In 1994 and 1995 two geneswere identified that
confer susceptibility to breast and ovarian
cancer when mutated, the breast cancer genes
BRCA1 and BRCA2. Both genes encode multifunctional
proteins with important cellular
functions in genomic stability, homologous recombination,
and double-stranded and transcription-
coupled DNA repair (see p. 80). The
BRCA1 and BRCA2 proteins interact and play a
role in cell cycle control (see p. 112) and in
development. An autosomal dominant susceptibility
mutant allele in one of these genes is
considered the main cause of the cancer in
about 5–10% of patients. Mutations in other
genes are involved in some cases. The direct
causative role of BRCA1 and BRCA2 mutations is
difficult to assess in individual patients. Different
mutations as well as polymorphic variants
occur throughout the genes.
Sunday, April 12, 2009
The breast cancer susceptibility gene BRCA1
The BRCA1 gene on chromosome 17 at q21.1
consists of 24 exons spanning 80 kb of genomic
DNA that encode a 7.8 kb mRNA transcript. The
protein has 1863 amino acids. Exon 11 is quite
large (3.4 kb). About 55% of all mutations occur
in exon 11. Although some mutations occur
relatively frequently in other exons, they tend to
be evenly distributed throughout the gene (only
some mutations are shown). The deletion of an
adenine (A) and a guanine (G) in nucleotide
position 185 (185delAG) and the insertion of a
cytosine in position 5382 (5382insC) account
for about 10% of mutations each. These mutations
are particularly frequent in the Ashkenazi
Jewish population.
The protein has five main functional domains.
The RING finger region near the N-terminus at
amino acids 1–112 defines a zinc-binding
domain of conserved cysteine and histidine residues
that mediate protein—protein or protein-
DNA interactions. This region is also the site of
heterodimerization of BRCA1 and BARD1
(BRCA1-associated RING domain 1). Other
functional domains define the central part of
the BRCA1 protein. These are two nuclear localization
signals (NLS) and two protein-binding
domains, one for p53 protein, retinoblastoma
(RB) protein, and RAD50 and RAD51. RAD50 and
51 are proteins involved in recombination
during mitosis and meiosis, and in recombinational
repair of double-stranded DNA breaks.
The C-terminus contains a region involved in
transcriptional activation and DNA repair.
consists of 24 exons spanning 80 kb of genomic
DNA that encode a 7.8 kb mRNA transcript. The
protein has 1863 amino acids. Exon 11 is quite
large (3.4 kb). About 55% of all mutations occur
in exon 11. Although some mutations occur
relatively frequently in other exons, they tend to
be evenly distributed throughout the gene (only
some mutations are shown). The deletion of an
adenine (A) and a guanine (G) in nucleotide
position 185 (185delAG) and the insertion of a
cytosine in position 5382 (5382insC) account
for about 10% of mutations each. These mutations
are particularly frequent in the Ashkenazi
Jewish population.
The protein has five main functional domains.
The RING finger region near the N-terminus at
amino acids 1–112 defines a zinc-binding
domain of conserved cysteine and histidine residues
that mediate protein—protein or protein-
DNA interactions. This region is also the site of
heterodimerization of BRCA1 and BARD1
(BRCA1-associated RING domain 1). Other
functional domains define the central part of
the BRCA1 protein. These are two nuclear localization
signals (NLS) and two protein-binding
domains, one for p53 protein, retinoblastoma
(RB) protein, and RAD50 and RAD51. RAD50 and
51 are proteins involved in recombination
during mitosis and meiosis, and in recombinational
repair of double-stranded DNA breaks.
The C-terminus contains a region involved in
transcriptional activation and DNA repair.
The breast cancer susceptibility gene BRCA2
The BRCA2 gene on 13q12 comprises 27 exons
spanning 80 kb of genomic DNA that encode a
10.4 kb mRNA transcript. Its protein has 3418
amino acids. Exon 11 is large (11.5 kb), as in
BRCA1. Mutations occur throughout the gene
(only some are shown). A deletion of thymine at
nucleotide position 6174 (6174delT) is relatively
(1%) frequent in the Ashkenazi Jewish
population.
