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© 2000 Oxford University Press
Human Molecular Genetics, 2000, Vol. 9, No. 8 1171–1175
Identical mutations in the CSB gene associated with
either Cockayne syndrome or the DeSanctis–
Cacchione variant of xeroderma pigmentosum
Stefano Colella, Tiziana Nardo, Elena Botta, Alan R. Lehmann1 and Miria Stefanini+
Istituto di Genetica Biochimica ed Evoluzionistica CNR, Via Abbiategrasso, 207-27100 Pavia, Italy and 1MRC Cell
Mutation Unit, Sussex University, Falmer, Brighton BN1 9RR, UK
Received 26 November 1999; Revised and Accepted 13 March 2000
Xeroderma pigmentosum (XP) and Cockayne
syndrome (CS) are two hereditary disorders in which
photosensitivity is associated with distinct clinical
and cellular phenotypes and results from genetically
different defects. We have identified the primary
molecular alteration in two patients in whom clinical
manifestations strongly reminiscent of a severe form
of XP were unexpectedly associated with the CS
cellular phenotype and with a defect in the CSB gene.
Sequencing of the CSB-coding region in both cDNA
and genomic DNA showed that these patients had
identical alterations to those in a patient with the clinical features of the classical form of CS. These data,
together with fluorescence in situ hybridization analysis, demonstrated that the two siblings with XP as
well as the CS patient were homozygous for the same
CSB mutated allele, containing a silent C2830T
change and a nonsense mutation C2282T converting
Arg735 to a stop codon. The finding that the same
inactivating mutation underlies different pathological
phenotypes indicates that there is no simple correlation between the molecular defect and the clinical
features. Therefore, alterations in the CSB gene give
rise to the same repair defect at the cellular level but
other genetic and/or environmental factors determine the pathological phenotype.
INTRODUCTION
Xeroderma pigmentosum (XP) and Cockayne syndrome (CS)
are two autosomal, recessive disorders in which hypersensitivity to UV light is associated with distinct clinical and
cellular phenotypes and results from genetically different
defects (see ref. 1 for a recent review). XP is characterized by
hypersensitivity to sun-exposure, pigmentary alterations and
premalignant lesions in sun-exposed areas of the skin, and an
extremely high incidence of skin cancer. Approximately 20%
of XP patients show neurological abnormalities of varying
severity due to primary neuronal degeneration. The biochemical defect in most XP patients is in nucleotide-excision repair
(NER) and typically results in a reduced capability to perform
+To
UV-induced DNA repair synthesis (UDS). A defect in postreplication repair is present in a minority of cases, the so-called
XP variant (XP-V) group characterized by a defect in the
ability to synthesize intact daughter DNA strands after UV
irradiation. Genetic analysis by somatic cell hybridization has
led to the identification in the NER-defective form of XP of
seven complementation groups, designated XP-A to XP-G.
CS is a multisystem disorder characterized by postnatal
growth failure, progressive neurological dysfunction due to
demyelination, premature ageing and otherwise clinically
heterogeneous features which commonly include cutaneous
photosensitivity but not cancer. Cells from CS patients are often
hypersensitive to UV light and are unable to recover normal
RNA synthesis rates following UV irradiation, despite having
normal levels of UDS. Two complementation groups, CS-A and
CS-B, have been identified; both groups are specifically defective in the sub-pathway of NER which rapidly removes damage
from the transcribed strand of active genes (2).
Rare patients have been described in which the cutaneous
alterations of XP are combined with some of the major clinical
symptoms of CS, namely pigmentary retinal degeneration,
primary demyelination, calcification of the basal ganglia and
cachetic dwarfism. All these patients, assigned to a clinical
entity designated XP/CS, show the cellular phenotype typical
of XP, i.e. reduced UDS level, and they have been assigned to
the XP-B, XP-D and XP-G groups (1).
