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Proc Natl Acad Sci U S A. Jan 21, 2003; 100(2): 616–621.
Published online Jan 13, 2003. doi:  10.1073/pnas.0236176100
PMCID: PMC141045

Global genome repair is required to activate KIN17, a UVC-responsive gene involved in DNA replication


UV light provokes DNA lesions that interfere with replication and transcription. These lesions may compromise cell viability and usually are removed by nucleotide excision repair (NER). In humans, inactivation of NER is associated with three rare autosomal recessive inherited disorders: xeroderma pigmentosum (XP), Cockayne syndrome, and trichothiodystrophy. The NER earliest step is lesion recognition by a complex formed by XPC and HHR23B proteins. In a subsequent step, XPA protein becomes associated to the repair complex. Here we investigate whether XPA and XPC proteins, involved in global genome repair, may contribute to a signal transduction pathway regulating the response to UVC-induced lesions. We monitored the expression of several UVC-induced genes in cells deficient in either a transduction pathway or mutated on an NER gene. Expression of the KIN17 gene is induced after UVC irradiation independently of p53 and of activating transcription factor 2. However, in human cells derived from XPA or XPC patients the UVC-induced accumulation of KIN17 RNA and protein is abolished. Our results indicate that the presence of functional XPA and XPC proteins is essential for the up-regulation of the KIN17 gene after UVC irradiation. They also show that the integrity of global genome repair is required to trigger KIN17 gene expression and probably other UVC-responsive genes.

Keywords: melanoma‖XPA‖XPC‖genotoxic stress ‖ nuclear proteins

Exposure of human cells to UV radiation provokes DNA lesions that interfere with DNA replication and transcription. Nucleotide excision repair (NER) is an efficient way to remove these lesions (13). The NER mechanism is well characterized, and various models have been proposed (46). In humans, NER inactivation is associated with three autosomal recessive inherited disorders: xeroderma pigmentosum (XP), Cockayne syndrome (CS), and trichothiodystrophy (7). NER possesses two major overlapping subpathways: global genome repair (GGR) and transcription-coupled repair (TCR). The earliest biochemical step of GGR is lesion recognition by a complex formed by XPC and HHR23B proteins (1). Thereafter, TFIIH is recruited at the DNA lesion followed by other NER proteins such as XPG, XPA, RPA, ERCC-1, and XPF. The sequential assembling of these components is due in part to the double role of XPA protein in facilitating the association of ERCC1–XPF and in activating the incision activity of XPG protein (4, 8, 9). This biochemical model is in accord with the fact that cells from XPC patients are GGR-deficient and TCR-proficient (1, 10, 11). After UVC irradiation, XPC-deficient cells display an almost normal massive activation of several genes and proteins, indicating that the DNA-repair deficiency of these cells does not affect the inducible response or that the normal TCR is sufficient to allow a nearly wild-type UV response (12, 13). Interestingly, UVC-irradiated GGR-deficient XPC cells also display the normal stabilization of p53 protein (14, 15), underlying the need to characterize the UVC-induced signal transduction pathway(s) further. Indeed, UVC activates receptors of tyrosine kinases and protein kinases of the plasma membrane leading to the activation of transcription factors (16). The direct effects of UVC in the nucleus, however, are less well characterized. Indeed, the presence of pyrimidine dimers in transcribed regions of the genome enhances the expression of several genes (13). However, functional DNA repair as well as the presence of repair intermediates do not seem essential for gene activation (2, 13, 1719).

We investigated whether XPA and XPC proteins may contribute to the activation of genes of DNA metabolism. We determined the expression pattern of P21 and GADD45, genes involved in the UVC response, in a p53-dependent manner together with that of c-fos and KIN17, a UVC-responsive gene with expression that is under a tight and complex regulation at several levels including mRNA and protein stabilization (ref. 19 and L. Miccoli, unpublished results). The KIN17 gene codes an evolutionarily conserved zinc-finger nuclear protein that is distributed in nucleoplasmic foci (20). The human kin17 protein physically interacts with simian virus 40 T-antigen and is a component of a high molecular weight complex that is essential for DNA replication (21). The direct association of human kin17 protein to chromosomal DNA during cell proliferation and its colocalization with RPA protein in nucleoplasmic foci in the presence of DNA damage after ionizing radiation suggest a role in DNA repair (22, 23). It therefore seemed important to understand better how the expression of kin17 protein is regulated after exposure of cells to UVC. Because activating transcription factor 2 (ATF2) mediates KIN17 response to ionizing radiation (22), a dominant mutant of ATF2 was used to test a possible contribution to KIN17 gene up-regulation after UVC irradiation. In parallel, we analyzed the levels of KIN17 RNA and protein in p53-, XPA-, XPC-, or CS-deficient cells after UVC irradiation. We show that XPA and XPC proteins may contribute to the regulation of gene expression, and the expression of the KIN17 gene is triggered by an ATF2- and p53-independent pathway. Our results indicate that two GGR proteins may participate in a signal transduction pathway.