The BRCA2 protein has a transcriptional activation
domain near the N-terminus and a nuclear
location signal (NLS) near the C-terminus. A
large central domain consists of eight copies of
a 30–80-amino-acid repeat, which are conserved
in all mammalian BRCA2 proteins (BRC
repeats).
spanning 80 kb of genomic DNA that encode a
10.4 kb mRNA transcript. Its protein has 3418
amino acids. Exon 11 is large (11.5 kb), as in
BRCA1. Mutations occur throughout the gene
(only some are shown). A deletion of thymine at
nucleotide position 6174 (6174delT) is relatively
(1%) frequent in the Ashkenazi Jewish
population.
The BRCA2 protein has a transcriptional activation
domain near the N-terminus and a nuclear
location signal (NLS) near the C-terminus. A
large central domain consists of eight copies of
a 30–80-amino-acid repeat, which are conserved
in all mammalian BRCA2 proteins (BRC
repeats).
BRCA1 and BRCA2 genes
The BRCA1 and BRCA2 genes are expressed
ubiquitously with the highest levels of expression
in thymus and testis. The spatial and temporal
expression patterns of Brca1 and Brca2 in
the mouse fetal and adult tissues are essentially
identical, with highest expression of both in
rapidly dividing tissues during differentiation,
especially in mammary epithelium. In the
mammary gland both genes are expressed
during puberty and pregnancy, and their expression
is reduced during lacation.
ubiquitously with the highest levels of expression
in thymus and testis. The spatial and temporal
expression patterns of Brca1 and Brca2 in
the mouse fetal and adult tissues are essentially
identical, with highest expression of both in
rapidly dividing tissues during differentiation,
especially in mammary epithelium. In the
mammary gland both genes are expressed
during puberty and pregnancy, and their expression
is reduced during lacation.
Retinoblastoma
Retinoblastoma (McKusick 180200) is the most
frequent tumor of the eye in infancy and early
childhood. It occurs in 1 of 15000–18000 live
births. This tumor results from loss of function
of both alleles of the retinoblastoma gene RB1.
Tumor initiation is preceded by two steps as A.
Knudson predicted in 1971 in his “two-hit” hypothesis
(tumor suppressor gene, p. 318). The
first predisposing mutation in one allele may
occur either in a retinoblast, an undifferentiated
retinal cell in the developing embryo, or
in the germline. The other allele is inactivated
by a second mutation.
frequent tumor of the eye in infancy and early
childhood. It occurs in 1 of 15000–18000 live
births. This tumor results from loss of function
of both alleles of the retinoblastoma gene RB1.
Tumor initiation is preceded by two steps as A.
Knudson predicted in 1971 in his “two-hit” hypothesis
(tumor suppressor gene, p. 318). The
first predisposing mutation in one allele may
occur either in a retinoblast, an undifferentiated
retinal cell in the developing embryo, or
in the germline. The other allele is inactivated
by a second mutation.
Phenotype
Retinoblastoma occurs in one eye or both eyes.
An important early sign is the so-called “cat’s
eye,” awhite shimmer out of the affected eye (1)
or the development of strabismus. One or
several tumors originate from the retina (2).
The tumor progresses rapidly (3). The relative
proportions of the genetic types of retinoblastoma
are about 60% somatic mutations (nonhereditary
form) and 40% germline mutations,
transmitted as an autosomal dominant trait
(hereditary form, in about 10–15%, due to
transmission from a parent; the remainder due
to a new mutation). New mutations usually affect
a paternal allele (about 10: 1). In about 10%
of carriers of a germline mutation no tumor
develops (nonpenetrance).
An important early sign is the so-called “cat’s
eye,” awhite shimmer out of the affected eye (1)
or the development of strabismus. One or
several tumors originate from the retina (2).
