Thrombospondin 3 (TSP3) is a secreted, pentameric glycoprotein whose regulation 
of expression and function are not well understood. Mouse Thbs3 is located just 
downstream from the divergently transcribed metaxin gene (Mtx), which encodes an 
outer mitochondrial membrane import protein. Although Thbs3 and Mtx share a 
common promoter region, previous studies showed that Mtx is regulated by 
proximal elements that had little effect on Thbs3 expression. In this study, 
transient transfection of rat chondrosarcoma cells and NIH-3T3 fibroblasts 
demonstrated that Thbs3 is regulated in a cell type-specific manner by a 
position- and orientation-independent far upstream enhancer located within 
intron 6 of Mtx. Despite its greater proximity to the transcription start site 
of Mtx, the Thbs3 enhancer did not have a significant effect on Mtx expression. 
Two DNA-protein complexes, which were both required for activity, were 
identified when nuclear extracts were assayed with a probe containing the 
enhancer sequence. The protein in one of these complexes was identified as Sp1, 
while the other DNA-protein complex remains uncharacterized. A 6-kilobase pair 
promoter containing the enhancer was able to direct specific expression of the 
E. coli lacZ gene in transgenic mice, whereas a 2-kilobase pair promoter that 
lacked the enhancer was inactive. Thus, despite their close proximity, the genes 
of the Mtx/Thbs3 gene cluster are regulated independently.