Clinical features reminiscent of the DeSanctis–Cacchione
syndrome (DSC), the form of XP with severe neurological
involvement, have been recently described in three siblings
from an Hispanic family. Detailed DNA repair investigations
in fibroblast strains from the two youngest patients, coded
GM10903 and GM10905, surprisingly showed that the cellular
response to UV light in both siblings was similar to that
typically described in CS without any evidence of defects
indicative either of the NER-defective or the variant form of
XP (3). Following genetic analysis they have been assigned to
the complementation group B of CS (4). Therefore, they represent the first XP cases assigned to this group, which comprises
about 30 subjects with CS features (5–10).
The cloning of the CSB gene has enabled mutations to be
identified in patients classified in the CS-B group (11). In the 22
CS cases previously analysed by us, many of the inactivating
whom correspondence should be addressed. Tel: +39 0382 546330; Fax +39 0382 422286; Email: [email protected]
1172 Human Molecular Genetics, 2000, Vol. 9, No. 8
Figure 1. Mutations found in the CSB cDNA and resulting protein alterations in the GM10903 and GM10905 patients. The diagram shows the CSB cDNA [white area:
open reading frame (ORF); black areas: untranslated regions] and the CSB protein with the predicted functional domains (11). Causative mutations are indicated in bold.
mutations resulted in severely truncated polypeptides because of
either stop codons, frameshifts or splice abnormalities (9,10).
In this paper we report the results of the molecular analysis
of the CSB gene in the patients GM10903 and GM10905.
RESULTS AND DISCUSSION
Sequence analysis of the CSB gene was performed on fibroblast strains obtained from the siblings GM10903 and
GM10905 (3). Genetic analysis by somatic cell hybridization
assigned both siblings to the CS-B complementation group (4).
To identify the molecular alterations, the sequence of the
coding region of the CSB gene was analysed by RT–PCR
followed by direct sequencing (nucleotides 80–4558 in ref.
11). Both patients showed in the whole of the amplified cDNA
population a silent C2830T change (Gly917) and a nonsense
mutation C2282T, which converts Arg735 to a stop codon
(opal), resulting in the synthesis of a truncated protein of 734
amino acids (Fig. 1).
We showed previously that the Turkish patient CS1TAN was
homozygous and the Caucasian patient 25627 heterozygous for
these same mutations at the cDNA level (9). Both these cases
showed the clinical features of the classical form of CS.
The patient CS1TAN was born from a consanguineous
marriage whereas consanguinity between GM10903 and
GM10905 parents could only be inferred from the recurrence
of the same family name in both maternal and paternal lines
and from the fact that both families had lived in the same small
Mexican village for many years (3). Since cells from the
parents were not available, we took different approaches to
demonstrate that the inactivating mutation was present on both
the CSB alleles of the patients. As detailed in Materials and
Methods, the identity of the cell strains was checked by both
karyotypic and fingerprint analysis. Sequencing of the relevant
genomic DNA region showed only the mutant T allele at position 2282 in all three patients (GM10903, GM10905 and
CS1TAN) suggesting that they were all homozygous for the
C2282T mutation (Fig. 2). To confirm that this was indeed the
case and to exclude the possibility of a deletion including the
CSB gene in one of the two alleles, we carried out fluorescence
in situ hybridization (FISH) analysis with a probe specific for
the CSB gene. In all three cases we obtained two positive
signals on nuclei and specific hybridization on two chromo-
Figure 2. Mutations found in the CSB genomic DNA of the patients GM10903 and
CS1TAN. Autoradiograph of the sequencing gel of the PCR products obtained
after amplification of the DNA region containing the C2282T mutation in a
control, GM10903 and CS1TAN.
somes (Fig. 3). These observations rule out the possibility of a
deletion in this region and confirm that GM10903, GM10905
and CS1TAN patients carry two CSB alleles, both containing
the same inactivating mutation. Therefore, the intriguing genotype–phenotype relationship in the siblings GM10903 and
GM10905 cannot be related simply either to the specific CSB
mutation or to the dosage of the mutated allele.
