We found that prostaglandin (PG) D(2) production was induced through transcriptional activation of cyclooxygenase (COX)-2 and lipocalin-type PGD synthase (L-PGDS) genes under serum-starved conditions in human brain-derived TE671 cells. Analysis of promoter and intron regions of the human L-PGDS gene demonstrated that an atypical E-box within intron 4 mediated serum starvation-induced up-regulation of L-PGDS gene expression. The results of electrophoretic mobility shift assay and chromatin immunoprecipitation assay showed that upstream stimulatory factor (USF) 1 bound to this atypical E-box. USF1 gene expression was also enhanced during serum starvation in TE671 cells through activation of p38 mitogen activated protein kinase, and the efficiency of the binding of USF1 to the atypical E-box was clearly increased by serum starvation. Administration of USF1 siRNA suppressed both L-PGDS and COX-2 gene expression and PGD(2) production. Moreover, NS-398, a COX-2 inhibitor and AT-56, an L-PGDS inhibitor, suppressed PGD(2) production in TE671 cells cultured under the serum-starved condition. These results indicate that PGD(2) production stimulated by serum starvation is mediated by both COX-2 and L-PGDS through enhancement of USF1 in TE671 cells.