Materials and Methods

Cell Lines and Culture Conditions.

Human melanoma MeWo and Δ196-4 cells were provided by Z. Ronai (The Ruttenberg Cancer Center, Mount Sinai Medical School, New York). Cells were cultured in DMEM (GIBCO/Life Technologies) supplemented with 10% FCS/penicillin (100 units/ml)/streptomycin (100 μg/ml), and in the case of Δ196-4 cells, 200 μg/ml G418 (geneticin, GIBCO/BRL) were added. Diploid fibroblasts from unexposed skin biopsy of a normal young child (405VI), XPA fetus (AS162 and AS456), or XPC fetus (XP202VI) were established as described (24). XP44RO cells came from the laboratory of D. Bootsma (Erasmus University, Rotterdam) and were cultured in MEM modified with l-glutamine and 15% FCS (25). XP44RO-XPC cells constructed in our laboratory were cultured with 500 μg/ml G418. MCF7 cells from a human breast carcinoma and their tumor necrosis factor-resistant derivative (26) were gifts from E. May (DRR, Commissariat à l'Energie Atomique) and S. Chouaib (Institute Gustave Roussy, Villejuif, France). MCF7/R-A1 carries an R280K mutation in the P53 gene (27). MCF7 and MCF7/R-A1 were grown at 37°C and 5% CO2 in DMEM (GIBCO) or RPMI medium 1640 (GIBCO), respectively, supplemented with 10% FCS/100 units/ml penicillin/100 μg/ml streptomycin/2 mM l-glutamine.

Genotoxic Treatment and Irradiation Conditions.

Asynchronous exponentially growing cultures (3.5 × 106 to 7 × 106 cells per dish) or cells serum-starved for 24 h in DMEM + 0.25% FCS in 100-mm dishes were washed with PBS, covered with 1 ml of PBS, and irradiated at 254 nm with a fluence rate of 0.3 J/m2s. Dosimetry was performed with the UV radiometer CX-254 (Viber Lourmat, Marne la Vallée, France). After irradiation, the original media were returned to the dishes and incubated further. Alternatively, a stock solution of 0.2 mg/ml mitomycin C (MMC) (Sigma) was diluted with media to the indicated final concentrations. UV- and MMC-treated cells were incubated for the indicated times at 37°C and harvested by using trypsin followed by a centrifugation at 250 × g. The pellets were washed twice with PBS and immediately stored at −80°C, or the total RNA was isolated by using the RNeasy Kit (Qiagen, Valencia, CA). RNA was monitored at 260 nm and integrity-controlled by gel electrophoresis.

DNA-Repair Analysis, Complementation Test, and Construction of the Complemented XPC Cell Line.

Diploid fibroblasts were examined for DNA repair after UVC irradiation as described (24). For the tumoral cell lines, hydroxyurea at a final concentration of 20 mM was added 1 h before UV irradiation and was present during the postirradiation incubation. The complementation group of the XP patient was identified by using the test already reported (28, 29). The XP44RO cell line was stably complemented by introducing the wild-type XPC cDNA as described (30). The complemented cells showed a full recovery of the wild-type unscheduled DNA synthesis (UDS) level and survival.

RT-PCR Analysis.

RT-PCR for ACTIN, β2-microglobulin (β2m), P21, and c-fos was performed as described (22). For GADD45 detection, the primers used (D, 5′-ACGAGGACGACGACAGAGAT-3′, and F, 5′-TCCCGCCAAAACAAATAAG-3′) amplified a 262-bp fragment by using an annealing temperature of 58°C. DNA fragments smaller than 400 bp were amplified as described (31).

Western Blot and Immunocytochemistry.

The proteins of 80,000 cells were separated onto a 12% SDS polyacrylamide gel and transferred onto a nitrocellulose filter as described (22). The supernatant of the hybridoma K58 containing an mAb directed against human kin17 protein was diluted 1/200 in TBS + 0.05% Tween-20 as described (21, 22). mAbK44, another mAb against human kin17 protein, systematically confirmed the specificity of the Western patterns. Immunocytochemical detection of kin17 protein was performed on cells that were UVC-irradiated or treated with chemicals as described above by using previously described conditions (22).


