CLONING AND ANALYSIS OF THE HUMAN PAX-5 GENE PROMOTER

The expression of Pax-5 gene is altered in human myeloma cells (malign ant plasma cells). This altered expression is considered to be closely involved in oncogenesis of human myeloma. To investigate the possible mechanism(s) underlying this alteration, we first cloned the 1,050 bp fragment in the 5' upstream region of human Pax-5 gene by PCR-mediate d gene walking method. The cloned fragment has predicted regulatory mo tifs for Lyf-1(Ik-1), Ik-2, bHLH, E-47, Sox-5, Oct-1, GATA-1,-2, and 3 , but it lacks a TATA box. By constructing deletion mutants of this fr agment, its basal promoter activity was analyzed by transfecting these mutants to Cos 7 cells. The maximal promoter activity was recovered b y the fragment that extends between -70 to -820 upstream of the transc ription start site. Also, three DNA fragments from this cloned sequenc e were used as templates in gel shift assay; these fragments covered m ost of the predicted regulatory sires. Specific binding activities wer e found in each DNA fragment. Therefore, we could clone the functional ly active fragment of 5' upstream region of human Pax-5 gene. (C) 1996 Academic Press, Inc.