The mutation in the CSB alleles in these patients results in an
altered cellular response to UV, typical of CS but it underlies a
clinical phenotype suggestive of DSC. As detailed by Moriwaki et al. (12), DSC cases can be distinguished from those
with CS by the absence of retinal degeneration, decreased to
absent deep tendon reflexes, primary neuronal degeneration,
and absence of calcification of the basal ganglia and other
brain structures. Patients with CS, in contrast, do not show XPtype skin freckling or skin cancer but do have increased or
normal deep tendon reflexes, signs of primary demyelination
and often calcification of the brain.
As well as showing the absence of the major clinical criteria
for the diagnosis of CS, extensive clinical evaluation of the
siblings GM10903 and GM10905 strongly suggested they
have DSC (3). The DSC patients so far described all show the
cellular phenotype typical of XP and are mutated in the XPA or
Human Molecular Genetics, 2000, Vol. 9, No. 8 1173
Figure 3. Fluorescence in situ hybridization with the cDNA of the CSB gene on
mitotic cells and interphase nuclei from the patients GM10903 (upper panels) and
CS1TAN (lower panels). The hybridization signals correspond to the yellow spots.
Arrows point to the chromosomes showing the hybridization signal.
XPD gene. Cellular and genetic DNA repair investigations in
the siblings GM10903 and GM10905 argue against the presence of an additional mutation in one of the XP genes identified so far (3,4). Therefore, GM10903 and GM10905 represent
the first cases in which the severe neurological alterations
diagnostic for DSC are associated with mutations in the CSB
gene. Interestingly, despite the presence of the neurological
anomalies typical of XP, in the patients GM10903 and
GM10905, the cutaneous symptoms of XP are limited to
actinic skin changes. Therefore, in these patients, as in CS
patients, the presence of mutated CSB alleles does not result in
any increased incidence of cancer. It has been suggested
recently that the CSB protein might have an additional function in transcription beyond its involvement in coupling repair
to blocked transcription (13,14). We are tempted to speculate
that mutations in the CSB gene might interfere with some of
the steps leading to neoplastic transformation and progression,
which may be dependent on the transcriptional role of CSB
protein.
Although there are a few reports in the literature of the same
mutation being linked to two different clinical entities (e.g. refs
15,16), this is unprecedented for syndromes associated with
DNA repair defects. Molecular analysis of NER-defective XP
patients defective in the XPB, XPD and XPG genes has shown
that specific mutations are associated with particular pathological phenotypes. This holds true for patients with trichothiodystrophy (TTD) or XP/CS altered in the XPB gene (17,18), for
XP, TTD and XP/CS cases altered in the XPD gene (19–27), and
for XP and XP/CS altered in the XPG gene (28,29). The
patients reported in this paper provide the first evidence that
the same inactivating mutation in the CSB gene underlies
distinct pathological phenotypes diagnostic, respectively, for
CS (CS1TAN) and for a severe form of XP (GM10903 and
GM10905). These observations imply that there is not any
obvious and direct correlation between the molecular defect in
the CSB gene and the clinical features. Our previous molecular
analysis of the CSB gene in several CS patients with clinically
heterogeneous features has already provided data suggesting
that other factors, besides the site of mutation, influence the
type and severity of the CS pathological phenotype (9,10).
This notion is further extended by the results reported in this
paper showing that the same set of mutated CSB alleles is associated with distinct clinical entities.
What might the alterations be in the genetic background of
the GM10903 and GM10905 patients that hamper the expression of the mutated CSB alleles at the phenotypic but not at the
cellular level? Despite the presence of the neurological abnormalities typical of XP, two pieces of experimental evidence
argue against the presence of an altered XP gene. The patients
show the clinical symptoms but not the cellular features of XP,
as clearly demonstrated by the results reported in ref. (3).
Furthermore, at least in the transgenic mouse system, the association of XP and CS defects (CSB–/–XPA–/– and CSB–/–XPC–/–
double knock-out mice) has a strong synergistic effect at the
phenotypic level and leads to dramatically pronounced CS
features (30; G.T.H. van der Host, unpublished data).