UVC and MMC Increase KIN17 Gene Expression in Human Melanoma-Derived Cells.

UVC irradiation of human primary fibroblasts at early passages up-regulates the KIN17 gene within 16 h as shown by Northern blot (20). We checked whether this may be extended to the radioresistant tumor cell line MeWo. These cells established from a late-stage human melanoma are highly metastatic, radioresistant, and DNA repair-proficient (32). Levels of KIN17 RNA were monitored by RT-PCR (22). The amount of β2m RNA varied <5% in all the experiments presented here and was used to normalize the variation of KIN17 RNA. MeWo cells growth-arrested by a 24-h serum deprivation were UVC-irradiated at 15 J/m2, and RNA was monitored during 24 h. We observed a 3-fold increase in KIN17 RNA 16 h after irradiation at a dose of 15 J/m2 (Fig. (Fig.11A). The kinetics were similar to those of primary human fibroblasts (20). Doses ranging from 5 to 10 J/m2 led to small variations in KIN17 RNA levels; only doses higher than 10 J/m2 provoked a significant increase (Fig. (Fig.11A).

Figure 1
UVC and MMC up-regulate KIN17 gene expression in melanoma MeWo cells. Cells serum-starved for 24 h were treated as indicated below. (A) Time course of KIN17 RNA levels after UVC irradiation. After irradiation with 15 J/m2, cells were harvested ...

We further asked whether MMC, an antitumor drug producing inter- and intrastrand covalent bonds in DNA molecules that can also be repaired by NER, was able to up-regulate KIN17 gene expression. Doses ranging from 1 to 5 μg/ml did not produce variations in KIN17 RNA levels; only doses higher than 7 μg/ml provoked a significant increase. KIN17 RNA levels increased 2.5-fold after 30 min of incubation with 10 μg/ml MMC and slowly decreased during the following 24 h. The up-regulation of KIN17 RNA within 30 min was observed for concentrations starting at 7 μg/ml (Fig. (Fig.11B). Under the same conditions, P21 and GADD45 RNA increased within 3 h after incubation (Fig. (Fig.11B), in agreement with the expression profile previously reported for these genes (3335). This result shows that at least one of the adducts formed after MMC activation was recognized by a DNA-damage sensor pathway that finally up-regulated KIN17 RNA and suggests that the response to interstrand covalent bonds is faster than the response to remaining pyrimidine dimers. Our data agree with the previous finding that MMC adducts are differentially recognized in human cells (36). We conclude that in MeWo cells, the KIN17 gene is up-regulated after genotoxic treatment as it is in primary human fibroblasts, and MeWo cells may be used to further characterize the stress response.

The Transcription Factors p53 and ATF2 Are Irrelevant for the UVC-Induced Up-Regulation of the KIN17 Gene.

p53 transcriptionally regulates nearly 100 genes of the UVC response (18, 37, 38). Some of these genes present kinetics similar to those of KIN17 after irradiation of DNA repair-proficient cells. We asked whether p53 may regulate KIN17 gene expression. We compared KIN17 RNA levels in MCF7, an established human cell proficient for p53 with those of MCF7/R-A1 cells carrying a mutated allele of the p53 gene (26, 27). KIN17 RNA levels increased 6-fold 16 h after UVC irradiation at 15 J/m2 in both types of cells (Fig. (Fig.22A). In parallel, we detected a 5-fold increase in P21 RNA levels in p53-proficient cells, whereas no significant increase was observed in p53-deficient cells (Fig. (Fig.22A). Our results in human cells confirm and extend previous observations on the expression of the KIN17 gene in mouse embryo fibroblasts from mice with an inactivated p53 gene (19).

Figure 2
The transcription factors p53 and ATF2 are irrelevant for the UVC-induced up-regulation of the KIN17 gene. (A) UV-induced up-regulation of KIN17 gene expression in p53-deficient cells. Asynchronous exponentially growing MCF7 (p53+/−) and ...

