greig cephalopolysyndactyly syndrome
DESCRIPTION
A presentation on Greig syndromeTRANSCRIPT
Greig Cephalopolysyndactyly Syndrome
Akpene Gbegnon, MS4Department of Pathology
June 5th , 2008
Outline
Greig syndrome Genetic abnormalities in GCPS GLI3 – mutated gene in GCPS Association of GLI3 with limb development Case with GCPS & brain tumor Relationship between GLI3 and Sonic
hedgehog Greig syndrome and medulloblastomas
Greig Cephalopolysyndactyly Syndrome (GCPS)
First described by David Greig in 1926 The incidence is less than 10/1,000,000 It is an autosomal dominant mutation
Phenotype can include: Craniofacial abnormalities
Hypertelorism (increased inter-pupillary distance) Macrocephaly with frontal bossing
Limb abnormalities Syndactyly (webbed fingers or toes) Polydactyly (extra digits)
Developmental delay (uncommon)
Craniofacial and limb findings vary significantly
Cases of Greig Syndrome
1926 – Greig describes digital malformations of mother and daughter with polysyndactyly and high forehead
1978 – McKusick studied a family in which 10 members of 4 generations were affected in a pattern consistent with a fully penetrant autosomal dominant trait.
1981 – Merlob reported a female infant with polydactyly, syndactyly, and craniofacial dysmorphism with frontal bossing.
The father had a high forehead and mild hypertelorism
Genetic Abnormalities Found in Greig Syndrome
Translocation t(3;7) (p21.1;p13) Translocation t(6;7) (q27;p13) Deletion of 7p13-p11.2 Deletion of 7p14-p12.3
Greig syndrome results from a mutation in chromosome 7p13
GLI3 is a gene on chromosome 7p13 that is mutated in Greig syndrome
(1991) GLI3 considered candidate gene since it had been mapped to chromosome 7p13
(1991) Vortcamp demonstrated that 2 of 3 translocations found to be associated with Greig syndrome interrupt the GLI3 gene.
(2000) Infant with Greig syndrome (syndactyly, macrocephaly) with a
father and grandfather with digital abnormalities.
Infant had 2 dominant mutations in the GLI3 gene which was also present in the father and grandfather.
Mutations of the GLI3 gene is associated with Greig syndrome
What is GLI3?
The GLI3 gene: Is one of the three zinc-finger GLI family proteins
(Gli1,Gli2,Gli3) is located on chromosome 7p13 functions as a transcription factor
Is a negative regulator of Sonic hedgehog (Shh) signaling
Is there an association between GLI3 and limb development?
Yes A mouse model of Greig Cephalopolysyndactyly syndrome: the
extra-toes mutation contains an intragenic deletion of the GLI3 gene. Hui et al. Nature Genetics. 1993.
Mouse model is called: extra-toes (Xt)
Phenotype: Craniofacial defects (enlarged interfrontal bone) Polydactyly and syndactyly
Heterozygotes are viable and breed freely Homozygous lethal (polydactyly, syndactyly, malformation of
brain)
Mouse Model of Greig Syndrome
Results The extra-toes (Xt) mutants have decreased
GLI3 expression secondary to a deletion in the 3’ end.
In the early embryo of mice, GLI3 expression is most prominent in the developing limb buds and facial primordia.
In situ hybridization analysisof GLI3 transcript in developing limb.Idm = interdigital mesenchymeIzm = interzonal mesenchyme
Mouse Model of Greig Syndrome
Conclusion Structures affected in the mouse mutant and human
syndrome correlate with the expression of GLI3.
This supports the hypothesis that GLI3 plays a role in limb development, and its mutation leads to the abnormal limb development seen in Greig syndrome.
In situ hybridization analysisof GLI3 transcript in developing head and limbNt = neural tubeHlb = hindlimb bud.
Mouse Model of Greig Syndrome(Litingtung et al., 2002; Welscher et al., 2002)
This supports the hypothesis that GLI3 plays a role in limb development, and its mutation leads to the abnormal limb development seen in Greig syndrome.
Skeleton morphology
GLI3 and limb development(Lallemand et al, 2006)
In the limb, although the three Gli genes are expressed, only Gli3 appears to be necessary for patterning
Case (cont.)
