1.1. The nuclear envelope
The nucleus is the defining characteristic organelle of the eukaryotes, and contains the nuclear genome. It is segregated from the cellular cytoplasm by the bilayer nuclear envelope (Figure 1), which consists of concentric inner and outer nuclear membranes, between which lies the perinuclear space. The outer nuclear membrane is contiguous with the rough endoplasmic reticulum, like which it is studded with protein producing ribosomes, and the perinuclear space is contiguous with the lumen of the endoplasmic reticulum. Transport across the nuclear envelope is accommodated by nuclear pore complexes (NPCs). The NPCs are the site where the inner and outer nuclear membranes are connected, as their shared lipid bilayers are united at that point. These NPCs are large, complex and heterogeneous protein structures, made up of multiple copies of approximately 30 different proteins, called nucleoporins . NPCs span the inner and outer nuclear membranes, and allow the regulated relocation of molecules between the nucleoplasm and cytoplasm. While smaller molecules, such as small metabolites or proteins under 40 kDa, are passively transported through the NPCs, larger molecules such as mRNAs, tRNAs, ribosomes and signalling molecules can be actively transported from the nucleus, while signalling molecules, proteins, lipids and carbohydrates are actively transported both into and out of the nucleus [2,3].
The inner nuclear membrane is embedded by various inner nuclear membrane proteins, such as LAP1, LAP2 and MAN1, which are involved in cell cycle control, linking the nucleus to the cytoskeleton and chromatin organisation [4,5]. Underlying and connected by various nuclear envelope proteins to the inner nuclear membrane are the nuclear lamina, a thin (30-100nm) and densely woven fibrillar mesh of intermediate filaments, composed of evolutionarily conserved lamins A, B1, B2 and C, and lamin associated proteins. These proteins are closely associated with the NPCs (Figure 1). This assembly of outer nuclear membrane, inner nuclear membrane, NPCs, and the lamina can be thought of as complex interface, coupling the nuclear genome to the rest of the cell, allowing for a sophisticated means of regulated traffic between inner and outer nuclear space, while compartmentalising DNA replication, RNA transcription and mRNA editing from translation at the ribosomes .
The nuclear lamins are type V intermediate filaments (IFs), and are closely related to the cytoplasmic intermediate filaments (types I-IV, which include the keratins), differing by the presence of a nuclear localisation signal (NLS) located in the initial section of the tail domain . Physically, these lamins have the characteristic tripartite assemblage of intermediate filaments; a short globular N-terminal head domain and a long C-terminal tail domain containing an immunoglobin-like domain, separated by a conserved central alpha-helical rod domain (Figure 4). Coiled-coil homodimers of A- and B-type lamins are formed by interaction between adjacent heptad hydrophobic repeats on the central rod domain, and charged residues along the centre of this dimer promote further assembly between dimers, leading to assembly of filamentous fibrils, whereas the N and C terminal endings facilitate head-to-tail polymerisation [6-8]. The nuclear lamina has been shown to have a major role in nuclear structure, heterochromatin organisation and gene regulation [8-11].
1.2. The lamins
A-type lamins are expressed only in differentiated cells, suggesting that they have a role in stabilising differential gene expression [15,16,19,20]. The main products in somatic cells are lamins A and C, with C2 and AΔ10 being less common isoforms, lamin C2 being specific to the testes [6,13,16,21]. The first 566 amino acids of lamins A and C are identical. However, at the C-terminals lamin A has 98 unique amino acids, and as with lamin B1 and B2, ends in a CaaX box motif, whilst lamin C has 6 unique terminal amino acids.
The second family of lamins, the B-type lamins, consist of lamin B1 encoded by the
The maturation process for lamin A, lamin B1 and B2 is detailed below, with these post-translational modifications taking place in the nucleus .
Prenylation: A farnesyl or geranylgeranyl isoprenoid group is covalently attached to the cysteine of the CaaX motif of prelamin A, lamin B1 and B2 by farnesyltransferase or geranylgeranyltransferase-I, respectively.
Cleavage: The terminal -aaX amino acids are removed by RCE1 and FACE1 for prelamin A, and by RCE1 alone for lamin B1 and B2.