In MeWo cells, ATF2 protein binds to regulatory elements (39) and modulates the expression of a set of genes involved in the response to ionizing radiation including KIN17 (22). This prompted us to test whether ATF2 regulates KIN17 gene expression after UVC irradiation. We compared the MeWo cell line with its derivative Δ196-4 cell line expressing a truncated ATF2 protein that lacks amino acids 1–195 that forms the transactivation domain (39). These two cell lines present similar basal levels of KIN17 RNA (22), although Δ196-4 cells were more sensitive to UVC than the parental MeWo cells (39). The levels of KIN17 RNA increased 3-fold 16 h after irradiation with a dose of 15 J/m2 in both MeWo and Δ196-4 cells. Therefore, the up-regulation of the KIN17 gene took place in the presence of the dominant negative truncated form of ATF2 (Fig. (Fig.22B). P21 and c-fos RNAs increased within 3 and 0.5 h, respectively, after irradiation of both MeWo and Δ196-4 cells, with kinetics similar to those reported for other human cells (12, 19, 35). We conclude that ATF2 and p53 are not involved in the up-regulation of the KIN17 gene after UVC irradiation.

GGR-Deficient Human Primary Fibroblasts Failed to Up-Regulate KIN17 Gene Expression After UVC Irradiation.

We asked whether the up-regulation of the KIN17 gene depended on the capacity to eliminate DNA damage. We therefore compared the expression profile of the KIN17 gene in nontransformed diploid XPC-deficient fibroblasts (XP202VI) with that of fibroblasts from a DNA repair-proficient individual (405VI) after irradiation at 7 and 15 J/m2, respectively. The dose of 7 J/m2 on XPC cells induced a similar cell killing as 15 J/m2 in wild-type cells. P21 RNA levels increased with similar kinetics in the two cell types (Fig. (Fig.33A). The induction kinetics of P21 RNA were very similar to those reported previously for other XPC primary fibroblasts (12). XPC-proficient cells (405VI) presented KIN17 RNA levels that started to increase 8 h after irradiation. A maximal 3-fold increase was observed at 16 h and a slight decrease within 24 h (Fig. (Fig.33A). These expression kinetics are similar to those reported for human primary fibroblasts and mouse cells (19, 20). XP202VI cells deficient for XPC were irradiated under the same conditions and displayed a maximum 1.5-fold increase in KIN17 RNA (Fig. (Fig.33A). Thereafter, we checked whether nontransformed diploid XPA-deficient primary fibroblasts (AS162) were able to up-regulate the KIN17 gene. AS162 cells were analyzed together with 405VI and XP202VI fibroblasts (NER-proficient and XPC-deficient, respectively). After irradiation with increasing UVC doses and incubation for 16 h, cells were harvested and RNA was analyzed. KIN17 RNA levels increased 5-fold at 10 J/m2, 4-fold at 15 J/m2 in 405VI fibroblasts, and only 1.9-fold at 5 J/m2 and 2.3-fold at 10 J/m2 in XP202VI cells. In contrast, a small decrease in KIN17 RNA levels was detected for AS162 cells. P21 RNA was induced in the range of 0–15 J/m2 for the wild-type 405VI and 0–10 J/m2 for XP202VI, whereas in irradiated AS162 cells KIN17 RNA was barely detectable and the P21 level remained nearly constant (Fig. (Fig.33B). Similar results were obtained with AS456, another nontransformed XPA-deficient fibroblast (data not shown). We assumed that the extreme UV sensitivity of XPA-deficient cells was the cause of the observed decrease in KIN17 RNA levels. Indeed, at 5 J/m2 the survival of AS162 cells was <0.2% compared with 100% and 50% for 405VI and XP202VI, respectively (data not shown). It has been shown that XPA-deficient cells present a maximal UVC induction of collagenase I and metallothionein mRNA exclusively at 2 J/m2 as compared with other XP- or CS-deficient primary fibroblasts. Indeed, irradiation of XPA-deficient cells at doses of 5 J/m2 or higher leads to a drastic decrease of these mRNAs (13). We therefore analyzed AS162 cells after irradiation at 2 J/m2. The level of KIN17 RNA determined by RT-PCR systematically varied <10% (G.P.-L., unpublished results). In the same samples the levels of kin17 protein normalized to the actin level varied <10% after irradiation, whereas p53 and proliferating cell nuclear antigen levels increased 200% and 300%, respectively. Immunocytochemical detection failed to show any significant difference in the subcellular distribution of the kin17 protein (G.P.-L., unpublished results). In contrast, under these experimental conditions UV irradiation of CS cells (CS539VI), known to be GGR-proficient and TCR-deficient, provoked an up-regulation of KIN17 RNA similar to that of wild-type cells, indicating that the increase in gene expression does not require TCR (data not shown). We conclude that the presence of the wild-type XPA and XPC proteins correlates with the capacity to accumulate KIN17 RNA after UVC irradiation. The absence of XPA or XPC proteins clearly decreases the amplitude of this response. In contrast, the presence of CS proteins is not required for the up-regulation of this gene. These results indicate the participation of XPA and XPC proteins in a UVC-induced signaling pathway in human primary fibroblasts.

