Transcript Example of a poster - University of Florida

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PCB 3063, University of Florida, Gainesville, FL
INTRODUCTION
Hutchinson-Gilford Progeria Syndrome, an
egregious laminopathy, develops in 1 in 4 million
infants between ages 12 and 24 months. A single
de novo autosomal dominant mutation in the LMNA
gene can lead to pleiotropic defects in array of
1C
different tissues, all stemming from nuclearmembrane buckling in somatic cells. Affected
children suffer accelerated aging, cardiovascular
defects, atherosclerosis, sclerotic skin, joint
contractures, osteodysplasia, osteolysis, pathologic
fractures, alopecia, growth retardation,
Figure 1. Phenotypic effects of
micrognathia, and a dearth of subcutaneous fat.
HGPS 12
A. Picture of HGPS child manifesting
Patients are not cognitively impaired and remain
classic signs of an undersize jaw,
mentally youthful as their bodies age. They most
alopecia, lack of subcutaneous fat,
often die from myocardial infarction or stroke
and accelerated age.
around thirteen years.
B. Picture of normal nuclear lamina.
1A
1B
C. Picture of “progerin-laden” nucleus
with characteristic abnormal
morphology.
LMNA GENE
GENE THERAPY
MODES OF INHERITANCE
Classical HGPS is primarily inherited in a de novo dominant autosomal manner with a
G608G mutation; despite this, affected individuals usually die before reaching
reproductive age. The de novo nature of this mutation is unsurprising given that the
cysteine of the CpG dinucleotide is easily methylated and deaminated to produce TpG.4
Atypical HGPS was observed in a study performed by Plasilova et al.11 on a
consanguineous Indian family that supported a recessive mode of inheritance. The
heterozygous offspring exhibited the affected phenotype (Figure 4).The investigators
used genome wide linkage analysis to restrict the gene locus to 1p13.3–1q23.3, and
additional microsatellite markers for further specificity. LMNA mutation analysis was used
to confirm a G to C transversion that resulted in a missense mutation; in this K542N, a
charged amino acid, lysine, was replaced by uncharged asparagine within the DNA
binding domain of the protein (Figure 5).
Figure 4. Pedigree of four generations of an
Indian family.11
Illustrates recessive inheritance. Consanguineous
parents, I-1 and I-2, did not exhibit the disorder, and
generated 5 heterozygous affected and 2 healthy
offspring.
The mapping of the LMNA gene, (Figure 2) was done by fluorescence in situ
hybridization with a DAPI counter-stain. Thus, specific assignment was made based
upon location or distance from DAPI bands and G-bands.14
MUTATIONS
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•Despite the synonymous nature of this substitution, the mutation results in an
unstable form of lamin A which generates a cryptic splice site that translates to an
in-frame deletion of 150 nucleotides or 50 amino acids.
•The LMNA gene actually codes for both lamin A and lamin C, but the mutation
only affects the structure of lamin A because exon 11 of LMNA is not present in
lamin C mRNA.2
Eriksson et al.4 was able to isolate the c.1824C>T mutation. Samples from 23
people diagnosed with classical HGPS underwent PCR amplification followed by
direct sequencing for all of the exons of the LMNA gene. The resulting sequences
revealed that a significant 18 people out of 23 people were found to be
heterozygous for the c.1824C>T mutation.
Figure 6. Contrasting posttranscriptional processing. The
mutant pre-lamin A has a 50AA deletion
and subsequently lacks the final
cleavage step that is characteristic to
normal mature lamin A production prior
to incorporation into the nuclear lamina.
http://www.ncbi.nlm.nih.gov/pmc/articl
es/PMC2846822/
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The most promising treatment aims to reverse the HGPS-cell phenotype via
farnesyl transferase inhibitors (FTIs)
FTIs bind to the CSIM sequence of pre-lamin A, blocking farnesyl transferase’s
target and preventing farnesyl addition.
Lacking the bulky substituent, progerin remains free from the INM.