Methylation: The now exposed C-terminal farnesylcysteine undergoes a methylation step, performed by a carboxymethyltransferase, isoprenylcysteine carboxyl methyltransferase (ICMT) . This is the final post-translational step for B-type lamins, therefore they retain the farnesylcysteine α-methyl ester at the C-terminus.
Second cleavage (for prelamin A only): FACE1 cleaves the carboxy-terminal 15 amino acids, including the farnesylcysteine methyl ester group, at the NM . This final modification step completes the post-translational modification of prelamin A to mature lamin A. This maturation is thought to aid localisation of lamin A to the nuclear rim [30,31].
Diseases caused by mutations in the
Laminopathies are caused by a heterogeneous set of pleiotropic mutations affecting universally expressed genes. However, their effects can be tissue specific to a degree, allowing for categorisation into five groups (Table 1). Striated muscles are affected in muscular dystrophies, peripheral nerves are affected in neuropathies, adipose tissue in lipodystophies, several tissues affected with premature development of multiple markers of senescence in segmental progeriod diseases, and finally diseases displaying symptoms from more than one category are known as overlapping syndromes.
2.1. Muscular dystrophies
Within this following section, selected muscular dystrophies will be detailed, while Table 2 shows a complete listing of known muscular dystrophy laminopathies, at the time of writing.
2.1.1. Emery-dreifuss muscular dystrophy
Emery-Dreifuss muscular dystrophy (EDMD), first described in 1955 , is the most prevalent laminopathy, affecting 1 in 100,000 births. It is also a prototypical laminopathy, occurring both as a primary and secondary laminopathy. The most commonly occurring form is autosomal dominant (AD-EDMD). It also occurs as an autosomal recessive (AR-EDMD) or X-linked (XL-EDMD) form [38,39]. Mutations in the emerin gene are responsible for XL-EDMD [40-43], while mutations in the
|Cardiomyopathy, dilated, 1A||CMD1A||115200||1q22|
|Emery-Dreifuss muscular dystrophy 1, X-linked||EDMD1||310300||Xq28|
|Emery-Dreifuss muscular dystrophy 2, AD||EDMD2||181350||1q22|
|Emery-Dreifuss muscular dystrophy 3, AR||EDMD2||181350||1q22|
|Emery-Dreifuss muscular dystrophy 4, AD||EDMD4||612998||6q25.1-q25.2|
|Emery-Dreifuss muscular dystrophy 5, AD||EDMD5||612999||14q23.2|
|Emery-Dreifuss muscular dystrophy 6, X-linked||EDMD6||300696||Xq26.3|
|Heart-hand syndrome, Slovenian type||HHS-S||610140||1q22|
|Muscular dystrophy, congenital||MDC||613205||1q22|
|Muscular dystrophy, limb-girdle, type 1B||LGMD1B||159001||1q22|
|Acquired partial lipodystrophy||APLD||608709||19p13.3|
|Lipodystrophy, familial partial, 2||FPLD2||151660||1q22|
|Mandibuloacral dysplasia with type A lipodystrophy||MADA||248370||1q22|
|Mandibuloacral dysplasia with type B lipodystrophy||MADB||608612||1p34.2|
|Adult-onset autosomal dominant leukodystrophy||ADLD||169500||5q23.2|
|Charcot-Marie-Tooth disease, type 2B1||CMT2B1||605588||1q22|
|Hutchinson-Gilford progeria syndrome||HGPS||176670||1q22|
|Atypical Werner syndrome||AWRN||277700||8p12|
|Hydrops-Ectopic calcification-moth-eaten skeletal dysplasia||HEM||215140||1q42.12|
|Melorheostosis with osteopoikilosis||MEL||155950||12q14.3|
EDMD is characterised by an onset in the teenage years of a slow, progressive wasting of skeletal muscle tissue in the shoulder girdle and distal leg muscles. This atrophy leads to muscle weakness around the humerus and fibula (a pattern described as scapulo-humero-peroneal), early contractures of the
2.1.2. Limb-girdle muscular dystrophy, type 1B
Limb-girdle muscular dystrophy, type 1B (LGMD1B) is a slowly progressive variant caused by an autosomal dominant mutation of the