Figure 3
Human primary XPA and XPC fibroblasts are unable to up-regulate KIN17 gene expression after UVC irradiation. (A) Time course of KIN17 RNA levels in irradiated fibroblasts. 405VI and XP202VI cells in the fourth passage were serum-starved for 24 h and irradiated ...

Complementation of XPC-Deficient Melanoma Restores the UV-Induced Up-Regulation of the KIN17 Gene.

To test further the role of XPC, we used XP44RO cells, the only tumor cell line established from a testicular melanoma from an XPC patient (25). A retrovirus carrying the wild-type XPC cDNA under the control of a viral LTR was transduced in XP44RO cells. Among the antibiotic-resistant colonies, we selected and characterized a cell line presenting a wild-type phenotype named here XP44RO-XPC. The determination of UDS levels and survival further confirmed that XP44RO-XPC cells are NER-proficient cells (Fig. (Fig.44A and data not shown). We compared the up-regulation of P21, GADD45, and KIN17 RNAs in XP44RO-XPC and XP44RO cells for a 24-h period after irradiation at 15 J/m2. P21 and GADD45 RNAs presented a maximal 7-fold increase 8 h after irradiation in both cell types (Fig. (Fig.44B), although XPC-deficient cells displayed slightly longer kinetics consistent with previously reported data (12). In XP44RO-XPC cells, KIN17 RNA levels started to increase 8 h after irradiation. A maximal 3-fold increase was observed at 16 h and a slight decrease within 24 h (Fig. (Fig.44B). This expression pattern was similar to that of XPC-proficient melanoma (Fig. (Fig.11A) and to those reported previously in mouse embryo fibroblasts (19). Interestingly, under the same conditions XP44RO cells displayed a low and nearly constant level of KIN17 RNA (Fig. (Fig.44B). We determined protein levels by Western blot. Cells irradiated at 15 J/m2 were harvested at different times, and proteins were detected with mAbs. The level of kin17 clearly increased within 8 h followed by a 3-fold maximal level at 16 h in XP44RO-XPC (Fig. (Fig.44B). Under the same conditions, the XP44RO cells displayed low and constant levels of kin17 protein (Fig. (Fig.44B). These data indicate that the increase in KIN17 RNA was followed by enhanced levels of kin17 protein only in GGR-proficient cells. Because immunocytochemical detection showed that UVC irradiation provokes the intranuclear redistribution of human kin17 protein (20), we used mAbK58 to detect kin17 protein in the XP44RO cells after irradiation at 15 J/m2. At the beginning, XP44RO and XP44RO-XPC cells displayed a similar diffuse nucleoplasmic staining (Fig. (Fig.44C, 0 h). We observed a clear increase in staining intensity in the cytoplasm and nucleus of XP44RO-XPC cells 8 h after UV irradiation (Fig. (Fig.44C, 8 h). The nucleoplasm presented readily distinguishable foci of diverse diameters (Fig. (Fig.44C, 8 h). Under these conditions, XP44RO cells presented a weaker staining pattern (Fig. (Fig.44C, 8 h). Twenty-four hours after irradiation, the staining was similar to that observed at the beginning of the experiment (data not shown). Therefore, the immunocytochemical detection of kin17 protein confirms the increased expression of the KIN17 gene in XP44RO-XPC cells after irradiation. Our data also suggest that the nucleoplasmic relocalization of kin17 protein takes place by a mechanism that is independent of the NER status of the cell. It has been shown that lower doses are needed to activate UVC-responsive genes in NER-deficient cells (12). Taking into account this observation, we compared KIN17 RNA and protein levels 16 h after irradiation at doses ranging from 0 to 5, 10, 15, and 20 J/m2. XP44RO cells presented low and constant levels of KIN17 RNA and protein, indicating that XPC protein contributes to a signaling pathway (Fig. (Fig.44D). Interestingly, XPC-deficient primary fibroblasts (XP202VI, Fig. Fig.33B) still presented a small increase in the KIN17 RNA level compared with transformed XP44RO cells (Fig. (Fig.44D). The precise nucleotide mutated in XP44RO and XP202VI cells remains to be determined. It is likely that these XPC mutations are different because these cells were established from a testicular metastasis and fetus, respectively. This idea is supported also by the fact that the encoded XPC proteins were barely detected by Western blot, suggesting that the mutations carried by these two cell lines resulted in truncated unstable XPC proteins (data not shown). Complemented XP44RO-XPC cells displayed a clear enhanced expression of the KIN17 gene after 15 and 20 J/m2. This expression pattern was very similar to those of DNA-proficient melanoma-derived cells (Figs. (Figs.11A and and44D). All these results show that XPC-deficient primary fibroblasts and cells derived from human melanoma from an XPC patient are unable to up-regulate the KIN17 gene after UVC irradiation. Complementation of these cells by expression of XPC cDNA fully restored their DNA-repair capacity and the UVC responsiveness of the KIN17 gene. These data suggest a role of XPC protein in a signaling pathway leading to the activation of a set of genes of the UVC response.

