Activation of human histidine decarboxylase gene promoter activity by gastrin is mediated by two distinct nuclear factors

The human histidine decarboxylase gene is regulated by gastrin through a cis-acting element known as the gastrin response element (GAS-RE) that was initially localized to a site (+2 to +24) downstream of the transcriptional start site. Electrophoretic mobility shift assays using sequentially deleted DNA probes and nuclear extracts from AGS-B gastric cancer cells showed that the GAS-RE is actually composed of two overlapping binding sites (GAS-RE1, +1 +10; and GAS-REB, +11 to +27) that bind distinct nuclear factors. Reporter gene assays demonstrated that each element alone could confer gastrin responsiveness, but the presence of both elements was required for complete gastrin response. Stimulation of AGS-B cells with gastrin for 10-20 min resulted in a >2-fold increase in factor binding. The binding was inhibited by pretreatment of AGS-B cells with cycloheximide and the MEK1 inhibitor PD98059, indicating a requirement for protein synthesis and also indicating that activation occurs through the MEK/mitogen-activated protein kinase pathway. UV cross-linking and Southwestern blot analysis showed that GAS-RE1 bound a 52-kDa protein, whereas GAS-RE2 bound a 35-kDa protein. Hence, activation of histidine decarboxylase gene promoter activity by gastrin is most likely mediated by two separate nuclear factors.