Patients display a classic limb-girdle pattern of muscle atrophy, with a proximal lower limb muscular weakness starting by age 20. By the 30s and 40s upper limb muscles also gradual weakened . As in EDMD, serum creatine kinase levels were normal or elevated. The late occurrence or absence of spinal, elbow and Achilles contractures distinguishes LGMD1B from EDMD. Cardiac conduction abnormalities with dilated cardiomyopathy also occur. One neonatally lethal case of LGMD1B was found to be caused by a homozygous
2.1.3. Dilated cardiomyopathy with conduction defect 1
Dilated cardiomyopathy with conduction defect 1 (CMD1A) is a highly heterogeneous disease, both genetically and phenotypically, with 16 genes currently found to be causatively mutated in cases of CMD1A . Five heterozygotic missense mutations in the
Dilated cardiomyopathy is a serious cardiac condition, in which the heart becomes weakened and enlarged, with downstream effects on the lungs, liver and other organs. Conduction problems and dilated cardiomyopathy arise, leading to frequent heart failure and sudden death events. Affected family members have little or no associated skeletal myopathy.
2.1.4. Malouf syndrome
Malouf syndrome (MLF) is an extremely rare disorder with only a handful of cases described in the literature. The disease has been found to be caused by one of two mutations in exon 1 of the
In males primary testicular failure, and in females premature ovarian failure, is a characteristic feature of the disease. Mild to moderate dilated cardiomyopathy also occurs. Micrognathia and sloping shoulders can give an atypical progeroid phenotype, however in patients suffering from MLF there is no severe growth failure, alopecia, or atherosclerosis .
2.1.5. Heart-hand syndrome, Slovenian type
The heterogeneous family of genetic diseases characterised by both congenital cardiac disease with limb deformities are known as Heart–hand syndromes (HHS). The Heart-hand syndrome, Slovenian type (HHS-S) disorder has been shown to be caused by a mutation (IVS9-12T-G) in intron 9 of the
The characteristic changes to the hands and feet include short distal, and proximal phalanges, as well as webbing or fusion of the fingers or toes. Dilated cardiomyopathy, with an adult-onset progressive conduction disorder is also present, with sudden death due to ventricular tachyarrhythmia [56,57].
Within this section, selected lipodystrophies were detailed, while Table 2 shows a complete listing of known lipodystrophy laminopathies, at the time of writing.
2.2.1. Familial partial lipodystrophy type 2
Familial partial lipodystrophy type 2 (FPLD2; Dunnigan variety of familial partial lipodystrophy) is an autosomal dominant lipodystophy, caused by a heterozygotic mutation in the
FPLD2 shows the characteristic lypodystrophy reduction or loss of subcutaneous adipose tissue in certain regions, starting in childhood, puberty or early adulthood. Patients gradually lose fat from the upper and lower limbs, buttocks and trunk. However intramuscular and bone-marrow fat are preserved. Adipose tissue may increase around the face, neck, back and intra-abdominally . Insulin resistance can occur with consequent complications of diabetes, dyslipidaemia, hypertension and hepatic steatosis. Clinical features may also include abnormalities of the menstrual cycle, hirsutism, and acanthosis nigricans.
2.2.2. Mandibuloacral dysplasia, type A and B
Mandibuloacral dysplasia (MAD) is an autosomal recessive disease, with strongly heterogeneous clinical features. It is categorised into type A (MADA), which is caused by mutations in the
Patients with MADA exhibit an acral loss of adipose tissue and a normal or increased fatty layer in the face, neck and trunk, whereas MADB is marked by a severe progressive glomerulopathy, and generalised lipodystrophy affecting the extremeties, but also the face. Growth retardation, osteolysis of the digits, pigmentary changes, mandibular hypoplasia and skeletal anomalies occur in both variants. Patients may also display some symptoms of progeria, and metabolic disorders such as insulin-resistant diabetes [63,66].