Figure 4
XPC dependence of KIN17 expression after UVC irradiation. (A) UDS analysis of NER efficiency in human cells. UDS was measured as described in Materials and Methods and quantified as the number of silver grains in the nucleus of irradiated over nonirradiated ...


The role of the human XPA and XPC proteins in the detection of DNA lesions during NER has been characterized in vitro and in vivo (1, 9, 10). We show here that the activity of XPA and XPC proteins is required to trigger one of the signaling pathways leading to the up-regulation of UVC-responsive genes. Several groups have shown that UVC irradiation of NER-deficient human primary fibroblasts enhances the levels of proteins such as p53 and p21 or RNAs such as GADD45, collagenase I, and metallothionein. All these results indicate that the kinetics and amplitudes are characteristic for each complementation type and that signal transduction is elicited mostly by cyclobutane pyrimidine dimers preferentially located on the transcribed strand. It has been suggested also that neither specific DNA intermediates nor ongoing repair are required for signal generation and transduction to promoters (12, 13, 40). We confirm and extend these observations to the human KIN17 gene, which participates in a conserved response to several genotoxic agents (20, 22, 23).

We have characterized the UVC response of melanoma cells from an XPC patient presenting a severe NER deficiency (UDS <10% of the proficient cells). These cells were fully complemented after introduction of XPC cDNA. Complemented cells are normal as judged by sensitivity to UVC irradiation and UDS and are stable at least for a period of >3 months in culture. Despite the striking differences in their DNA-repair capacity, XP44RO and XP44RO-XPC present similar expression profiles of P21 and GADD45 RNA after UVC irradiation (Fig. (Fig.44B), confirming previous observations (12, 13). These results further support the idea that the activation of the UVC-responsive genes partially depends on TCR efficiency, which is normal in XPC cells. This view is consistent with the fact that UVC irradiation activates several distinct regulatory mechanisms involving not only p53 but also mitogen-activated protein kinases, nuclear factor κB, and other proteins that may activate genes with protective effects (18, 19, 41, 42). Interestingly, the analysis of XPC-deficient melanoma and XPC-deficient human primary fibroblasts shows that the up-regulation of the KIN17 gene after UVC irradiation depends on the presence of a normal XPC gene. Other physical and chemical genotoxics such as ionizing radiation, bleomycin, and MMC also up-regulate the KIN17 gene, although with different kinetics and probably by different mechanisms (refs. 22 and 23 and Fig. Fig.1).1). These results indicate that the KIN17 gene may help to overcome damage provoked by several types of genotoxic stress and provide further support to the idea that kin17 protein may contribute to bypass the perturbation of DNA replication produced by unrepaired lesions (22, 23). The human and mouse NER-proficient cells analyzed thus far present a UVC-induced accumulation of KIN17 RNA very similar to that reported previously for wild-type primary fibroblasts (19, 20), thus highlighting the evolutionary conservation of this response.

The idea that the integrity of GGR is crucial for the up-regulation of the KIN17 gene and probably other UV-responsive genes is supported by the following observations.

  • (i) The up-regulation of the KIN17 gene starts 8 h after irradiation, by which time 99% of pyrimidine-pyrimidone(6-4) photoproducts are already eliminated, but >50% of cyclobutane pyrimidine dimers, preferentially repaired by GGR, still remain (2, 43, 44).
  • (ii) GGR-proficient and TCR-deficient human CS cells present a UVC-induced accumulation of KIN17 RNA similar to that of NER-proficient cells.
  • (iii) GGR- and TCR-deficient human XPA cells (AS162 and AS456) are unable to up-regulate KIN17 RNA levels and after irradiation at low UVC doses present constant levels of kin17 protein.

