Gene imprinting(edited)

VBMS 7720 Developmental Molecular BiologyPresented by: Chris Ramhold Ph.D. Student

2/25/15

GENE IMPRINTING

• What is imprinting?– Expand your vocabulary

• Inheritance– Mendelian/Non-Mendelian– Pattern of Imprinting

• Importance• Mechanisms

– Big Picture• Disease

– Detection of imprinted genes• Disclaimer *Your results may vary*

OVERVIEW

• Epigenetics- “On genes,” modifications to DNA that retain sequence fidelity yet alter gene expression

• Imprinting- Gamete-specific differential modification– Unique to placental mammals, marsupials and flowering

plants– Typically marked with DNA methylation– Often occur in clusters– Gene is silenced

• Epigenetic memory- The set of epigenetic modifications inherited in descendant cells

WHAT IS IMPRINTING?

• 2 broad categories of inheritance– Mendelian– Non-mendelian

• Law of Segregation- Each parent contributes a single gamete containing single alleles from either maternal or parental chromosomes

MENDELIAN GENETICS…. INHERITANCE

IMPRINTING INHERITANCE PATTERN (GAMETES)

http://hihg.med.miami.edu/code/http/modules/education/Design/Print.asp?CourseNum=2&LessonNum=2

• Imprinting established in gametes

IMPRINTING INHERITANCE PATTERN (POST FERTILIZATION)


• Imprinting maintained during embryogenesis

• Genome-wide epigenetic reprogramming

• Paternal X inactivation

IMPRINTING INHERITANCE PATTERN (DIFFERENTIATION)


• Imprinting maintained in adult somatic tissues

IMPRINTING INHERITANCE PATTERN ( BACK TO GAMETES)


• Imprinting erased and modified based on sex

• Imprinted genes– Are crucial for normal development– Bypass epigenetic reprogramming– Are vulnerable to epigenetic copying machinery

• Roles in– Growth– Behavior– Stem cells– Disease

IMPORTANCE

• Paternal expression– Placental development– Enhance growth– Large offspring (benefit for father)

• Maternal expression– Suppress growth– Limit expression of paternal genes– Small offspring (benefit for mother)• Resource use

IMPORTANCE

Moon, Y. S., Smas, C. M., Lee, K., Villena, J. A., Kim, K.-H., Yun, E. J., & Sul, H. S. (2002). Mice lacking paternally expressed Pref-1/Dlk1 display growth retardation and accelerated adiposity. Molecular and Cellular Biology, 22, 5585–5592. doi:10.1128/MCB.22.15.5585-5592.2002

• Beautiful buttocks• Callipyge gene• Paternally expressed

IMPORTANCE

MECHANISM- HOW DOES IT WORK?

Zakhari, S. (2013). Alcohol metabolism and epigenetics changes. Alcohol Research : Current Reviews, 35, 6–16.

• DNA Methylation– CpG- Cytosine-

phosphate-guanine

– DNMT- DNA methyltransferase

– 5’Methyl-cytosine can easily be deaminated to form thymine

• CpG Islands- Regions with high concentrations of Cpg motifs– Promoter region– ~70% of human promoter regions

contain CpG islands• Imprinting Control Regions (ICR) &

Differentially Methylated Regions (DMR)– Cis-acting– Control gene clusters ~3.0 Mb away– Deletion of ICR leads to loss of

imprinting

DNA METHYLATION

http://biol10005.tumblr.com/post/31972466118/epigenetics

DNA METHYLATION

Li, E. (2002). Chromatin modification and epigenetic reprogramming in mammalian development. Nature Reviews. Genetics, 3, 662–673. doi:10.1038/nrg887

• Dnmt3 family facilitates imprinting in gametes

• Dnmt1 maintains methylation states post-embryogenesis

• 98% of genome 1 CpG/100bp- Me

• <2% of genome 1 CpG/10bp- Un-Me

• Chromatin condensation• Gene imprinting on rare

occasions will modify histones through acetylation/methylation

DNA METHYLATION- QUICK STEP BACK

Duygu, B., Poels, E. M., & da Costa Martins, P. A. (2013). Genetics and epigenetics of arrhythmia and heart failure. Frontiers in Genetics, 4, 219. doi:10.3389/fgene.2013.00219

• Acetylation generally leads to increased expression

• Methylation generally leads to decreased expression

• Ideas on why imprinting occurs in clusters?