The tumor progresses rapidly (3). The relative
proportions of the genetic types of retinoblastoma
are about 60% somatic mutations (nonhereditary
form) and 40% germline mutations,
transmitted as an autosomal dominant trait
(hereditary form, in about 10–15%, due to
transmission from a parent; the remainder due
to a new mutation). New mutations usually affect
a paternal allele (about 10: 1). In about 10%
of carriers of a germline mutation no tumor
develops (nonpenetrance).
Retinoblastoma locus on chromosome 13
The RB1 locus at 13q14.2 was first defined with
cytogenetically visible interstitial deletions.
cytogenetically visible interstitial deletions.
Retinoblastoma gene BR-1 and the pRB protein
The RB1 gene is organized into 27 exons spanning
183 kb of genomic DNA (1). The RB1 gene is
ubiquitously expressed and transcribed into a
4.7 kb mRNA (2). The gene product (pRB protein)
has 928 amino acids (3). It is a 100 kD
phosphoprotein with important functions in
the regulation of the cell cycle. It is activated by
phosphorylation (P) during cell cycle progression
from G0 to G1 (p. 112) at about 12 distinct
serine and threonine residues. Three
functional domains, A, B, and C, and a nuclear
localization signal (NLS) can be distinguished.
183 kb of genomic DNA (1). The RB1 gene is
ubiquitously expressed and transcribed into a
4.7 kb mRNA (2). The gene product (pRB protein)
has 928 amino acids (3). It is a 100 kD
phosphoprotein with important functions in
the regulation of the cell cycle. It is activated by
phosphorylation (P) during cell cycle progression
from G0 to G1 (p. 112) at about 12 distinct
serine and threonine residues. Three
functional domains, A, B, and C, and a nuclear
localization signal (NLS) can be distinguished.
D. Diagnostic principle
Molecular diagnosis of retinoblastoma greatly
contributes to its early recognition and to the
correct assessment of individual risks within
families. In about 3–5% of patients an interstitial
deletion 13q14 or a larger deletion is visible
by chromosomal analysis (1). In familial retinoblastoma
indirect DNA diagnosis can be
achieved by segregation analysis using DNA
markers at the RB1 locus (2). In the example
shown, the affected girl (II-1) has inherited haplotype
a from the unaffected father and haplotype
c from the unaffected mother. In tumor
cells, obtained after one eye had to be removed,
haplotype a only is present (loss of heterozygosity,
LOH, see p. 318). This reveals that haplotype
a represents the mutation-carrying RB1 allele.
In the family shown (3), I-2 and
II-2 are affected (3). Sequence analysis reveals a
C-to-T transversion in codon 575 in the two affected
individuals (CAA glutamine to TAA stop
codon). The mutational spectrum in hereditary
retinoblastoma involves deletions (~26%), insertions
(~9%), and point mutations (~65%), including
splice-site mutations.
contributes to its early recognition and to the
correct assessment of individual risks within
families. In about 3–5% of patients an interstitial
deletion 13q14 or a larger deletion is visible
by chromosomal analysis (1). In familial retinoblastoma
indirect DNA diagnosis can be
achieved by segregation analysis using DNA
markers at the RB1 locus (2). In the example
shown, the affected girl (II-1) has inherited haplotype
a from the unaffected father and haplotype
c from the unaffected mother. In tumor
cells, obtained after one eye had to be removed,
haplotype a only is present (loss of heterozygosity,
LOH, see p. 318). This reveals that haplotype
a represents the mutation-carrying RB1 allele.
In the family shown (3), I-2 and
II-2 are affected (3). Sequence analysis reveals a
C-to-T transversion in codon 575 in the two affected
individuals (CAA glutamine to TAA stop
codon). The mutational spectrum in hereditary
retinoblastoma involves deletions (~26%), insertions
(~9%), and point mutations (~65%), including
splice-site mutations.