2.3.1. Adult-onset autosomal dominant leukodystrophy
Adult-onset autosomal dominant leukodystrophy (ADLD) is an adult-onset neuropathy, caused by a heterozygous tandem genomic duplication resulting in a duplication of the lamin B1 gene, and a corresponding over-expression of lamin B1 [67,68].
ADLD is slowly progressive, with symptoms becoming apparent in the 40s and 50s, and are markedly similar to progressive multiple sclerosis. These symptoms include symmetric demyelination of the brain and spinal cord, autonomic abnormalities, as well as pyramidal and cerebellar dysfunction. Pathological examination reveals that ADLD differs from progressive multiple sclerosis with a lack of astrogliosis and a preservation of oligodendroglia in the presence of subtotal demyelination .
2.3.2. Charcot-Marie-Tooth disorder
Charcot-Marie-Tooth disorder (CMT) disorder was described simultaneously by Charcot, Marie and Tooth in 1886. Today the disease is considered a spectrum of phenotypically and genetically heterogeneous inherited neuropathies, with over 40 genes known to be associated with the disorder (www.molgen.ua.ac.be/CMTMutations). The autosomal recessive variant, CMT2B1 (AR-CMT2A or CMT4C1) (OMIM: 605588), is known to be caused by a mutation in
Sufferers of CMT2B1 display an early onset muscle wasting in the distal lower limbs (peroneal muscular atrophy syndrome), high arched feet (
2.4. Segmental progeroid diseases
2.4.1. Hutchinson-Gilford progeria syndrome
Hutchinson-Gilford progeria syndrome (HGPS) is an extremely rare, fatal genetic disorder that displays a marked phenotype of premature senility (see chapter on Hutchinson-Gilford progeria syndrome). At least 90% of all HGPS cases are caused by a
Individuals with HGPS are born normally but they present failure to thrive and sclerodermatous skin with loss of subcutaneous fat usually before one year of age. The early symptoms of HGPS also include short stature, and low body weight, which is followed by the occurrence of a tight skin over the abdomen and thighs beginning at the age one or two. Alopecia, scleroderma and the loss of subcutaneous fat also occur at early stages of the disease, succeeded by thin epidermis, fibrosis in the dermis and a loss of skin appendages. Patients often show micrognathia, prominent eyes and veins along with a small beaked nose. Atherosclerosis and calcification of the thoracic aorta is recurrent and death occurs in the early teenage years, most commonly due to cardiovascular complications [76-80].
2.4.2. Restrictive dermopathy
Restrictive dermopathy (RD) is a rare lethal autosomal recessive disease most often caused by loss of function mutations of the
Intrauterine growth retardation is an early sign of RD, along with decreased foetal movement. Thin, translucent, tight skin, as well as joint contractures, respiratory insufficiency, a small pinched nose, micrognathia and mouth in a characteristic fixed ‘o’ shape are the signs of the disease at birth. Usually respiratory failure due to the tight skin leads to a neonatal death within a few weeks of birth [81,82].
2.4.3. Atypical Werner syndrome
First described in 1904 by Otto Werner, Werner syndrome (WS) is caused by mutations in the
WRN is known as ‘progeria of the adult’ and symptoms, such as pubertal growth failure, begin to emerge in the early teenage years. Then in the late teenage years or early 20s, skin atrophy and ulcers, cataracts, type 2 diabetes mellitus, osteoporosis, atherosclerosis, hair greying and alopecia follow. Lipoatrophy and a mild axonal sensorimotor polyneuropathy can also occur. There is also an increased risk of malignancies, reduced fertility and gonadal atrophy. Severe coronary, and peripheral artery disease is also present, and the most common causes of death are myocardial infarction and cancer by a median age of 54 [85,86].
