In Mlo-y4 Cbfal Induced Expression Of The Separation Of The New Gene

The development of the skeleton is a complicated process involving many factors. Transcription factors play a vital role in the developing process. To date, Cbfal (Core Binding Factor 1) and Osx (Osterix) have been identified as transcription factors which specifically express in the osteoblast lineage cells. In 1997, Cbfal which can activate the earlier differentiation stage of the osteoblast was proved to be a key factor for bone formation. Moderate expression of Cbfal can promote the differentiation of the osteoblasts, but overexpression of Cbfal can suppress the mutation of the osteoblasts. It was in 2002 that Osx was identified by Kazuhisa as another transcription factor by Suppression Subtractive Hybridization (SSH). Cbfal and Osx act at different stages during the differentiation of the osteoblasts. Cbfal has an essential role in the stage that Multipotential mesenchymal progenitors differentiate into preosteoblasts, and Osx is required in preosteoblasts differentiating into functional osteoblasts.The direct purpose of this article is to separate some new genes induced by Cbfal, and its long-term aim is to identified some other transcription factors. We transfected Cbfal to MLO-Y4 (Murine Long Bone Osteocyte-Y4) which has the characters of osteocyte, and then separate induced genes from it by SSH. We have constructed a library containing genes induced by Cbfal successfully. We found that there are twenty-four Cbfal, nineteen known and forty unkown gene segments in the eighty-three positive clones which we selected randomly form the library. The unkown gene segments have a proportion of 67.8%. We checked one gene segment which has no homologous sequence in the mouse genome by Virtual Northern blot and found that it only expresses in theMLO-Y4 cells which were transfected with Cbfal. Having found that there are Osx gene segments among the known genes induced by Cbfal in our SSH result, then we checked it in MLO-Y4 cells by RT-PCR and performed similar experiments in the two osteoblasts: MC3T3-E1 and U2OS. We have got the same conclusion that Cbfal do induce the expression of Osx in these cells.