Nationwide Children s Hospital August 25, 2015 Nuclear Envelope and Muscular Dystrophy Howard J. Worman, M.D. Columbia University Columbia University Medical Center
The Nuclear Envelope By D. W. Fawcett
The Nuclear Envelope: Nuclear Lamina Dauer & Worman, 2009
Nuclear Lamins: A Brief Ancient History Dwyer& Blobel (1975) Aebi et al. (1986) Gerace, Blum & Blobel (1978) Aebi et al. (1986) Goldman et al. (1986) McKeon et al. (1986) Fisher et al. (1986)
HUMAN NUCLEAR LAMINS LOCUS CHROMOSOME PROTEINS CELL-TYPES EXPRESSED LMNA 1q21.2-21.3 Lamin A Differentiated Somatic Lamin C Lamin A D10 Lamin C2 Differentiated Somatic Differentiated Somatic Germ LMNB1 5q23.2-31.1 Lamin B1 Apparently All Somatic LMNB2 19p13.3 Lamin B2 All or Most Somatic Lamin B3 Germ
LMNA Encoding A-type Lamins Lin and Worman, JBC, 1993 Localization to chromosome 1q21.2 q21.3 Wynder et al., Genomics, 1995
Mutations in LMNA Cause Diseases ( Laminopathies ) with Four Major Tissue-Selective Phenotypes Dauer and Worman Dev. Cell (2009)
LMNA Genotype- Phenotype Correlations Dominant mutations causing muscle diseases are missense, splicing and small deletions and even haploinsufficiency Recessive missense mutation peripheral neuropathy Dominant mutations cluster in a small region encoded of exon 8 in partial lipodystrophy Recessive missense in MAD Dominant G608G or S608G in HGPF Worman and Courvalin (2005)
Hutchinson-Gilford Progeria Syndrome (HGPS) Normally, lamin A starts as a precursor prelamin A, which is fanesylated. The farnesylated precursor is recognized by ZMPSTE24 endoprotease and cleaved; mature lamin A is not farnesylated. From Worman and Courvalin. (2005) farnesyl The LMNA mutation causing HGPS leads to expression of a truncated prelamin A ( progerin ), which remains farnesylated. Paradisi et al. (2005) Modified from Eriksson et al. Nature (2003)
Evidence that Abnormal Farnesylation of Progerin is Involved in Pathogenesis of HGPS Treatment of cultured fibroblasts from model mice and affected human with protein prenylation inhibitors reverse abnormal nuclear shape characteristic of cells expressing progerin Treatment of mice with a targeted HGPS mutation with farnesyltransferase inhibitors improve phenotype Based on above, clinical trials in affected children have been conducted but unfortunately uncontrolled ones There are some LMNA mutations that cause progeroid phenotypes that apparently do not alter prenylation
LMNA Mutations Causing Striated Muscle Disease Versus LMNA Mutations Causing Lipodystrophy Dunnigan-type Familial Partial Lipodystrphy Emery-Dreifuss Muscular Dystrophy and Variant Disorders
The vast majority of LMNA mutations in Dunnigan-type partial lipodsytrophy change the surface charge of an Ig fold in the lamin A/C tail domain Cter Nter Nter Cter W520 R527 Y481 R453 R482 V442, V452, I469, I497, T528, L530 N456 G465 K486 Krimm et al. Stucture (2001)
Recessive LMNA Mutations Charcot-Marie-Tooth type 2B1 Mandibuloacryl dysplasia Tazir et al. (2004) Novelli et al. (2002) LMNA R298C LMNA R2527H
Integral Inner Nuclear Membrane Protein Gene Mutations EDM emerin Emery-Dreifuss Muscular Dystrophy (X-linked) and variants LBR lamin B receptor Pelger-Huët Anomaly (Heterozygous) HEM/Greenberg Skeletal Dysplasia (Homozygous) MAN1/LEMD3 Osteopoikilosis, Buschke-Ollendorff Syndrome
Outer Nuclear Membrane Protein/Perinuclear Space Gene Mutations SYNE1 nesprin-1 Autosomal Recessive Cerebellar Ataxia Arthrogryposis Myopathies SYNE2 nesprin-2 Myopathies SYNE4 nesprin-4 High Frequency Hearing Loss TOR1A torsina DYT1 Dystonia Gundersen and Worman Cell (2013)
LMNA Cardiomypathy and Muscular Dystrophy
LMNA Linked Myopathies Emery-Dreifuss Limb-girdle Type 1B Dilated Cardiomyopathy 1A All three clinical phenotypes can occur in members of the same family with the same mutation and there is overlap between phenotypes: LMNA dilated cardiomyopathy with variable skeletal muscle involvement.
