sequences predicted from coding exons of the FGF4 gene in five pigs of two breeds, and HispFGF4, a 6× histidine-tagged porcine FGF4, was produced in Escherichia coli. HispFGF4 was purified efficiently from the supernatant of cell lysate by heparin column chromatography.

Severe inhibition of in vitro cardiomyogenesis in mouse embryonic stem cells ectopically expressing EGAM1C homeoprotein.

Embryoid bodies were prepared from mouse embryonic stem cells expressing exogenous EGAM1C to analyze their ability to differentiate toward terminally differentiated cell types. The generation of cardiomyocytes was severely suppressed in Egam1c transfectants without upregulation of Nkx2-5, a crucial gene for cardiomyogenesis. These results indicate that EGAM1C is capable of affecting terminal differentiation in mouse embryonic stem cells.

Preferential emergence of cell types expressing markers for primitive endoderm lineages in mouse embryonic stem cells expressing exogenous EGAM1 homeoprotein.

Embryonic stem (ES) cells have been considered as a valuable renewable source of materials in regenerative medicine. Recently, we identified the homeoprotein EGAM1 both in preimplantation mouse embryos and mouse ES cells. Expression of the Egam1 transcript and its encoded protein was detectable in differentiating mouse ES cells, while it was almost undetectable in undifferentiated cells.

Effect of ectopic expression of homeoprotein EGAM1C on the cell morphology, growth, and differentiation in a mouse embryonic stem cell line, MG1.19 cells.

The homeoprotein EGAM1C was identified in preimplantation mouse embryos and embryonic stem (ES) cells. To explore the impact of EGAM1C on the hallmarks of mouse ES cells, MG1.19 cells stably expressing EGAM1C at levels similar to those in blastocysts were established using an episomal expression system.