Patient with Greig syndrome and pilocytic astrocytoma
Brain
Tumor (2.7cm X 2.0cm) involving the left cerebellopontine angle, dorsal medulla, and middle cerebellum
Hemorrhage at the resection site
Tumor is present in the cerebellum Asymmetry of the
cerebellum
Shift from left to right side
Left cerebellum is enlarged
Dentate nucleus seen on the right, but not on the left
Upper part of the left also looks enlarged
Left Right
FS of brain tumor showing Pilocytic Astrocytoma
increased cellularity, elongated nuclei fibrillary processes present Rosenthal fibers present
Is there an association between Greig syndrome and tumors?
Unclear…..
There are <200 articles published on Greig syndrome to date < 20 of those discuss Greig syndrome AND cancer
Cases: Medulloblastoma (2) Acute lymphoblastic leukemia (1)
Homer-wright rosettes
Medulloblastoma, WHO grade IV
Tumor of the cerebellumDense cellularityCircular arrangement of cells in rosettesSmall blue cell tumor
GLI3, Shh, and medulloblastomas
Sonic hedgehog (Shh) plays a major role in the developing cerebellum
Activation of the Shh pathway is associated with hereditary and sporadic medulloblastomas.
GLI3 is a negative regulator of Shh signaling
GLI3 and Sonic Hedgehog (Shh)
In Drosophila, hedgehog signaling is mediated by the Cubitus interruptus (Ci) protein. Ci75 is a hedgehog repressor Ci155 is a transcription activator
In birds and mammals, the orthologues of Cubitus interruptus are the three member Gli family (Gli1, Gli2, and Gli3)
GLI3 and Sonic hedge hog
Truncated form of GLI3 (GLI3R) is a repressor of Shh
Another form is possibly an activator
Complicated interaction between GLI3 and Shh, involving other proteins including Hand 2, Gremlin, Bmps, etc.
The relationship between GLI3 and Shh(Robert et al, 2006)
GLI3 is a negative regulator of Shh signaling
Is GLI3 involved in medulloblastomas?
GLI3 is not mutated commonly in sporadic Medulloblastomas. Erez et al. ACS. 2002.
Question: Are mutations in the GLI3 gene associated with medulloblastomas?
Why? Because there were 2 patients with Greig syndrome who had developed medulloblastomas.
Is GLI3 involved in medulloblastomas?
GLI3 is not mutated commonly in sporadic Medulloblastomas. Erez et al. ACS. 2002.
Method: The authors sequenced the GLI3 gene, including all exon-intron boundaries in: 10 medulloblastomas samples from 10 patients 2 human medulloblastoma cell lines 30 control DNA blood samples from healthy donors 1 blood sample from a patient with Greig syndrome and
medulloblastoma. (tumor sample was not available)
Is GLI3 involved in medulloblastomas? Results:
Authors found several new polymorphisms on exon 5 and 15 of GLI3 (CGCCAC, CTGCCG, etc)
Those polymorphisms also occurred in normal individuals with a frequency of > 25%
There were no tumor-associated mutations in the sporadic tumors
Authors found a new nonsense germline mutation (introduction of a stop codon) leading to a severely truncated GLI3 protein in the child with Greig syndrome and medulloblastoma
Conclusion:
GLI3 is “rarely” mutated in medulloblastoma
Summary
Greig syndrome is a rare genetic disorder Patients with Greig syndrome have hypertelorism,
macrocephaly, syndactyly and polydactyly Greig syndrome is caused by a mutation in the GLI3
gene, which is a gene located on chromosome 7p13 GLI3 is a transcription factor expressed in the limb and
skull during development GLI3 is a suppressor of the Sonic hedge hog pathway The association between GLI3 and neoplasms is not
well understood.
References A mouse model of Greig Cephalopolysyndactyly syndrome: the
extra-toes mutation contains an intragenic deletion of the GLI3 gene. Hui et al. Nature Genetics. 1993.
GLI3 is not mutated commonly in sporadic medulloblastomas. Erez et al. ACS. 2002.
Acute Lymphoblastic Leukemia in a Patient with Greig Cephalopolysyndactyly and Interstitial Deletion of Chromosome 7 del(7)(p11.2 p14) Involving the GLI3 and ZNFNIAI Genes. Mendoza et al. Genes, Chromosomes, & Cancer. 2005.