The GM10903 and GM10905 patients might be mutated in a
gene, or a panel of genes, that is able to confer neurological
abnormalities indistinguishable from those resulting from XP
defects but does not have any effect on repair. In addition, as
suggested above, the absence of a functional CSB protein
might per se be a sufficient condition to prevent carcinogenesis. Current hypotheses propose that the neurological abnormalities typical of CS result from subtle transcription
alterations and/or from an inability to repair oxidative damage
(reviewed in ref. 31). It is possible that the genetic background
in the GM10903 and GM10905 patients suppresses or
compensates for the deleterious effects of the CSB defect on
transcription or repair of oxidative damage, thereby preventing
expression of the neurological abnormalities typical of CS.
MATERIALS AND METHODS
Cells and culture conditions
The study was performed on cells from patients GM10903 and
GM10905 purchased from the NIGMS Human Genetic Mutant
Cell Repository (Camden, NJ). Fibroblasts were routinely
grown in Ham’s F-10 medium (Gibco-BRL, Rockville, MD)
supplemented with 12% fetal calf serum (FCS; Irvine, Santa
Ana, CA) and subcultured by trypsinization. The identity of
the cell strains was ascertained both at the cellular and molecular levels. The analysis of four hypervariable regions in the
genomic DNA samples used for sequencing showed that
GM10903 and GM10905 have the same set of FGA and TH01
alleles (23–26 and 7–8, respectively) and share one allele of
VWA (GM10903, 18–19; GM10905, 15–18) and D21S11
(GM10903, 31.2–32.2; GM10905, 30–32.2). A different
combination of alleles for all the four analysed regions was
observed in CS1TAN, namely FGA, 20–24; TH01, 8–9;
VWA, 17–18 and D21S11, 29–31.2. Molecular analysis of a
locus allowing for sex-discrimination (amelogenin) and karyotype analysis on cell samples parallel to those used for FISH
indicated that GM10903 is a female whereas CS1TAN and
GM10905 are males, as expected.
1174 Human Molecular Genetics, 2000, Vol. 9, No. 8
Identification of the molecular alteration
ACKNOWLEDGEMENTS
RNA extraction, cDNA synthesis and PCR amplification of
the cDNA of the CSB gene were carried out using protocols
and primers described in detail in ref. (9). Briefly, 2 µg of RNA
in a total volume of 10 µl were heated to 90°C for 2 min and
incubated at 37°C for 1 h following addition of 30 µl of a mix
containing 1× first strand cDNA buffer (Gibco-BRL), 10 mM
DTT, 1mM dNTPs, 25 ng oligo(dT)15 (Promega, Madison,
WI) and 200 U Moloney Murine Leukemia Virus Reverse
Transcriptase (M-MuLV RT, Gibco-BRL). Samples were then
heated to 95°C for 5 min, made up to 100 µl with water and
stored at –20°C. cDNA was amplified by PCR into six overlapping fragments; PCR mixtures contained 5–25 µl of cDNA, 1×
Gene Amp buffer II (Perkin Elmer, Norwalk, CT), 2 mM
MgCl2, 0.2 mM dNTPs, 20 pmol each of the required primers
and 2U AmpliTaq (Perkin Elmer) in a total volume of 50 µl.
Amplification was carried out for 35 cycles of 94°C for 1 min,
annealing at 65°C (fragment I), 63°C (fragment II) or 67°C
(fragment III–VI) for 1.5 min and 72°C for 3 min.
Genomic DNA was isolated from samples of 5 × 106 fibroblasts by phenol extraction and isopropanol precipitation,
following an overnight incubation at 37°C with 6 ml lysis
buffer (150 mM NaCl, 10 mM Tris–HCl, pH 8.0; 1 mM
EDTA, 150 µg/ml proteinase K, 0.5% SDS) and RNase treatment for 1 h at 37°C. Genomic DNA amplification of the
region containing the identified mutation was carried out on
0.5 µg samples using the primer pairs A (5′-TAATGTTCCCTTCTCTGCTCTTATTAAAGG-3′ in Intron 10) and B (5′-ATCTGGCAAAGAAAGGCTCATCTTGACATC-3′ in Exon 11).
The fragment was amplified using AmpliTaq Gold (Perkin
Elmer) and the following conditions: 1 cycle at 95°C for 4 min
and 35 cycles at 95°C for 1 min, 60°C for 10 min.