2.5. Overlapping syndromes
2.5.1. Hydrops-Ectopic calcification-moth-eaten skeletal dysplasia
Hydrops-Ectopic calcification-moth-eaten (HEM) skeletal dysplasia is an extremely rare, autosomal recessive lethal chondrodystrophy, which was first described by Greenberg in 1988, in an examination of two sibling foetuses. A 7-bp, homozygous 1599–1605 TCTTCTArCTAGAAG substitution in exon 13 of the lamin B receptor gene (
2.5.2. Pelger-Huet anomaly
Pelger-Huet anomaly (PHA) is a benign, autosomal dominant blood disorder, with characteristic misshapen, hypolobulated nuclei and abnormally course chromatin in blood granulocytes, caused by a mutation in the
Heterozygous patients are clinically normal, while homozygosity has been associated with skeletal dysplasia and early lethality in animal models, although at least one case of non-lethal homozygotic PHA has been found in humans .
2.5.3. Reynolds syndrome
Reynolds syndrome (RP) is caused by a heterozygous mutation in the
RP displays a highly heterogeneous set of clinical features similar to the elements of CREST syndrome (CREST is an acronym that stands for calcinosis, Raynaud's phenomenon, esophageal dysmotility, sclerodactyly, and telangiectasia). These symptoms include scleroderma, liver disease, telangiectasia, eosophageal varicies and Raynaud’s phenomenon .
2.5.4. Osteopoikilosis/Buschke-Ollendorff syndrome
Osteopoikilosis/Buschke-Ollendorff syndrome (BOS) is a highly penetrant, benign, rare, autosomal dominant bone disorder. It is caused by a mutation in the
The osteopoikilosis is revealed by radiographs as numerous and widespread grain- to pea-sized areas of increased bone density, most often in the cancellous bone regions of the epiphyses and metaphyses, although they are found in almost all bones in the body, with the exception of the cranium where they are rarely found. The skin phenotype is manifested as firm lesions, which histologically are revealed to be either elastic-type (juvenile elastoma) or collagen-type (dermatofibrosis lenticularis disseminata) nevi. Joint stiffness may also be present .
2.5.5. Melorheostosis with osteopoikilosis
Melorheostosis with osteopoikilosis (MEL) has been thought to be caused by a mutation in the
MEL is characterised by the flowing hyperostosis of the tubular bone cortices, and sometimes accompanied by abnormalities in surrounding soft-tissue, such as muscle atrophy, joint-contractures, epidermal lesions or hemangiomas .
3. Linking genotype and phenotype of laminopathies
A marked change in heterochromatin is one of the most apparent features noted when examining cells affected by laminopathies, from loci of diminished or clumped heterochromatin to total loss of peripheral heterochromatin [99-103]. This alteration of normal heterochromatin, coupled with the known interactions between lamins and gene regulatory proteins, defines a major constituent for the molecular mechanism behind laminopathies . Lamins have been shown to interact with proteins of the inner nuclear membrane (emerin, myne-1, nesprin, LAP1 and LAP2, LBR and MAN1), and chromatin-associated proteins (H2a, H2B, H3-H4, Ha95, HP1 and BAF) [105-109]. These associations allow for gene silencing by means of heterochromatin reorganisation, which could be a causative factor for phenotypic changes . Recruiting genes selectively to the inner nuclear membrane has also been shown to result in their transcriptional repression . The tissue specific gene regulatory role of lamins is thought to underlie the tissue-specific symptoms observed in laminopathies [34,110]. Tissue specific regulation of lamin A expression may also be an explanatory factor for tissue-specific symptoms. Low-level of prelamin A expression in the brain has been shown to be due to a brain-specific microRNA, miRNA-9 , and miR-9 overexpression has been shown to alleviate nuclear blebbing in non-neural cells .
