Characterization of two distinct families of transcription factors that bind to the CCAAT box region of the human COL1A2 gene

Abstract Both the mouse and human α2(I) procollagen promoters contain an inverted CCAAT box at −80, but only the human promoter contains an additional regulatory element, the collagen modulating element (CME), immediately downstream of the CCAAT box [Collins et al. (1997): Biochem J 322:199–206]. In this study, the transcription factors that bind to the G/CBE and CME within the human promoter were characterized in SVWI‐38 and CT‐1 nuclear extracts. Two distinct proteins bind to the CME, and both were identified as heat‐labile factors that were sensitive to high ionic strengths and required Zn 2+ for DNA‐binding activity. These proteins had Stokes radii of 4.12 and 3.15 nm, sedimentation coefficients of 3.9 and 3.2 S and native molecular weights of 66 and 41 kDa, respectively. On the basis of biochemical and DNA‐binding properties, the CME binding proteins are probably novel factors involved in the regulation of the human α2(I) procollagen gene. By contrast, the G/CBE binding proteins were more resistant to heat, ionic strength, and divalent metal ion chelators, demonstrating that the G/CBE and CME binding proteins had distinct DNA‐binding properties. The above properties suggest that this factor is a member of the previously characterized family of CCAAT box‐binding factors, CBF, NF‐Y, CP‐1 and α‐CP1. Taken together, these physicochemical properties of the COL1A2 CCAAT box and CME‐binding proteins demonstrated that they were distinct unrelated transcription factors. These results also suggest that there is a distinct difference in the DNA‐binding activity between the equivalent region of the mouse and human α2(I) procollagen promoters. J. Cell. Biochem. 70:455–467, 1998. © 1998 Wiley‐Liss, Inc.