Robert B, Lallemand Y.Anteroposterior patterning in the limb and digit specification: contribution of mouse genetics. Dev Dyn. 2006 Sep;235(9):2337-52. Review.
Acknowledgement
Dr. Tim Rand Dr. Han Lee Dr. Scott Oakes
Dr. Philip Ursell
And..
All of the awesome path residents and fellows and attendings…
Extra Slides
.
What happens when you knockout Shh?
In the Shh null mutant, no digit is detectable in the forelimb and
a single digit
remains in the hindlimb.
This digit hasa digit1 (hallux)
identity according to molecular markers
Thus, despite the absence of Shh, the limb phenotype of the Shh:Gli3 double null mutants is indistinguishable from that of single Gli3 homozygotes.
This finding suggests that,in the limb, the main if not exclusive role of Shh is to prevent Gli3 processing
What happens if you have a Shh:Gli3 double knock-out?
Greig Syndrome and Acute Lymphoblastic Leukemia
Acute lymphoblastic leukemia in a patient with Greig cephalopolysyndactyly and interstitial deletion of
chromosome 7 del(7)(p11.2 p14) involving the GLI3 and ZNFNIAI Genes. Mendoza et al. Genes,
Chromosomes, & Cancer. 2005.
Case: 9 y/o boy with Greig syndrome with recurrent ALL.
Greig Syndrome and Acute Lymphoblastic Leukemia
Question: Is GLI3 related to ZNFNIAI?
ZNFNIAI a lymphoid-restricted zinc finger transcription factor which
has been associated with childhood leukemia
Decrease in ZNFNIAI activity leads in mice leads to T-cell hyperproliferation, aberrant expansion of thymic clones, and T-cell neoplasia
Mice that express non-DNA binding ZNFN1A1 isoforms develop an aggressive form of lymphoblastic leukemia
Results: Karyotype showed deletion of 7p11.2p14 in ~ 40% of
cells taken from a skin biopsy, or bone marrow aspirates FISH analysis shows the GLI3 gene was deleted in 74%
of cells
FISH (fluorescence in situ hybridization) of bone marrow cells using a GLI3 gene probe (red) showed normal hybridizationin 26% of the cells (b) and deletion in 74% of the cells (c).(Green labels the centromere of chromosome 7)
Present 26% Absent 74%
Patient with Greig syndrome and ALL has chromosome deletion that includes both GLI3 and ZNFNIAI
FISH (fluorescence in situ hybridization) of bone marrow cells using a ZNFNIAI gene probe (red) confirmed that the gene was within the deleted segment.
(Green labels the centromere of chromosome 7)
Patient with Greig syndrome and ALL has chromosome deletion that includes both GLI3 and ZNFNIAI
Conclusion: The deletion of a region in chromosome 7
resulted in Greig Syndrome (deletion of GLI3) and predisposed the patient to developing leukemia (deletion of ZNFN1A1).
Patient with Greig syndrome and ALL has chromosome deletion that includes both GLI3 and ZNFNIAI
The story of Sonic hedgehog
The zone of polarizing activity is a region of the limb bud mesenchyme and specifies the identity of each digit by threshold values of a morphogen
That morphogen is Sonic hedgehog Shh is a secreted protein Shh is expressed in the ZPA Cells expressing Shh can induce mirror-image
duplications of the digits Mice bearing a null mutation of Shh have forelimbs
that lack all digits in the forelimb and a single digit in the hindlimb
The story of Sonic Hedgehog
Mouse mutations leading to polydactyly are characterized by an ectopic Shh expression domain in the limb These mice include:
Hemimelic extra-toes, Sasquatch, Strong’s Luxoid (Alx4), extra-toes (Gli3)
The story of Sonic Hedgehog
SHH and GLI3 In Drosophila, hedgehog signaling is mediated
by the Cubitus interruptus (Ci) protein. Ci155 is a transcription activator Ci75 is a repressor
In birds and mammals, the orthologues of Cubitus interruptus are the three member Gli family (Gli1, Gli2, and Gli3)
In the limb, although the three Gli genes are expressed, only Gli3 appears to be necessary for patterning
The story of Sonic Hedgehog
SHH and GLI3 One form of GLI3 is a repressor of Shh Another form is possibly an activator Complicated interaction between GLI3 and
Shh, involving other proteins including Hand 2, Gremlin, Bmps, etc.