HISTONE MODIFICATION

Duygu, B., Poels, E. M., & da Costa Martins, P. A. (2013). Genetics and epigenetics of arrhythmia and heart failure. Frontiers in Genetics, 4, 219. doi:10.3389/fgene.2013.00219

• Imprinted genes contain regions of ncRNAs

• Air paternally expressed

• Repress flanking protein-coding genes in cis

LNC-RNA

Fatica, A., & Bozzoni, I. (2014). Long non-coding RNAs: new players in cell differentiation and development. Nature Reviews. Genetics, 15, 7–21. doi:10.1038/nrg3606

• CTCF- 11-zinc finger protein

• Binds Imprinting Control Element

• Prevents methylation of ICE and H19

• Ins2- Insulin 2• H19- lnc-RNA,

negative regulator of growth and proliferation

INSULATOR MODEL

• “Random” X-chromosome inactivation– Cell-to-cell basis– Actually regulated by several cis-Elements (Choice elements)

• Xist- X-Inactive Specific Transcript- encodes lnc-RNA• Tsix- Antisense to Xist- negative regulator of Xist

X CHROMOSOME INACTIVATION CO-EVOLUTION?

Reik, W. & Lewis, A. Co-evolution of X-chromosome inactivation and imprinting in mammals. Nature Rev. Genet. 6, 403–410 (2005).

• Xist


• Xist lncRNA acts cis- coating chromosome• Histone modification silences coated chromosome• Very similar to imprinting



• DNA methylation steps identical to imprinting


• Xist


• Methylation is maintained in placenta• Lost in embryonic tissue


• Paternal genome undergoes rapid de-methylation

• Maternal genome de-methylates more slowly

• Imprinted genes escape methylome reprogramming

UNDERSTANDING THE TIMELINE OF METHYLATION

Vertino, P. (2011), "DNA methylation in cancer", in Issa, J. (ed.), DNA Methylation: Physiology, pathology and disease, The Biomedical & Life Sciences Collection, Henry Stewart Talks Ltd, London

THE METHYLOME AND YOU (BIG PICTURE)

Vertino, P. (2011), "DNA methylation in cancer", in Issa, J. (ed.), DNA Methylation: Physiology, pathology and disease, The Biomedical & Life Sciences Collection, Henry Stewart Talks Ltd, London

THE METHYLOME AND YOU (ADULT LIFE)

THE METHYLOME AND YOU (DISEASE)

• Angelman Syndrome (AS)– 1/20k births– Severe mental retardation– Lack of speech– Ataxic gait– Unnaturally happy disposition• Random bouts of laughter

– Hand flapping

DISEASE

• Prader-Willi Syndrome (PWS)– 1/25k births– Mild mental retardation– Chronic hunger- obesity– Stunted height– Most common syndromal cause of

human obesity

DISEASE

• Loss of function• Chromosome 15

– Loss of expression in 15q11-13

• AS- Maternal• PWS- Paternal

DISEASE

• Beckwith-Wiedemann Syndrome– Microcephaly- small head– Macroglossia- enlarged tongue– Visceromegaly- enlarged organs– Macrosomia- large body size– Umbilical hernia

• Loss of imprinting on chromosome region 11p15.5

DISEASE

http://health.shorehealth.org/imagepages/17076.htm

• Cancer– Wilms’ tumor• Embryonic kidney cancer

– Loss of imprinting control on H19– Hypermethylation of imprinting control

region– Overexpression of IGF2

DISEASE

Martens-Uzunova ES, Bottcher R, Croce CM, Jenster G, Visakorpi T, Calin GA. Long noncoding RNA in prostate, bladder, and kidney cancer. Eur Urol. 2013;65:1140–51. doi: 10.1016/j.eururo.2013.12.003.