FTIs are inevitably toxic as many proteins require farnesylation to maintain
proper functioning. Thus, FTIs are capable of disrupting normal cellular
processes.
CONCLUSION
Figure 5 . Significance of K542N
position. K542N mutation (yellow)
results in an AA substitution of an
uncharged residue in the place of a
charged residue within the DNA
binding domain, which is largely
positively charged (blue).
http://www.ncbi.nlm.nih.gov/pmc/a
rticles/PMC2846822/
•Classical HPGS is most commonly caused by a c.1824C>T mutation. This C>T
transition results in a silent Gly1824Gly mutation within the pre-lamin
polypeptide.
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The LMNA gene produces a polypeptide that requires post-translational processing to
produce the mature lamin A protein, which functions as a nuclear protein scaffold
significant to the integrity of the nuclear structure. In a study conducted by De SandreGiovannoli et al.2 on classical HGPS, a reverse transcriptase PCR isolated a normal
transcript and a truncated mRNA transcript from lymphocytes derived from an affected
child. The truncated transcript translated to a mutated form of pre-lamin A with an
internal deletion of 50 amino acids, including the Zmpste24 cleavage site, due to
activation of a cryptic splicing site. Thus, as seen in (Figure 6), the final cleavage of the
18 codons at the C terminus of the pre-lamin polypeptide cannot occur. Instead the
resultant protein, progerin remains farnesylated (with a hydrocarbon at CAAX motif)
and accumulates in the nuclear periphery.4 Progerin is then able intercalate into the
nuclear membrane and dimerize with normal lamin A to form a protein complex that
disrupts the intended protein scaffolding function; this results in the abnormal nuclear
morphology characteristic of HGPS.1
Figure 3 . Table of HPGS mutations. c.1824C>T
represents the most common classical mutation for
HGPS. Atypical mutations are significantly more rare,
but,are also presented. They can result in amino acid
substitutions as well as alternative splice cites.
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1735
754/pdf/v041p00e67.pdf
Because HGPS’s effect is not limited to specific tissue, but is instead universal to
somatic cells, there is no way to selectively repair the deleterious mutations
throughout the body.
I
PROTEIN
Figure 2. LMNA gene position. mapped the LMNA gene to 1q21.2–q21.3, with emphasis on
1q21.3 as indicated by the black triangle.
http://www.genecards.org/cgi-bin/carddisp.pl?gene=LMNA
Figure 7. Immunofluorescence staining
of fibroblasts.1 The three pictures depict a
normal skin cell (far left), an HGPS skin cell
(middle), and an FTI-treated HGPS skin cell
(far right). Notice the near-restoration of the
FTI-treated nuclear lamina’s spherical shape
from the collapsed “progerin-laden” HGPS
cell.
Although rare, HGPS remains a great concern for its array of debilitating effects. Even
more frustrating perhaps is its resistance to therapies. Despite knowing its exact
location, 1q21.2, its somatic virulence eludes direct combat, relegating most medical
interventions to high-calorie diets, careful playing with other children, and persistence.
The most promising treatment, FTIs, offer the hope of rescuing the nuclear lamina
from distortion in affected cells, but, again, at the risk of compromising other, healthy
protein and cellular processes in the body. Nevertheless, HGPS offers intriguing insight
into the massive epigenetic consequences of a single transition, swapping a cysteine
for a thymine.
REFERENCES
1. Capell, B.C., Erdos, M.R., Madigan, J.P., Fiordalisi, J.J., Varga, R., Conneely, K.N., Gordons, L.B., Der, C.J.,
Cox, A.D., Collins, F.S. (2005). Proc. Natl. Acad. Sci. USA. 102, 12879-12884.
2. Csoka, A.B., Cao, H., Sammak, P.J., Caonstantinescu, D., Schatten, G.P., Hegele, R.A. (2004). Novel
lamin A/C gene (LMNA) mutations in atypical progeroid syndromes. J. Med. Genet. 41, 304308.