LMNA Cardiomyopathy Approximately 6% to 9% of all dilated cardiomyopathies Up to 30% of dilated cardiomyopathies with AV block and skeletal muscle involvement van Berlo et al. J. Mol. Med. 2004; 83:79-83 Taylor et al. J. Am. Coll. Cardiol. 2003;41: 771-780
LMNA H222P/H222P Mouse Model of Emery-Dreifuss Muscular Dystrophy (Arimura et al. 2005) Heart disease progression from 8 to 20 weeks of age in male mice Heart structure and function Significant LV dilatation and decreased fractional shortening by echocardiography
Abnormal Signaling in Hearts of Lmna H222P/H222 Mice HEART at 10 weeks AFFYMETRIX 430 2.0 MAP Kinase AKT/mTOR
Early Activation of ERK1/2 in Hearts of Lmna H222P/H222P Mice > ERK1/2 and downstream genes are abnormally activated prior to (4 weeks and 7 weeks) and concurrent with (10 weeks) the onset of cardiomyopathy in hearts of Lmna H222P/H222P mice 4 weeks Choi et al. Sci Transl Med 2012 Muchir et al. J Clin Invest 2007
MEK1/2 Inhibitor Protocol D r u g A d m i n i s t r a t i o n MEK1/2 ERK ERK P P PD98059 Selumetinib CIP-137401 1 6 w e e k s 2 0 w e e k s E n d p o i n t s -E c h o c a r d i o g r a p h y -B i o c h e m i c a l M a r k e r s -C a r d i a c F i b r o s i s
M E K 1 / 2 I n h i b i t o r T r e a t m e n t Target Inhibition decreased phosphorylated ERK1/2 (perk1/2) in heart after selumetinib treatment
M E K 1 / 2 I n h i b i t o r T r e a t m e n t Muchir et al. 2012
M E K 1 / 2 I n h i b i t o r T r e a t m e n t Fibrosis decreased fibrosis tissue and collagen gene expression 20% 12%
M E K 1 / 2 I N H I B I T I O N Survival prolonged median survival (proprietary MEK1/2 inhibitor)
MEK1/2 Inhibitor Treatment Blocks AKT/mTOR in Hearts of Lmna H222P/H222P Mice Chhoi et al. Sci Transl Med (2012)
Increased AKT/mTOR Activity Correlates with Decreased Autophagy Lmna Mice Human Subjects Chhoi et al. Sci Transl Med (2012)
Temsirolimus Blocks mtor Activity Improving Heart Function and Enhancing Autophagy Chhoi et al. Sci Transl Med (2012)
One Model of LMNA Cardiomyopathy Normal Heart Outstanding Practical Question: Can use of these drugs be optimized to treat human subjects? CIP-137401 Outstanding Theoretical Question: How do LMNA mutation activate these pathways? Dilated Heart
Emerin and LAP1 and Muscular Dystrophy
Mutations in EMD Encoding Emerin Phenocopy Myopathies Caused by LMNA Mutations X-linked Emery-Dreifuss Muscular Dystrophy (and related myopathies) Emerin binds to A-type lamins In most cases, emerin is lacking from the nuclear envelope Muscle Frozen Sections Anti-emerin Ab Normal X-EDMD X-EDMD Modified from A.E.H. Emery Nagano et al. Nat Genet. 1996;12:254-259
Emerin as a Candidate LAP1 Binding Protein Lamina-associated polypeptide 1 (LAP1) integral inner nuclear membrane protein of unclear function In an unbiased proteomics screen for interacting proteins in collaboration with Bill Dauer, emerin was found to be top candidate
Emerin Binds to LAP1 Biochemical Assays FRET Shin et al. Dev Cell (2013)
Loss of Emerin: of Mice and Men Pacemaker Emery-Dreifuss Muscular Dystrophy A.E. H. Emery (2000) Apparently Normal Mouse Melcon et al. Hum Mol Genet 2006;15:637-651 Ozawa et al. Am J Pathol 2006;168:907-917
Why Don t Emerin Null Mice Get Sick? Mouse skeletal muscle has diminished emerin compared to human. Conversely, LAP1 is significantly higher in mouse than human striated muscle. Shin et al. Dev Cell (2013)
Therefore, We Depleted LAP1 from Mouse Skeletal Muscle Shin et al. Dev Cell (2013)
Mice Lacking LAP1 in Skeletal Muscle Shin et al. Dev Cell (2013)
LAP1 Depletion Causes Muscular Dystrophy Shin et al. Dev Cell (2013)
Depletion of LAP1 Striated Muscle on Emerin Null Background Shin et al. Dev Cell (2013)
Control: Deplete LAP1 from Hepatocytes Shin et al. Dev Cell (2013)
Deplete LAP1 from Hepatocytes Minimal Pathology Shin et al. Dev Cell (2013)
Depletion of LAP1 from Heart Shin et al. Nucleus (2014)
Depletion of LAP1 from Heart Decreases Left Ventricular Fractional Shortening Shin et al. Nucleus (2014)
Disruption of Emerin-Lamin A/C-LAP1 Complex: Cardiomyoapthy and Muscular Dystrophy Disruption Emerin-Lamin A/C-LAP1 Complex ERK1/2 and AKT/mTOR Striated Muscle Disease
Columbia Past & Present Revekka Boguslavsky Jason Choi Gregg Gundersen Alan Herron Stephen Lehnart Caroline LeDour Feng Lin Angelika Lüdtke Ivan Méndez-López Antoine Muchir Cecilia Östlund Paul Pavlides Jian Shan Ji-Yeon Shin Bruno Soullam Yuexia Wang Wei Wu Quan Ye Collaborators Elsewhere Takuro Arimura Gisèle Bonne Jean-Claude Courvalin Bill Dauer Yukiko Hayashi Micheline Paulin-Levasseur Sophie Zinn-Justin
Funding BioAccelerate NYC Prize Los Angeles Thoracic and Cardiovascular Foundation