PCR products were purified by agarose gel electrophoresis
and directly sequenced using the ThermoSequenase cycle
sequencing kit (Amersham Pharmacia Biotech, Uppsala,
Sweden).
We are grateful to Dr Wim Vermeulen (Erasmus University,
Rotterdam) for providing us with the pSLME6 plasmid and to
Dr Carlo Previderè (Department of Legal Medicine, University, Pavia) for fingerprint analysis. This work was supported
in part by Telethon grant E.550 to M.S. and EC contracts
BMH4-CT98-3045 and QLG1-1999-00181 to A.R.L. and
M.S.
Fluorescence in situ hybridization
The pSLME6 plasmid containing the whole of the CSB cDNA
(11) was used as probe. Plasmid DNA was labelled with
biotin-16-dUTP by nick translation according to the supplier’s
instructions (Gibco-BRL) and purified through a Sephadex
G50 column (Amersham Pharmacia Biotech). Hybridization
was carried out according to methods described by Lichter and
Cremer (32). Metaphase spreads and nuclei were denatured for
3 min at 70°C in 70% formamide, 2× SSC and then the slides
were dehydrated at 4°C. The labelled probe (200 ng/slide) was
resuspended in a hybridization mixture (30 µl/slide) containing
50% deionized formamide, 2× SSC, 10% dextran sulfate, 100×
excess of salmon sperm DNA, denatured at 70°C for 10 min
and then applied to slides under a 24 × 50 mm coverslip. After
incubation for 16 h at 37°C the slides were washed three times
for 5 min in 50% formamide, 2× SSC and three times for 5 min
in 2× SSC. All washes were at 42°C. Hybridization signals
were revealed using fluoresceinated avidin. Metaphases and
nuclei were counterstained with propidium iodide and
examined using a Leitz Orthoplan fluorescence microscope;
photographs were taken on colour print film and digitized
using the Adobe Photoshop 4.0 software.
REFERENCES
1. Bootsma, D., Kraemer, K.H., Cleaver, J. and Hoeijmakers, J.H.J. (1998)
Nucleotide excision repair syndromes: xeroderma pigmentosum, Cockayne
syndrome and trichothiodystrophy. In Vogelstein, B. and Kinzler, K.W.
(eds), The Genetic Basis of Human Cancer. McGraw-Hill, New York, NY,
pp. 245–274.
2. van Hoffen, A., Natarajan, A.T., Mayne, L.V., Van Zeeland, A.A.,
Mullenders, L.H.F. and Venema, J. (1993) Deficient repair of the
transcribed stand of active genes in Cockayne’s syndrome cells. Nucleic
Acids Res., 21, 5890–5895.
3. Greenhaw, G.A., Hebert, A., Duke-Woodside, M.E., Butler, I.J., Hecht,
J.T., Cleaver, J.E., Thomas, G.H. and Horton W.A. (1992) Xeroderma
pigmentosum and Cockayne syndrome: overlapping clinical and
biochemical phenotypes. Am. J. Hum. Genet., 50, 677–689.
4. Itoh, T., Cleaver, J.E. and Yamaizumi, M. (1996) Cockayne syndrome
complementation group B associated with xeroderma pigmentosum
phenotype. Hum. Genet., 97, 176–179.
5. Tanaka, K., Kawai, Y., Kumahara, Y., Ikenaga, M. and Okada, Y. (1981)
Genetic complementation groups in Cockayne syndrome. Somat. Cell.
Genet., 7, 445–455.
6. Lehmann, A.R. (1982) Three complementation groups in Cockayne
syndrome. Mutat. Res., 106, 347–356.
7. Miyauchi, H., Horio, T., Akaeda, T., Asada, Y., Chang, H.R., Ishizaki, K.
and Ikenaga, M. (1994) Cockayne syndrome in two adult siblings. J. Am.
Acad. Dermatol., 30, 329–335.
