Murine Laminin B1 Gene Regulation during the Retinoic Acid- and Dibutyryl Cyclic AMP-induced Differentiation of Embryonic F9 Teratocarcinoma Stem Cells

Abstract

Retinoic acid (RA) and cyclic AMP analogs cause the differentiation of F9 embryonic teratocarcinoma stem cells into parietal endoderm, an epithelial cell of the early mouse embryo. Laminin B1 is induced in this differentiation process, but is not transcriptionally activated until 24-48 h after RA addition and is not maximally induced until approximately 72 h. Cyclic AMP analogs enhance this transcriptional activation. Although several DNase I hypersensitive sites (DHSS) were observed in the LAMB1 5 -flanking DNA, one of the sites, DHSS2, was detected only after 72 h of RA treatment. Transient transfections have demonstrated that the DHSS2 region functions as a "late-acting RA-inducible enhancer," and motifs in this enhancer contain the homeobox protein-binding site TTATTAACA. Greater binding is observed at these sites by electrophoretic mobility shift assay when cells are cultured with RA and cyclic AMP analogs versus RA alone, and no binding is seen in extracts from RA-treated F9 RAR gamma-/- cells which lack RAR gamma mRNA and protein. Laminin B1 mRNA is not induced by RA in the RAR gamma-/- cells (Boylan, J. F., Lohnes, D., Taneja, R., Chambon, P., and Gudas, L. J. (1993) Proc. Natl. Acad. Sci. U. S. A. 90, 9601-9605). Our data show that these DNA regulatory elements contribute to the transcriptional activation of the LAMB1 gene during the later stages of the differentiation process.

Extracellular matrix signaling: integration of form and function in normal and malignant cells

“…Similar studies in F9 teratocarcinoma cells have linked retinoic-acid-induced differentiation to upregulation of Hoxa1 and a Hox-dependent upregulation of laminin promoter activity [32,35]. Several papers in the past year have extended this relationship to both epithelial and endothelial cells (also summarized in Table 1).…”

Identification of an Enhancer That Controls Up-regulation of Fibronectin during Differentiation of Embryonic Stem Cells into Extraembryonic Endoderm

“…While extraembryonic endoderm cells are first comprised of primitive endoderm upon segregation from the embryonic ICM cells, the primitive endoderm differentiates further into parietal endoderm (PE) and visceral endoderm (VE) in vivo (1). F9 cells are able to mimic this process in culture when specific inductive factors are added (1,25,26). Indeed VE-like cells can be obtained by culturing F9 EC cells with retinoic acid (RA), while the addition of both RA and dibutyryl cAMP converts them to PE-like cells.…”

The topographical regulation of embryonic stem cell differentiation

“…Importantly, it was established that endodermal cells were not already present within the aggregates, ruling out the possibility that endodermal cells migrated to the periphery in response to retinoic acid (Becker et al 1992). Furthermore, if the F9 cells were cultured in a monolayer, retinoic acid induced the whole population to become endoderm (Li & Gudas 1996). These observations show that the way in which F9 cells respond to retinoic acid depends upon their interaction with other cells; being centrally located within an aggregate renders F9 cells incapable of responding to endoderm-inducing cues.…”