3. De Sandre-Giovannoli, A., Bernard, R., Cau, P., Navarro, C., Amiel, J., Boccaccio, S.L., Stewart, C.,
Munnich, A., Le Merrer, M., Lévy, N. (2003). Lamin A Truncation in Hutchinson-Gilford
Progeria. Science. 300, 2055.
4. Eriksson, M., Brown, W.T., Gordon, L.B., Glynns, M.W., Singer, J., Scott, L., Erdos, M.R., Robbins, C.M.,
Moses, T.Y., Berglund, P., et al., (2003). Recurrent de novo point mutations in lamin A cause
Hutchinson-Gilford progeria syndrome. Nature. 423, 293-298.
5. Garg, A., Subramanyam, L., Agarwal, A.K., Simha, V., Levine, B., D’ Apice, M.R., Novelli, G., Crow, Y.
(2009). Atypical Progeroid Syndrome due to Heterozygous Missense LMNA Mutations. J. Clin.
Endocrinol. Metlab. 94, 4971-4983.
6. Gordon, L.B. (2011). Hutchinson-Gilford Progeria Syndrome.
http://www.ncbi.nlm.nih.gov/books/NBK1121/#hgps.Management
7. Gordon, L.B., McCarten, K.M., Giobboe-Hurder, A., Machan, J.T., Campbell, S.E., Berns, S.D., Kieran, M.W.
(2007). Disease Progression in Hutchinson-Gilford Progeria Syndrome: Impact on Growth
and Development. Pediatrics. 120, 824-833.
8. Glynn, M.W., Glover, T.W. (2005). Incomplete processing of mutant lamin A in Hutchinson-Gilford
progeria leads to nuclear abnormalities, which are reversed by farnesyltransferase inhibition.
Human Molecular Genetics. 14, 2959-2969.
9. Marji, J., O’Donoghue, S.I., McClintock, D., Satagopasm, V.P., Schneider, R., Ratner, D., Womran, H.J.,
Gordon, L.B., Djabali, K. Defective Lamin A-Rb Signling in Hutchinson-Gilford Progeria
Syndrome and Reversal by Farnesyltransferase Inhibition. PLoS Biol. 5, 1-14
10. McKusick, V.A. (2010). Hutchinson-Gilford Progeria Syndrome; HGPS.
http://www.ncbi.nlm.nih.gov/omim/176670#BiochemicalFeatures-176670.
11. Plasilova, M., Chattopadhyay, C., Pal, P., Schaub, N.A., Buechner, S.A., Mueller, H., Miny, P., Ghosh, A.,
Heinimann, K. (2004). Homozygous missense mutation in the lamin A/C gene causes
autosomal recessive Hutchinson-Gilford progeria syndrome. 41, 609-614.
12. Raska, I. (2010). Importance of molecular cell biology investigations in human medicine in the story of
the Hutchinson-Gilford progeria syndrome. Interdisc Toxicol. 3, 89-93.
Scaffidi, P., Gordon, L., Misteli, T. (2005). The Cell Nucleus and Aging: Tantalizing Clues and Hopeful
Promises. PLoS Biol. 3, 1855-1859.
13. Wydner, K.L., McNeil, J.A., Lin, F., Worman, H.J., Lawrence, J.B. (1996). Chromosomal Assignment of
Human Nuclear Envelope Protein Genes LMNA, LMNB1, and LBR by
FluorescenceinsityHybridization. Genomics. 32, 474-478.
14. Yang, S.H., Chang, A.Y., Ren, S., Wang, Y., Andres, D.A., Spielmann, H.P., Fong, L.G., Young, S.G.
(2011). Absence of progeria-like disease phenotypes in knock-in mice expressing a nonfanesylated version of progerin. Human Molecular Genetics. 20, 436-444.
lts in an unstable form of Lamin A which generates a cryptic site within the precursor mRNA for lamin A. The LMNA gene codes for both lamin A and lamin C, but the mutation only affects the structure of lamin A because exon 11 of LMNA is not present in lamin C mRNA m (Csoka, J Me
e through a mutation on chromosome 1q at position 1822 of a Guanine to Adenine, a mutation on chromosome 1q at position 1921 of a Guanine to Adenine, a mild mutation: 35 amino acid deletion, a mutation at E145K, and a joint mutation of R471C and R527C (Csoka, J Med Genet 2
and severe disease phenotypes in affected patients. Retention of farnesyl group causes progerin to become permanently anchored in the nuclear membrane and unable to be released. The central rod domain of progerin then allows dimerization with mature nonfarnesylated LA and asse
The LMNA gene was mapped to 1q21.2–q21.3, with positional emphasis on
1q21.3, as determined by fluorescence in situ hybridization with a DAPI
counter-stain.