8. Stefanini, M., Fawcett, H., Botta, E., Nardo, T. and Lehmann, A.R. (1996)
Genetic analysis of UV hypersensitivity in twenty-two patients with
Cockayne syndrome. Hum. Genet., 97, 418–423.
9. Mallery, D.L., Tanganelli, B., Colella, S., Steingrimsdottir, H., van Gool,
A.J., Troelstra, C., Stefanini M. and Lehmann, A.R. (1998) Molecular
analysis of mutations in the CSB (ERCC6) gene in patients with Cockayne
syndrome. Am. J. Hum. Genet., 62, 77–85.
10. Colella, S., Nardo, T., Mallery, D., Borrone, C., Ricci, R., Ruffa, G.,
Lehmann, A. R. and Stefanini, M. (1999) Cellular and molecular analysis of
three non photosensitive Italian patients with the severe form of Cockayne
syndrome (CS) and alterations in the CSB gene. Hum. Mol. Genet., 8, 935–
941.
11. Troelstra, C., van Gool, A.J., de Wit, J., Vermulen, W., Bootsma, D. and
Hoeijmakers, J.H.J. (1992) ERCC6, a member of a subfamily of putative
helicase, is involved in Cockayne’s syndrome and preferential repair of
active genes. Cell, 71, 939–953.
12. Moriwaki, S., Stefanini, M., Lehmann, A.R., Hoeijmakers, J.H.J., Robbins,
J.H., Rapin, I., Botta, E., Tanganelli, B., Vermeulen, W., Broughton, B.C.
and Kraemer, K.H. (1996) DNA repair and ultraviolet mutagenesis in cells
from a new patient with xeroderma pigmentosum group G and Cockayne
syndrome resemble xeroderma pigmentosum cells. J. Invest. Dermatol.,
107, 647–653.
13. Friedberg, E.C. (1996) Cockayne syndrome—a primary defect in DNA
repair, transcription, both or neither? BioEssays, 18, 731–738.
14. vanGool, A.J., van der Horst, G.T.J., Citterio, E. and Hoeijmakers, J.H.J.
(1997) Cockayne syndrome: defective repair of transcription? EMBO J., 16,
4155–4162.
15. Rutland, P., Pulleyn, L.J., Reardon, W., Baraitser, M., Hayward, R., Jones,
B., Malcolm, S., Winter, R.M., Oldridge, M., Slaney, S.F. et al. (1995)
Identical mutations in the FGFR2 gene cause both Pfeiffer and Crouzon
syndrome phenotypes. Nature Genet., 9, 173–176.
16. Wilkie, A.O.M., Slaney, S.F., Oldridge, M., Poole, M.D., Ashworth G.J.,
Hockley A.D., Hayward, R., David D.J., Pulleyn, L.J., Rutland, P. et al.
(1995) Apert syndrome results from localized mutations of FGFR2 and is
allelic with Crouzon syndrome. Nature Genet., 9, 165–172.
Human Molecular Genetics, 2000, Vol. 9, No. 8 1175
17. Vermeulen, W., Scott, R.J., Rodgers, S., Muller, H.J., Cole, J., Arlett, C.F.,
Kleijer W.J., Bootsma, D., Hoeijmakers, J.H.J. and Weeda, G. (1994)
Clinical heterogeneity within xeroderma pigmentosum associated with
mutation in the DNA repair and transcription gene ERCC3. Am. J. Hum.
Genet., 54, 191–200.
18. Weeda, G., Eveno, E., Donker, I., Vermulen, W., Chevalier-Lagente, O.,
Taieb, A., Stary, A., Hoeijmakers, J.H.J., Mezzina, M. and Sarasin, A.
(1997) A mutation in the XPB/ERCC3 DNA repair transcription gene,
associated with trichothiodystrophy. Am. J. Hum. Genet., 60, 320–329.
19. Broughton, B.C., Steingrimsdottir, H., Weber, C.A. and Lehmann, A.R.
(1994) Mutations in the xeroderma pigmentosum group D DNA repair/
transcription gene in patients with trichothiodystrophy. Nature Genet., 7,
189–194.
