Endocrine Abstracts

Bone morphogenetic protein-2 (BMP-2) signal transduction requires heterocomplexing between type II receptors and different type I (α or β) receptors resulting in the activation of different signalling pathways. Previously, we have used αT3-1 and Lβt-2 cell lines, which are representative of different stages of gonadotroph development, to demonstrate differential expression of the receptor isoforms during gonadotroph maturation. BMPR-Iα and II mRNA and protein were detected in both cell types but BMPR-Iβ was present only in the less differentiated αT3-1s.

To investigate activation of the Smad signalling pathway in these gonadotroph cell lines we have employed the Xvent2-luc plasmid. This plasmid contains a region of the BMP responsive Xenopus Vent2 promoter cloned into a luciferase reporter plasmid and has been demonstrated to respond to Smad activation following BMP treatment ¹.

αT3-1 and LβT-2 cells were transiently transfected with Xvent2-luc and incubated with 10, 50, 100 and 200 ng/ml of BMP-2 for 4 h. αT3-1 cells showed fold increases of 1.35±0.19 (50 ng/ml) and 1.35±0.10 (100 ng/ml) in Xvent2-luc activity above basal. In contrast, transfected LβT-2 cells failed to respond to BMP-2 at any concentration. Thus, we have demonstrated transcriptional stimulation of Xvent2-luc to BMP-2 in αT3-1 s but not in the fully differentiated LβT-2 s. This may relate to the absence of the type Iβ? receptor in the LβT-2 cell line.

Acknowledgements: This project is supported by the BBSRC.