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1. De Sandre-Giovannoli, A., Bernard, R., Cau, P., Navarro, C., Amiel, J., Boccaccio,
S.L., Stewart, C., Munnich, A., Le Merrer, M., Lévy, N. (2003). Lamin A Truncation
in Hutchinson-Gilford Progeria. Science. 300, 2055.
2. Domingo, D.L., Trujillo, M.I., Council, S.E., Merideth, M.A., Gordon, L.B., Wu, T.,
Introne, W.J., Gahl, W.A., Hart, T.C. (2009). Hutchinson-Gilford progeria
syndrome: Oral and craniofacial phenotypes. Oral Dis. 15, 187-195.
3. Eriksson, M., Brown, W.T., Gordon, L.B., Glynns, M.W., Singer, J., Scott, L.,
Erdos, M.R., Robbins, C.M., Moses, T.Y., Berglund, P., et al., (2003). Recurrent de
novo point mutations in lamin A cause Hutchinson-Gilford progeria syndrome.
Nature. 423, 293-298.
4. Garg, A., Subramanyam, L., Agarwal, A.K., Simha, V., Levine, B., D’ Apice, M.R.,
Novelli, G., Crow, Y. (2009). Atypical Progeroid Syndrome due to Heterozygous
Missense LMNA Mutations. J. Clin. Endocrinol. Metlab. 94, 4971-4983.
5. Gordon, L.B. (2011). Hutchinson-Gilford Progeria Syndrome.
http://www.ncbi.nlm.nih.gov/books/NBK1121/#hgps.Management
6. Gordon, L.B., McCarten, K.M., Giobboe-Hurder, A., Machan, J.T., Campbell, S.E.,
Berns, S.D., Kieran, M.W. (2007). Disease Progression in Hutchinson-Gilford
Progeria Syndrome: Impact on Growth and Development. Pediatrics. 120, 824833.
7. Glynn, M.W., Glover, T.W. (2005). Incomplete processing of mutant lamin A in
Hutchinson-Gilford progeria leads to nuclear abnormalities, which are reversed by
farnesyltransferase inhibition. Human Molecular Genetics. 14, 2959-2969.
8.Marji, J., O’Donoghue, S.I., McClintock, D., Satagopasm, V.P., Schneider, R.,
Ratner, D., Womran, H.J., Gordon, L.B., Djabali, K. Defective Lamin A-Rb Signling
in Hutchinson-Gilford Progeria Syndrome and Reversal by Farnesyltransferase
Inhibition. PLoS Biol. 5, 1-14
9. McKusick, V.A. (2010). Hutchinson-Gilford Progeria Syndrome; HGPS.
http://www.ncbi.nlm.nih.gov/omim/176670#BiochemicalFeatures-176670.
10. Plasilova, M., Chattopadhyay, C., Pal, P., Schaub, N.A., Buechner, S.A., Mueller,
H., Miny, P., Ghosh, A., Heinimann, K. (2004). Homozygous missense mutation in
the lamin A/C gene causes autosomal recessive Hutchinson-Gilford progeria
syndrome. 41, 609-614.
11. Raska, I. (2010). Importance of molecular cell biology investigations in
human medicine in the story of the Hutchinson-Gilford progeria syndrome.
The majority of patients with HGPS
have de novo heterozygous dominant
mutations in the LMNA gene.
Presumably, patients with the disorder
do not survive long enough to
reproduce (Eriksson et al., 2003; Cao
and Hegele, 2003).