The naturally occurring mutants of DDB are impaired in stimulating nuclearimport of the p125 subunit and E2F1-activated transcription

The human UV-damaged-DNA binding protein DDB has been linked to the repair deficiency disease xeroderma pigmentosum group E (XP-E), because a subset o f XP-E patients lack the damaged-DNA binding function of DDB. Moreover, the microinjection of purified DDB complements the repair deficiency in XP-E c ells lacking DDB. Two naturally occurring XP-E mutations of DDB, 82TO and 2 RO, have been characterized. They have single amino acid substitutions (K24 4E and R273H) within the WD motif of the p48 subunit of DDB, and the mutate d proteins lack the damaged-DNA binding activity. In this report, we descri be a new function of the p48 subunit of DDB, which reveals additional defec ts in the function of the XP-E mutants. We show that when the subunits of D DB were expressed individually, p48 localized in the nucleus and p125 local ized in the cytoplasm. The coexpression of p125 with p48 resulted in an inc reased accumulation of p125 in the nucleus, indicating that p48 plays a cri tical role in the nuclear localization of p125. The mutant forms of p48, 2R O and 82TO, are deficient in stimulating the nuclear accumulation of the p1 25 subunit of DDB. In addition, the mutant 2RO fails to form a stable compl ex with the p125 subunit of DDB. Our previous studies indicated that DDB ca n associate with the transcription factor E2F1 and can function as a transc riptional partner of E2F1. Here we show that the two mutants, while they as sociate with E2F1 as efficiently as wild-type p48, are severely impaired in stimulating E2F1-activated transcription. This is consistent with our obse rvation that both subunits of DDB are required to stimulate E2F1-activated transcription. The results provide insights into the functions of the subun its of DDB and suggest a possible link between the role of DDB in E2F1-acti vated transcription and the repair deficiency disease XP-E.