Fusion Gene as Cause of Tumors: CML
Chronicmyeloid leukemia (CML) is amalignant
tumor that originates from a single cell of the
bone marrow in adulthood. The number of myelocytes
(white blood cells from the bone marrow)
is greatly increased. The disease follows a
chronic course. Acute crises develop intermittently
and terminally. In about 90% of the
patients, affected bone marrow cells contain a
chromosome 22 with a shortened long arm
(22q–, Philadelphia chromosome).
tumor that originates from a single cell of the
bone marrow in adulthood. The number of myelocytes
(white blood cells from the bone marrow)
is greatly increased. The disease follows a
chronic course. Acute crises develop intermittently
and terminally. In about 90% of the
patients, affected bone marrow cells contain a
chromosome 22 with a shortened long arm
(22q–, Philadelphia chromosome).
The Philadelphia chromosome (Ph1) in different forms of leukemia
A Philadelphia chromosome is present in the
bone marrow cells of most patients with the
chronic form of the disease (CML). If it is not
present, the illness progresses more rapidly
than usual and has a poorer prognosis. In addition,
the Philadelphia chromosome may be
found in some acute leukemias (acute lymphocytic
leukemia, ALL; acutemyelocytic leukemia,
AML) in adults and in children. Here, Ph1 indicates
a poor prognosis, whereas its absence is
favorable.
bone marrow cells of most patients with the
chronic form of the disease (CML). If it is not
present, the illness progresses more rapidly
than usual and has a poorer prognosis. In addition,
the Philadelphia chromosome may be
found in some acute leukemias (acute lymphocytic
leukemia, ALL; acutemyelocytic leukemia,
AML) in adults and in children. Here, Ph1 indicates
a poor prognosis, whereas its absence is
favorable.
The Ph1 translocation [t(9;22)(q34;q11)]
The Philadelphia chromosome arises by reciprocal
translocation between a chromosome 22
and a chromosome 9. The breakpoints are in
9q34 and 22q11. A good half of the long arm of a
chromosome 22 is translocated to the long arm
of a chromosome 9. A very small segment of the
distal long arm of a chromosome 9 (9q34), not
visible under the light microscope, is translocated
to a chromosome 22. The Philadelphia
chromosome (22q–) consists of the short arm
and the proximal one-third of the long arm of a
chromosome 22 and the small distal segment
from the long arm of a chromosome 9. For demonstration
of the Philadelphia translocation
translocation between a chromosome 22
and a chromosome 9. The breakpoints are in
9q34 and 22q11. A good half of the long arm of a
chromosome 22 is translocated to the long arm
of a chromosome 9. A very small segment of the
distal long arm of a chromosome 9 (9q34), not
visible under the light microscope, is translocated
to a chromosome 22. The Philadelphia
chromosome (22q–) consists of the short arm
and the proximal one-third of the long arm of a
chromosome 22 and the small distal segment
from the long arm of a chromosome 9. For demonstration
of the Philadelphia translocation
The Ph1 translocation leads to the fusion of two gen
The breakpoints of the Ph1 translocation are located
in the BCR gene of chromosome 22 and in
the ABL gene of chromosome 9. The translocation
leads to the fusion of these genes. The exact
locations of the breakpoints differ from patient
to patient, but in the BCR gene they are limited
to a small region of just 6 kb (thus the designation
BCR, or breakpoint cluster region). In CML,
the breakpoints lie in exons 10–12 of the BCR
gene; in acute Ph1-positive leukemias (e.g., ALL)
they lie further in the 5! direction in exon 1 or 2.
The breakpoint region in the ABL gene extends
over 180 kb between exons 1a and 1b, which are
separated by an intron.
in the BCR gene of chromosome 22 and in
the ABL gene of chromosome 9. The translocation
leads to the fusion of these genes. The exact
locations of the breakpoints differ from patient
to patient, but in the BCR gene they are limited
to a small region of just 6 kb (thus the designation
BCR, or breakpoint cluster region). In CML,
the breakpoints lie in exons 10–12 of the BCR
gene; in acute Ph1-positive leukemias (e.g., ALL)
they lie further in the 5! direction in exon 1 or 2.