A mouse model with the
The possibility of complex interactions between these different causative mechanisms, the complex multirole functionality of lamins, along with widely varying environmental and genetic co-factors affecting this spectrum of processes, would afford a possible explanation for the heterogeneity of disease effects amongst the sufferers of laminopathies [48,118,119]. This variance of disease is one of the most fascinating aspects of laminopathies, the disparity between how a very large family of mutations affecting many genes give rise to diseases with such interrelated clinical features, and on the other hand how even amongst members of a single family carrying the same mutation, disease manifestations are diverse and variable. In AD-EDMD, heterozygous mutations in the
The diversity of disease phenotypes in consanguineous patients with identical mutations, such as disease onset, severity and progress, indicates that laminopathies are strongly influenced by disease modifiers such as genetic or environmental factors. For example, female sufferers of FPLD2 exhibit a more pronounced phenotype than male , family members with BOS can have both or just one of the bone and skin manifestations of that disease. Different missense mutations at the same locus can also give rise to different laminopathies. For example, in the
An altered nuclear integrity, leading to a weakness in cell structure and a susceptibility to mechanical stress as a constituent of the causative mechanism for laminopathies is supported by the specificity of some laminopathies, such as HGPS or EDMD, to tissues affected by high levels of mechanical stress (the skin, muscles and aortic arch), as well as the similarity of muscular dystrophies caused by mutations in genes responsible for karyoskeleton, cytoskeleton and myotubule proteins to laminopathic muscular dystrophies. The unique expression pattern of lamins in muscle cells might also illuminate a causative system for laminopathies. As no lamin B1 is expressed in muscle cells at all, when
Various hypotheses have been put forward to account for the muscle cell specificity of EDMD . Muscle cells contain very low or undetectable amounts of lamin B1, whereas in most other cell types lamin B1 is a major lamin, leaving muscle cells more sensitive to loss of function of either emerin or lamin A/C [42,136]. Emerin may also interact with transcription factors or directly with DNA to cause specific gene regulation in muscle cells . Finally, muscle cells also undergo mechanical stress, and emerin, as part of a nucleo-cytoskeletal system may have a protective role against mechanical stress .
Lamin A mutations have also been shown to cause premature exhaustion of somatic stem cell populations, as well as stem cell dysfunction. As adult somatic stem cell population is depleted, tissues undergoing a high rate of turnover, such as the skin, would be affected first [139,140].
A consistent relationship between mutation location on the
It was suspected that a duplication of the
Finally, a link between levels of progerin produced in laminopathies that exhibit an accumulation of the mutant lamin A/C precursor, and both the severity and age of onset of the phenotype has been shown. RD is considered to be similar but more severe than HGPS, with a correspondingly higher rate of prelamin A accumulation [143,144]. Two cases of a Werner syndrome-like form of progeria displayed a progeria-like aspect with middle age onset coronary artery disease, with a level of progerin that was one quarter of that seen in HGPS cells . Further proof of the toxicity of accumulated progerin is shown by the decrease of progerin levels in cell cultures by treatment with rapamycin, with a resultant rescue of the phenotype . As allele dependant differences in expression of the
These details paint a complex picture of a heterogeneous family of mutations resulting in varying and overlapping phenotypes, with a diversity in severity and age of onset resulting from tissue specific gene regulation, site of mutation and various genetic and possibly environmental co-factors.
4. Mouse models
Mouse models have yielded invaluable knowledge about the functions of the
Mice that were thought to completely lack A-type lamin expression were created in order to study a model with no expressed lamin A/C. These
Mice with the
|These ||Postnatal lethality, with cardiomyopathy and muscular dystrophy||[101,147]|
|These mice have a total loss of lamin A/C.||Growth retardation, developmental heart defects, skeletal muscle hypotrophy, decreased subcutaneous adipose tissue. Death occurs at 2 to 3 weeks |||
|These lamin C only mice carry a mutant Lmna allele that yields lamin C exclusively, without lamin A.||No disease phenotypes and a normal lifespan.|||
|Mature lamin A only mouse, bypassing prelamin A synthesis and processing.||No detectable pathology, fibroblasts show misshapen nuclei.|||
|These mice are null for the endoprotease responsible for the final cleavage step in prelamin A maturation, leading to an accumulation of farnesylated pre-lamin A.||Mice have rib fractures, osteoporosis, muscle weakness and die at 6–7 months.|||
|Postnatal growth retardation, shortened lifespan, loss of fat layer and muscular dystrophy.|||
|These mice have a missense CDM1A-associated lamin A mutation, N195K.||Postnatal death associated with cardiomyopathy. MEFs showed nuclear abnormalities.|||