Considering that kin17 protein may participate in essential functions such as transcription and DNA replication and/or repair (21, 23, 45), it is reasonable to postulate that a subset of the complexes formed at the site of DNA lesions during GGR is the primary signal that leads to the modulation of KIN17 gene expression and probably of other UV-responsive genes. These results now enable dissection of the molecular chain of events going from DNA lesion formation to the activation of a particular set of genes of the UV response. The use of other human cells deficient in different DNA-repair pathways will help to shed some light on this GGR-dependent signaling pathway and to specify further the role of the KIN17 gene in the response to genotoxic agents.


We thank Z. Ronai for his MeWo cells and extensive advice and J. Grassi for support. We warmly thank A. Benoit for cell culture and M. C. Nevers, P. Lamourette, and M. Plaissance for antibody preparations. The support of B. Dutrillaux is gratefully acknowledged. C.M. received fellowships from the Commissariat à l'Energie Atomique, Electricité de France (EDF) and Association pour la Recherche sur le Cancer. This work was funded by EDF Contract 8702.


nucleotide excision repair
xeroderma pigmentosum
Cockayne syndrome
global genome repair
transcription-coupled repair
activating transcription factor 2
mitomycin C
unscheduled DNA synthesis


This paper was submitted directly (Track II) to the PNAS office.