20. Broughton, B.C., Thompson, A.F., Harcourt, S.A., Vermeulen, W.,
Hoeijmakers, J.H.J., Botta, E., Stefanini, M., King, M.D., Weber, C.A.,
Cole, J., Arlett, C.F. et al. (1995) Molecular and cellular analysis of the
DNA repair defect in a patient in xeroderma pigmentosum group D who has
the clinical features of xeroderma pigmentosum and Cockayne syndrome.
Am. J. Hum. Genet., 56, 167–174.
21. Frederik, G.D., Amirkhan, R.M., Shultz, R.A. and Friedberg, E.C. (1994)
Structural and mutational analysis of the xeroderma pigmentosum group D
(XPD) gene. Hum. Mol. Genet., 3, 1783–1788.
22. Takayama, K., Salazar, E.P., Lehmann, A.R., Stefanini, M., Thompson,
L.H. and Weber C.A. (1995) Defects in DNA repair and transcription gene
ERCC2 in the cancer prone disorder xeroderma pigmentosum group D.
Cancer Res., 55, 5656–5663.
23. Takayama, K., Salazar, E.P., Broughton, B.C., Lehmann, A.R., Sarasin, A.
and Thompson, L.H. (1996) Defects in the DNA repair and transcription
gene ERCC2 (XPD) in trichothiodistrophy. Am. J. Hum. Genet., 58, 263–
270.
24. Takayama, K., Danks, D.M., Salazar, E.P., Cleaver, J.E. and Weber, C.A.
(1997) DNA repair characteristic and mutations in the ERCC2 DNA repair
25.
26.
27.
28.
29.
30.
31.
32.
and transcription gene in a trichothiodistrophy patient. Hum. Mutat., 9, 519–
525.
Kobayashi, T., Kuraoka, I., Saijo, M., Nakatsu, Y., Tanaka, A., Someda, Y.,
Fukuro, S. and Tanaka, K. (1997) Mutations in the XPD gene leading to
xeroderma pigmentosum symptoms. Hum. Mutat., 9, 322–331.
Taylor, E.M., Broughton, B.C., Botta, E., Stefanini, M., Sarasin, A., Jaspers,
N.G.J., Fawcett, H., Harcourt, S.A., Arlett, C.F. and Lehmann, A.R. (1997)
Xeroderma pigmentosum and trichothiodistrophy are associated with
different mutations in the XPD (ERCC2) repair/transcription gene. Proc.
Natl Acad. Sci. USA, 94, 8658–8663.
Botta, E., Nardo, T., Broughton, B., Marinoni, S., Lehmann, A.R. and
Stefanini, M. (1998) Analysis of mutations in the XPD gene in Italian
patients with trichothiodystrophy: site of mutation correlates with repair
deficiency but gene dosage appears to determine clinical severity. Am. J.
Hum. Genet., 63, 1036–1048.
Nouspikel, T. and Clarkson, S.G. (1994) Mutations that disable the DNA
repair gene XPG in a xeroderma pigmentosum group G patient. Hum. Mol.
Genet., 3, 963–967.
Nouspikel, T., Lalle P., Leadon, S.A., Cooper, P.K. and Clarkson, S.G.
(1997) A common mutational pattern in Cockayne syndrome patients from
xeroderma pigmentosum group G: implications for a second XPG function.
Proc. Natl Acad. Sci. USA, 94, 3116–3121.
de Boer, J. and Hoeijmakers, J.H.J. (1999) Cancer from the outside, aging
from the inside: mouse models to study the consequences of defective
nucleotide excision repair. Biochimie, 81, 127–137.
de Boer, J. and Hoeijmakers, J.H.J. (2000) Nucleotide excision repair and
human syndromes. Carcinogenesis, 21, 453–460.
Lichter, P. and Cremer, T. (1992) Chromosome analysis by non-isotopic in
situ hybridization. In Rooney, D.E. and Czepulkowski, B.H. (eds), Human
Cytogenetics: A Practical Approach, 2nd edn. IRL Press, Oxford, Vol., 1,
pp. 157–192.
1176 Human Molecular Genetics, 2000, Vol. 9, No. 8