The breakpoint region in the ABL gene extends
over 180 kb between exons 1a and 1b, which are
separated by an intron.
The gene fusion leads to changes in transcription and gene products
The ABL gene codes formRNA transcripts of 7 kb
(exon 1b, 2–11) and 6 kb (exon 1a, 2–11) by
differential splicing; these in turn code for a
protein of about 145000 Da (p145abl). From the
fusion of the two genes in CML, an 8.5 kb mRNA
transcript results, which codes for a fusion protein
of 210000 Da (p210bcr/abl). In the acute form
of leukemia (ALL), a transcript results that codes
for a fusion protein of 190000 Da (p190bcr/abl). In
contrast to the normal protein, it has high tyrosinase
activity. This results in uncontrolled
cell division in the affected cells and tumor
growth.
(exon 1b, 2–11) and 6 kb (exon 1a, 2–11) by
differential splicing; these in turn code for a
protein of about 145000 Da (p145abl). From the
fusion of the two genes in CML, an 8.5 kb mRNA
transcript results, which codes for a fusion protein
of 210000 Da (p210bcr/abl). In the acute form
of leukemia (ALL), a transcript results that codes
for a fusion protein of 190000 Da (p190bcr/abl). In
contrast to the normal protein, it has high tyrosinase
activity. This results in uncontrolled
cell division in the affected cells and tumor
growth.
Genomic Instability Syndromes
Genomic instability, visible by light microscopy
as breaks and rearrangements in different chromosomes
in a variable proportion of metaphase
cells, is a hallmark of a group of characteristic
hereditary diseases. The underlying genetic defect
predisposes patients to different types of
cancer. Three important examples are presented
here.
as breaks and rearrangements in different chromosomes
in a variable proportion of metaphase
cells, is a hallmark of a group of characteristic
hereditary diseases. The underlying genetic defect
predisposes patients to different types of
cancer. Three important examples are presented
here.
Bloom syndrome (BS)
In Bloom syndrome (McKusick 210900) (1), prenatal
and postnatal growth deficiency is pronounced
(birth weight 2000 g, birth length
!40 cm, adult height around 150 cm). The
phenotype (2) includes a narrow face. Usually,
but not always, a sunlight-induced erythema
develops on the cheeks, eyelids, mouth, ears,
and back of the hands (a and b). The photograph
on the right (c) shows a boy with Bloom syndrome
and acute leukemia. Metaphase cells
show about a tenfold increase in the rate of sister
chromatid exchanges (SCE), !60 instead of
about 6 per metaphase in normal cells (3). (Sister
chromatid exchanges are explained in the
glossary, p. 423). Metaphases of patients contain
increased breaks in one or both chromatids
and exchanges between homologous chromosomes
in about 1–2% of cells. In Bloom syndrome
patients, different types of malignancies
occur in a distribution comparable to that of the
general population, but at a much earlier age
(mean age 24.7, range 2–48 years). Some
patients have multiple primary tumors, which
underlines the striking susceptibility to cancer
in Bloom syndrome. Chemotherapy is very
poorly tolerated.
and postnatal growth deficiency is pronounced
(birth weight 2000 g, birth length
!40 cm, adult height around 150 cm). The
phenotype (2) includes a narrow face. Usually,
but not always, a sunlight-induced erythema
develops on the cheeks, eyelids, mouth, ears,
and back of the hands (a and b). The photograph
on the right (c) shows a boy with Bloom syndrome
and acute leukemia. Metaphase cells
show about a tenfold increase in the rate of sister
chromatid exchanges (SCE), !60 instead of
about 6 per metaphase in normal cells (3). (Sister
chromatid exchanges are explained in the
glossary, p. 423). Metaphases of patients contain
increased breaks in one or both chromatids
and exchanges between homologous chromosomes
in about 1–2% of cells. In Bloom syndrome
patients, different types of malignancies
occur in a distribution comparable to that of the
general population, but at a much earlier age
(mean age 24.7, range 2–48 years). Some
patients have multiple primary tumors, which
underlines the striking susceptibility to cancer
in Bloom syndrome. Chemotherapy is very
poorly tolerated.
