Maintenance of telomeres is implicated in chromosome stabilization and cell immortalization. Telomerase, which catalyzes de novo synthesis of telomeres, is activated in germ cells and most cancers. Telomerase activity is regulated by gene expression for its catalytic subunit, TERT, whereas several lines of evidence have suggested a post‐translational regulation of telomerase activity. Here we identify the 14‐3‐3 signaling proteins as human TERT (hTERT)‐binding partners. A dominant‐negative 14‐3‐3 redistributed hTERT, which was normally predominant in the nucleus, into the cytoplasm. Consistent with this observation, hTERT‐3A, a mutant that could not bind 14‐3‐3, was localized into the cytoplasm. Leptomycin B, an inhibitor of CRM1/exportin 1‐mediated nuclear export, or disruption of a nuclear export signal (NES)‐like motif located just upstream of the 14‐3‐3 binding site in hTERT impaired the cytoplasmic localization of hTERT. Compared with wild‐type hTERT, hTERT‐3A increased its association with CRM1. 14‐3‐3 binding was not required for telomerase activity either in vitro or in cell extracts. These observations suggest that 14‐3‐3 enhances nuclear localization of TERT by inhibiting the CRM1 binding to the TERT NES‐like motif.

Human cells contain a protein that binds to UV-irradiated DNA with high affinity. This protein, damaged DNA-binding protein (DDB), is a heterodimer of two polypeptides, p127 and p48. Recent in vivo studies suggested that DDB is involved in global genome repair of cyclobutane pyrimidine dimers (CPDs), but the mechanism remains unclear. Here, we show that in vitro DDB directly stimulates the excision of CPDs but not (6 -4)photoproducts. The excision activity of cell-free extracts from Chinese hamster AA8 cell line that lacks DDB activity was increased 3-4-fold by recombinant DDB heterodimer but not p127 subunit alone. Moreover, the addition of XPA or XPA ؉ replication protein A (RPA), which themselves enhanced excision, also enhanced the excision in the presence of DDB. DDB was found to elevate the binding of XPA to damaged DNA and to make a complex with damaged DNA and XPA or XPA ؉ RPA as judged by both electrophoretic mobility shift assays and DNase I protection assays. These results suggest that DDB assists in the recognition of CPDs by core NER factors, possibly through the efficient recruitment of XPA or XPA⅐RPA, and thus stimulates the excision reaction of CPDs. Nucleotide excision repair (NER)1 is the major mechanism for removing bulky DNA lesions including cyclobutane pyrimidine dimers (CPDs) and (6 -4)photoproducts induced by sun light in humans (1-4). The basic reaction mechanism of NER is highly conserved from yeast to human. The core reaction from DNA damage recognition to excision of the damage is accomplished by six repair factors (XPA, RPA, XPC⅐HR23B, TFIIH, XPF⅐ERCC1, XPG), so called excision nuclease (excinuclease) (5-8). The recent development of reconstituted systems using highly purified proteins has enabled analyses of the detailed reaction mechanism of NER (7-10). In yeast, some accessory or regulatory factors such as Rad7⅐Rad16 complex (11-13), ABF1 (14), and MMS19 (15-16) have been reported to help mediate NER. The existence of such factors in humans, however, has not been clearly demonstrated.Damaged DNA-binding protein (DDB) is one of the candidates that might play such an accessory role in human cells. DDB is composed of two subunits, p127 and p48, and has a higher affinity for various types of DNA lesions compared with the damage recognition subunits (XPA, RPA, XPC⅐HR23B, and TFIIH) of the six core repair factors (17-19). DDB activity has been shown to be missing in a subset of xeroderma pigmentosum group E (XP-E) patients. XP is an autosomal recessive disease associated with abnormal sensitivity to UV lights and a high incidence of skin cancer, which is known to be caused by defects in NER. XP-E is one of eight genetic complementation groups (XP-A through XP-G and XP-V) of XP patients, and, along with XP-V, manifests the mildest clinical features and deficiency in NER. XP-E individuals lacking DDB activity were found to carry mutations in p48 gene (20 -23). Because of its high specificity for binding to damaged DNA and a defect in some XP-E cells, DDB has been postulated to be involved...