DISEASE

• Cancer– Bladder cancer

– Loss of imprinting on H19– Hypomethylation of imprinting control

region– Overexpression of IGF2

Martens-Uzunova ES, Bottcher R, Croce CM, Jenster G, Visakorpi T, Calin GA. Long noncoding RNA in prostate, bladder, and kidney cancer. Eur Urol. 2013;65:1140–51. doi: 10.1016/j.eururo.2013.12.003.

• Bisulfite sequencing– Cytosine converted to

uracil– Methylated cytosine

unaffected

DETECTION OF IMPRINTED GENES

• Sequencing after treatment should reveal no bands in lane C

• Bands in lane C represent methylated cytosines

DETECTION OF IMPRINTED GENES

Herman, J G et al. “Methylation-Specific PCR: A Novel PCR Assay for Methylation Status of CpG Islands.” Proc of Nat. Ac. of Sci. 93.18 (1996): 9821–9826.

• Hongerwinter 1944– Dutch period of starvation during WWII– Childen born were short, and

diagnosed with anemia, edema, diabetes, and depression

– The women who were born of this era were shown to have children that mimicked the same symptoms as their mothers.

– Hyopmethylation of IGF2 six decades later

– Implication in environmental imprinting

YOU ARE WHAT YOU DON’T EAT?

Veenendaal M, Painter R, de Rooij S, Bossuyt P, van der Post J, Gluckman P, et al. Transgenerational effects of prenatal exposure to the 1944-45 Dutch famine. BJOG (2013) 120(5):548–5410.1111/1471-0528.12136

• Duygu, B., Poels, E. M., & da Costa Martins, P. A. (2013). Genetics and epigenetics of arrhythmia and heart failure. Frontiers in Genetics, 4, 219. doi:10.3389/fgene.2013.00219

• Fatica, A., & Bozzoni, I. (2014). Long non-coding RNAs: new players in cell differentiation and development. Nature Reviews. Genetics, 15, 7–21. doi:10.1038/nrg3606

• Herman, J G et al. “Methylation-Specific PCR: A Novel PCR Assay for Methylation Status of CpG Islands.” Proceedings of the National Academy of Sciences of the United States of America 93.18 (1996): 9821–9826.

• http://health.shorehealth.org/imagepages/17076.htm• http://biol10005.tumblr.com/post/31972466118/epigenetics• Li, E. (2002). Chromatin modification and epigenetic reprogramming in mammalian development. Nature Reviews. Genetics, 3,

662–673. doi:10.1038/nrg887• Martens-Uzunova ES, Bottcher R, Croce CM, Jenster G, Visakorpi T, Calin GA. Long noncoding RNA in prostate, bladder, and kidney

cancer. Eur Urol. 2013;65:1140–51. doi: 10.1016/j.eururo.2013.12.003.• Moon, Y. S., Smas, C. M., Lee, K., Villena, J. A., Kim, K.-H., Yun, E. J., & Sul, H. S. (2002). Mice lacking paternally expressed

Pref-1/Dlk1 display growth retardation and accelerated adiposity. Molecular and Cellular Biology, 22, 5585–5592. doi:10.1128/MCB.22.15.5585-5592.2002

• Reik, W. & Lewis, A. Co-evolution of X-chromosome inactivation and imprinting in mammals. Nature Rev. Genet. 6, 403–410 (2005).• Veenendaal M, Painter R, de Rooij S, Bossuyt P, van der Post J, Gluckman P, et al. Transgenerational effects of prenatal exposure to

the 1944-45 Dutch famine. BJOG (2013) 120(5):548–5410.1111/1471-0528.12136• Vertino, P. (2011), "DNA methylation in cancer", in Issa, J. (ed.), DNA Methylation: Physiology, pathology and disease, The

Biomedical & Life Sciences Collection, Henry Stewart Talks Ltd, London• Zakhari, S. (2013). Alcohol metabolism and epigenetics changes. Alcohol Research : Current Reviews, 35, 6–16.

REFERENCES

Gene imprinting(edited)

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