|These mice have a missense EDMD-associated lamin A mutation, H222P.||These mice show a stiff walking posture and cardiac dysfunction. Death occurs by 9 months of age. MEFs showed nuclear abnormalities.|||
|These mice carry an||Heterozygous mice, |||
|These mice have a L530P mutation in the lamin A gene that is associated with EDMD in humans.||Homozygous mice display defects consistent with HGPS, and die within 4-5 weeks of birth.|||
|cDNA with mis-sense mutation expressed with a heart specific promoter.||Cardiomyopathy and early postnatal lethality|||
|The wild-type mouse ||Growth retardation, weight loss, cardiovascular problems and shortened lifespan.|||
|These mice have an insertional mutation in ||Mice survive embryonic development, however die at birth with lung and bone defects.|||
|These mice do not express emerin.||Mice overtly normal but with slightly retarded muscle regeneration.||[158,159]|
A mouse model for EDMD was created by knocking out the
Knock-in mouse models such as the
A mouse model where only lamin-C is produced (
In order to study early post-natal development effects caused by loss of lamin A/C, an
A mouse model was created using a heart-selective promoter (α-myosin heavy chain promoter) to control the expression of human normal lamin A, and lamin A containing the EDMD causing mutation M371K. Mice expressing the wild-type human lamin A were born at slightly less than expected rates, and had a normal lifespan. However, mice expressing mutant M371K lamin A exhibited a much higher risk of prenatal death, and were born at only a fraction (0.07) of the expected frequency. Those animals that were born died within 2-7 weeks, and displayed pulmonary and cardia edema. Cardiac cells from these mice showed abnormal, convoluted nuclear envelopes with clumped chromatin and intranuclear foci of lamins .
Mouse models of laminopathies are limited by the gross physiological differences between rodent (mouse models being the most relevant models used to investigate laminopathies) and human. However, despite the limitations of mouse models, the advantages are legion; being able to study very rare diseases at any stage of disease, with limitless sampling, temporal and physically controlled expression of mutant protein, and with the possibilities for testing different type of treatment.
Current treatments for laminopathies are largely symptomatic, controlling the secondary effects of the disease. Corrective surgery is used to treat the EDMD contractures, coronary artery bypass surgery for HGPS, pacemaker installation or heart transplantation for DCMI or LGMD1B patients . FPLD2 patients with diabetes mellitus and hypertension are treated with antidiabetic drugs, angiotensin converting enzyme inhibitors, calcium channel blockers and beta blockers [52,165,166]. The administration of a recombinant methionyl human leptin has been tried with some success in patients suffering from FPLD, giving rise to improved fasting glucose concentrations, insulin sensitivity, and triglyceride levels [167,168]. The impairment of pre-adipocyte differentiation, an impairment which is brought about by the negative effects of prelamin A accumulation on the rate of DNA-bound SREBP1, may also be treated with troglitazone, a PPAR-gamma ligand which promotes the adipogenic program .
Curative treatment for laminopathies that are autosomal-recessive involving loss-of-function of a protein, such as EDMD-AR, would require the expression of a healthy wild-type allele in the affected tissue. However, autosomal dominant laminopathies require a more complex treatment, in which the production, modification and/or the effect of the mutant protein also need to be eliminated. For example, in a phase II clinical trial with HGPS patients, lonafarnib, a farnesyl transferase inhibitor (FTI) is being given as treatment (see chapter on Hutchinson-Gilford progeria syndrome) . FTI is normally used as an anti-tumour treatment, but it also reduces the amount of progerin produced by inhibiting the farnesylation of prelamin A. Previous experiments with FTIs in cell cultures showed marked improvements, with a reduction of misshapen nuclei . With mouse models for HGPS an improvement in disease phenotype was noted, although no total reversal was apparent [172-176]. This may be due to the fact that although FTI treatment inhibits the farnesylation of prelamin-A by farnesyl transferase, a secondary modification pathway, a geranylgeranylation by geranylgeranyltransferase, allows prelamin A to be processed into progerin despite the FTI treatment . However, a combination of statins (a potent HMG-CoA reductase inhibitor, used to inhibit the production of cholesterol in the liver) and bisphosphonates (a class of drugs used to treat osteoporosis), was used to inhibit the synthesis of farnesyl pyrophosphate, a co-substrate of farnesyltransferase and a precursor of a substrate for geranylgeranyltransferase I. This combination inhibits prenylation, and when used to treat laminopathies, resulted in an increased longevity, reduced oxidative stress, cellular senescence and improved phenotype in mice [61,154,172,178,179]. A triple drug trial was initiated in 2009 to examine the efficacy of treatment involving an FTI, a statin and a bisphosphonate, however the results of this trial have not yet been made public.
