Protein stability and domain topology determine the transcriptional activity of the mammalian glial cells missing homolog, GCMb.

The glial cells missing (GCM) family of transcription factors consists of Drosophila GCM and the mammalian proteins GCMa and GCMb. They are expressed in a highly restricted manner during development and are known or assumed to be important regulators of developmental fate decisions. As the biochemical properties of GCMb have not been studied so far, we have undertaken a detailed structure-function analysis of the mouse GCMb (mGCMb) protein. DNA-binding specificity was very similar to that of other GCM proteins. Nevertheless, mGCMb was only a weak transcriptional activator in a number of different tissue culture systems. Interestingly, this was not due to an intrinsic absence of transactivation potential. In effect, we were able to identify two separate transactivation domains within mGCMb, one carboxyl-terminally adjacent to the DNA-binding domain and the second within the extreme carboxyl terminus. Activity of both transactivation domains was, however, modulated by an inhibitory region unique to mGCMb and located between the two transactivation domains. Furthermore, pulse-chase experiments proved that the mGCMb protein has a half-life approximately four times shorter than mGCMa. Introduction of the above mentioned inhibitory domain of mGCMb into mGCMa shortened the half-life of mGCMa to a value typical of mGCMb with a concomitant reduction in transactivation potential. Given the strong correlation between protein stability and transactivation potential, functional differences between the two mammalian GCM homologs are likely due to differences in stability with a single inhibitory region in mGCMb being involved in the reduction of both.