1. Sugasawa K, Ng J M, Masutani C, Iwai S, van der Spek P J, Eker A P, Hanaoka F, Bootsma D, Hoeijmakers J H. Mol Cell. 1998;2:223–232. [PubMed]
2. Friedberg E C, Walker G C, Siede W. DNA Repair and Mutagenesis. Washington, DC: Am. Soc. Microbiol.; 1995.
3. Hoeijmakers J H. Nature. 2001;411:366–374. [PubMed]
4. Batty D, Rapic'-Otrin V, Levine A S, Wood R D. J Mol Biol. 2000;300:275–290. [PubMed]
5. Batty D P, Wood R D. Gene. 2000;241:193–204. [PubMed]
6. Sugasawa K, Okamoto T, Shimizu Y, Masutani C, Iwai S, Hanaoka F. Genes Dev. 2001;15:507–521. [PMC free article] [PubMed]
7. de Boer J, Hoeijmakers J H. Carcinogenesis. 2000;21:453–460. [PubMed]
8. Yokoi M, Masutani C, Maekawa T, Sugasawa K, Ohkuma Y, Hanaoka F. J Biol Chem. 2000;275:9870–9875. [PubMed]
9. Volker M, Mone M J, Karmakar P, van Hoffen A, Schul W, Vermeulen W, Hoeijmakers J H, van Driel R, van Zeeland A A, Mullenders L H. Mol Cell. 2001;8:213–224. [PubMed]
10. Venema J, van Hoffen A, Karcagi V, Natarajan A T, van Zeeland A A, Mullenders L H. Mol Cell Biol. 1991;11:4128–4134. [PMC free article] [PubMed]
11. Venema J, van Hoffen A, Natarajan A T, van Zeeland A A, Mullenders L H. Nucleic Acids Res. 1990;18:443–448. [PMC free article] [PubMed]
12. Dumaz N, Duthu A, Ehrhart J C, Drougard C, Appella E, Anderson C W, May P, Sarasin A, Daya-Grosjean L. Mol Carcinog. 1997;20:340–347. [PubMed]
13. Blattner C, Bender K, Herrlich P, Rahmsdorf H J. Oncogene. 1998;16:2827–2834. [PubMed]
14. Ford J M, Hanawalt P C. J Biol Chem. 1997;272:28073–28080. [PubMed]
15. Smith M L, Ford J M, Hollander M C, Bortnick R A, Amundson S A, Seo Y R, Deng C X, Hanawalt P C, Fornace A J., Jr Mol Cell Biol. 2000;20:3705–3714. [PMC free article] [PubMed]
16. Devary Y, Gottlieb R A, Smeal T, Karin M. Cell. 1992;71:1081–1091. [PubMed]
17. Herrlich P, Sachsenmaier C, Radler-Pohl A, Gebel S, Blattner C, Rahmsdorf H J. Adv Enzyme Regul. 1994;34:381–395. [PubMed]
18. May P, May E. Oncogene. 1999;18:7621–7636. [PubMed]
19. Blattner C, Kannouche P, Litfin M, Bender K, Rahmsdorf H J, Angulo J F, Herrlich P. Mol Cell Biol. 2000;20:3616–3625. [PMC free article] [PubMed]
20. Kannouche P, Mauffrey P, Pinon-Lataillade, Mattei M-G, Sarasin A L, D-G, Angulo J F. Carcinogenesis. 2000;21:1701–1710. [PubMed]
21. Miccoli L, Biard D S F, Creminon C, Angulo J F. Cancer Res. 2002;62:5425–5436. [PubMed]
22. Masson C, Menaa F, Pinon-Lataillade G, Frobert Y, Pablo Radicella J, Angulo J F. Radiat Res. 2001;156:535–544. [PubMed]
23. Biard D S, Miccoli L, Despras E, Frobert Y, Creminon C, Angulo J F. J Biol Chem. 2002;277:19156–19165. [PubMed]
24. Sarasin A, Blanchet-Bardon C, Renault G, Lehmann A, Arlett C, Dumez Y. Br J Dermatol. 1992;127:485–491. [PubMed]
25. Keijzer W, Mulder M P, Langeveld J C, Smit E M, Bos J L, Bootsma D, Hoeijmakers J H. Cancer Res. 1989;49:1229–1235. [PubMed]
26. Cai Z, Stancou R, Korner M, Chouaib S. Int J Cancer. 1996;68:535–546. [PubMed]
27. Cai Z, Capoulade C, Moyret-Lalle C, Amor-Gueret M, Feunteun J, Larsen A K, Paillerets B B, Chouaib S. Oncogene. 1997;15:2817–2826. [PubMed]
28. Carreau M, Eveno E, Quilliet X, Chevalier-Lagente O, Benoit A, Tanganelli B, Stefanini M, Vermeulen W, Hoeijmakers J H, Sarasin A, et al. Carcinogenesis. 1995;16:1003–1009. [PubMed]
29. Zeng L, Sarasin A, Mezzina M. Methods Mol Biol. 1999;113:87–100. [PubMed]
30. Carreau M, Quilliet X, Eveno E, Salvetti A, Danos O, Heard J M, Mezzina M, Sarasin A. Hum Gene Ther. 1995;6:1307–1315. [PubMed]
31. Chevillard S, Muller A, Levalois C, Laine-Bidron C, Vielh P, Magdelenat H. Breast Cancer Res Treat. 1996;41:81–89. [PubMed]
32. Yang Y M, Ronai Z. Mol Carcinog. 1995;14:111–117. [PubMed]
33. Butz K, Geisen C, Ullmann A, Zentgraf H, Hoppe-Seyler F. Oncogene. 1998;17:781–787. [PubMed]
34. Butz K, Whitaker N, Denk C, Ullmann A, Geisen C, Hoppe-Seyler F. Oncogene. 1999;18:2381–2386. [PubMed]
35. Wang W, Furneaux H, Cheng H, Caldwell M C, Hutter D, Liu Y, Holbrook N, Gorospe M. Mol Cell Biol. 2000;20:760–769. [PMC free article] [PubMed]
36. Abbas T, Olivier M, Lopez J, Houser S, Xiao G, Kumar G S, Tomasz M, Bargonetti J. J Biol Chem. 2002;277:40513–40519. [PubMed]
37. Maltzman W, Czyzyk L. Mol Cell Biol. 1984;4:1689–1694. [PMC free article] [PubMed]
38. Smith M L, Fornace A J., Jr Proc Natl Acad Sci USA. 1997;94:12255–12257. [PMC free article] [PubMed]
39. Ronai Z, Yang Y, Fuchs S Y, Alder V, Sardana M, Herlyn M. Oncogene. 1998;16:523–531. [PubMed]
40. Lu X, Lane D P. Cell. 1993;75:765–778. [PubMed]
41. Fornace A J, Amundson S A, Bittner M, Myers T G, Meltzer P, Weinsten J N, Trent J. Gene Expression. 1999;7:387–400. [PubMed]
42. Schmidt-Ullrich R K, Dent P, Grant S, Mikkelsen R B, Valerie K. Radiat Res. 2000;153:245–257. [PubMed]
43. Eveno E, Bourre F, Quilliet X, Chevallier-Lagente O, Roza L, Eker A P, Kleijer W J, Nikaido O, Stefanini M, Hoeijmakers J H, et al. Cancer Res. 1995;55:4325–4332. [PubMed]
44. Emmert S, Kobayashi N, Khan S G, Kraemer K H. Proc Natl Acad Sci USA. 2000;97:2151–2156. [PMC free article] [PubMed]
45. Timchenko T, Bailone A, Devoret R. EMBO J. 1996;15:3986–3992. [PMC free article] [PubMed]

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