Long-term treatment with FTIs is not without risks. All CaaX box/motif proteins would have their farnesylation processing inhibited, which would mean an inhibition of lamin-B maturation. Non-farnesylated lamins might also accumulate in the cell, with unexpected effects. In a mice model where non-farnesylated prelamin-A was solely expressed, with the CaaX motif/box mutated to SAAX, a cardiomyopathy was observed to occur . In HIV treatment, acquired lipodystrophy is a possible side-effect of the use of HIV protease inhibitors, which cause pre-lamin A accumulation . This pre-lamin accumulation was also observed in fibroblasts from FPLD2 patients, further hinting at the toxicity of pre-lamin A accumulation .
Rapamycin, an immunosuppressant antibiotic drug, has also been examined as a possible treatment in laminopathies. Rapamycin treatment in HGPS cell cultures resulted in reduced nuclear blebbing and decreased rates of senescence, as well as a marked reduction of progerin and prelamin A levels, a restoration of wildtype LAP2α, BAF and trimethylated H3K9 organisation, and a rescue of the normal chromatin phenotype. These effects come about by means of autophagic degradation of prelamin A, triggered by inactivation of the inhibitory mammalian target of rapamycin (mTOR) dependent pathway [145,182]. In an
Pre-lamin A antisense oligonucleotides were used to reduce pre-lamin A levels, with a resultant decrease in misshapen nuclei. The most common HGPS point mutation causes an increased usage of a cryptic splice site in exon 11, CAG#GTGGGC, which is also used at near-undetectable levels in wild-type cells. Antisense morpholino oligonucleotides directed to this site resulted in an improvement of HGPS fibroblast disease phenotype . RNA interference has also been used to successfully improve proliferation and nuclear morphology, as well as reducing senescence in fibroblasts expressing mutant lamin A . In another experiment exon 11 splice donor site antisense oligonucleotides were also used to promote the alternative splice pathway, leading to an increased in progerin production in fibroblast cells, and short hairpin RNA (shRNA) were then used to diminish this production in fibroblasts, leading to an improvement of phenotype . Morpholinos have also been used to target the cryptic splicing event in mouse. The use of antisense morpholinos to the exon 10 lamin A splice donor site and the c.1827C>T;p.G609G mutation of the
During the last decade the number of diseases found to be caused by mutations in lamin or lamin associated genes has increased significantly. These phenotypically diverse diseases have been categorised both phenotypically and genetically, and today research is focused on both deciphering the pathogenic mechanisms behind their pathophysiological processes, as well as understanding how such diverse pathologies can arise from this related family of mutations. During that time the appreciated role for lamins has changed from being regarded merely as a structural scaffold for the nucleus, to a key element in DNA replication and transcription, chromatin organisation, cell replication and differentiation. Future research is sure to continue at an ever-increasing pace, especially as the development and integration of next generation sequencing technologies and technologies that allows for global analysis of the genome and epigenome into both research and clinical settings. For researchers this level of genomic interrogation brings about unprecedented access to new information about our genome, which will be valuable for the creation of maps of genetic and possibly epigenetic variation that influence disease.
The laminopathies described in this review are without a doubt, exceedingly rare. However by researching these rare conditions, it is hoped that we can shed light on their all too common clinical symptoms, such as cardiac disease, metabolic disorders such as insulin resistance, and even ageing itself.
Our work is supported by a VINNMER fellow grant from VINNOVA, and an Innovator grant from The Progeria Research Foundation. We thank the patients, and Dr. Nicola Carboni and Dr Marco Mura for contributing photos of patients. Primary fibroblast cultures were obtained from the Aging Repository of the Coriell Cell Repository.