Homozygosity
Homozygosity for mutations in the Bloom syndrome
gene (BLM) results in an increased rate of
somatic mutations, a manifestation of genomic
instability. The BLM gene on chromosome 15 at
q16.1 encodes a member of the RecQ family of
DNA helicases. The 1417-amino-acid protein
shows homology to the yeast SGS1 gene product
(slow growth suppressor) and the human WRN
gene product (Werner syndrome, McKusick
277700). Allozygous nonsense mutations (two
different mutations in the two alleles of the
same gene) are frequent in the BLM gene. A
characteristic homozygous 6 bp deletion/7 bp
insertion at nucleotide 2281 occurs in Ashkenazi
Jewish
gene (BLM) results in an increased rate of
somatic mutations, a manifestation of genomic
instability. The BLM gene on chromosome 15 at
q16.1 encodes a member of the RecQ family of
DNA helicases. The 1417-amino-acid protein
shows homology to the yeast SGS1 gene product
(slow growth suppressor) and the human WRN
gene product (Werner syndrome, McKusick
277700). Allozygous nonsense mutations (two
different mutations in the two alleles of the
same gene) are frequent in the BLM gene. A
characteristic homozygous 6 bp deletion/7 bp
insertion at nucleotide 2281 occurs in Ashkenazi
Jewish
Fanconi anemia (FA)
Fanconi anemia (hereditary pancytopenia)
(McKusick 227650) is a malformation syndrome
(1) with variable clinical expression.
Growth deficiency (2), hypoplastic or absent
thumbs (3), and short or absent radii are characteristic
physical signs.
FA cells are hypersensitive to DNA-crosslinking
agents, such as diepoxybutane (DEB). Several
complementation groups can be distinguished.
Three FA genes have been identified, at chromosome
16q24.3 (FAA), 9q22.3 (FAC), and 3p22–
26 (FAD). FAA is the most prevalent group in
60–65% of patients.
(McKusick 227650) is a malformation syndrome
(1) with variable clinical expression.
Growth deficiency (2), hypoplastic or absent
thumbs (3), and short or absent radii are characteristic
physical signs.
FA cells are hypersensitive to DNA-crosslinking
agents, such as diepoxybutane (DEB). Several
complementation groups can be distinguished.
Three FA genes have been identified, at chromosome
16q24.3 (FAA), 9q22.3 (FAC), and 3p22–
26 (FAD). FAA is the most prevalent group in
60–65% of patients.
Ataxia telangiectasia
Ataxia telangiectasia (McKusick 208900) is a
pleiotropic, variable disease due to mutations in
the ATM gene on chromosome 11q23. Preferential
reciprocal translocations between chromosomes
7 and 14 with breakpoints at 7p14, 7q14,
14q11, and 14q32 occur in a small proportion of
metaphases. The ATM gene has 66 exons spanning
more than 150 kb of genomic DNA. From
its 13-kb transcript (and smaller alternatively
spliced products), a 3056-amino-acid protein
kinase ATM(350 kDA) is translated. ATM is activated
in response to double-strand DNA breaks.
It is part of a network of proteins that regulate
cellular responses to DNA damage (p. 80). Clinically
different disorders are related at the cellular
level
pleiotropic, variable disease due to mutations in
the ATM gene on chromosome 11q23. Preferential
reciprocal translocations between chromosomes
7 and 14 with breakpoints at 7p14, 7q14,
14q11, and 14q32 occur in a small proportion of
metaphases. The ATM gene has 66 exons spanning
more than 150 kb of genomic DNA. From
its 13-kb transcript (and smaller alternatively
spliced products), a 3056-amino-acid protein
kinase ATM(350 kDA) is translated. ATM is activated
in response to double-strand DNA breaks.
It is part of a network of proteins that regulate
cellular responses to DNA damage (p. 80). Clinically
different disorders are related at the cellular
level
Subscribe to:
Comments (Atom)
