Cardiovascular Outcomes in Aortopathy: GenTAC Registry of Genetically Triggered Aortic Aneurysms and Related Conditions.

BACKGROUND: The GenTAC (Genetically Triggered Thoracic Aortic Aneurysm and Cardiovascular Conditions) Registry enrolled patients with genetic aortopathies between 2007 and 2016. OBJECTIVES: The purpose of this study was to compare age distribution and probability of elective surgery for proximal aortic aneurysm, any dissection surgery, and cardiovascular mortality among aortopathy etiologies. METHODS: The GenTAC study had a retrospective/prospective design. Participants with bicuspid aortic valve (BAV) with aneurysm (n = 879), Marfan syndrome (MFS) (n = 861), nonsyndromic heritable thoracic aortic disease (nsHTAD) (n = 378), Turner syndrome (TS) (n = 298), vascular Ehlers-Danlos syndrome (vEDS) (n = 149), and Loeys-Dietz syndrome (LDS) (n = 121) were analyzed. RESULTS: The 25% probability of elective proximal aortic aneurysm surgery was 30 years for LDS (95% CI: 18-37 years), followed by MFS (34 years; 95% CI: 32-36 years), nsHTAD (52 years; 95% CI: 48-56 years), and BAV (55 years; 95% CI: 53-58 years). Any dissection surgery 25% probability was highest in LDS (38 years; 95% CI: 33-53 years) followed by MFS (51 years; 95% CI: 46-57 years) and nsHTAD (54 years; 95% CI: 51-61 years). BAV experienced the largest relative frequency of elective surgery to any dissection surgery (254/33 = 7.7), compared with MFS (273/112 = 2.4), LDS (35/16 = 2.2), or nsHTAD (82/76 = 1.1). With MFS as the reference population, risk of any dissection surgery or cardiovascular mortality was lowest in BAV patients (HR: 0.13; 95% CI: 0.08-0.18; HR: 0.13; 95%: CI: 0.06-0.27, respectively). The greatest risk of mortality was seen in patients with vEDS. CONCLUSIONS: Marfan and LDS cohorts demonstrate age and event profiles congruent with the current understanding of syndromic aortopathies. BAV events weigh toward elective replacement with relatively few dissection surgeries. Nonsyndromic HTAD patients experience near equal probability of dissection vs prophylactic surgery, possibly because of failure of early diagnosis.

Elevated expression levels of lysyl oxidases protect against aortic aneurysm progression in Marfan syndrome.

Patients with Marfan syndrome (MFS) are at high risk of life-threatening aortic dissections. The condition is caused by mutations in the gene encoding fibrillin-1, an essential component in the formation of elastic fibers. While experimental findings in animal models of the disease have shown the involvement of transforming growth factor-ß (TGF-ß)- and angiotensin II-dependent pathways, alterations in the vascular extracellular matrix (ECM) may also play a role in the onset and progression of the aortic disease. Lysyl oxidases (LOX) are extracellular enzymes, which initiates the formation of covalent cross-linking of collagens and elastin, thereby contributing to the maturation of the ECM. Here we have explored the role of LOX in the formation of aortic aneurysms in MFS. We show that aortic tissue from MFS patients and MFS mouse model (Fbn1(C1039G/+)) displayed enhanced expression of the members of the LOX family, LOX and LOX-like 1 (LOXL1), and this is associated with the formation of mature collagen fibers. Administration of a LOX inhibitor for 8weeks blocked collagen accumulation and aggravated elastic fiber impairment, and these effects correlated with the induction of a strong and rapidly progressing aortic dilatation, and with premature death in the more severe MFS mouse model, Fbn1(mgR/mgR), without any significant effect on wild type animals. This detrimental effect occurred preferentially in the ascending portion of the aorta, with little or no involvement of the aortic root, and was associated to an overactivation of both canonical and non-canonical TGF-ß signaling pathways. The blockade of angiotensin II type I receptor with losartan restored TGF-ß signaling activation, normalized elastic fiber impairment and prevented the aortic dilatation induced by LOX inhibition in Fbn1(C1039G/+) mice. Our data indicate that LOX enzymes and LOX-mediated collagen accumulation play a critical protective role in aneurysm formation in MFS.

GenTAC registry report: gender differences among individuals with genetically triggered thoracic aortic aneurysm and dissection.

Previous data suggest women are at increased risk of death from aortic dissection. Therefore, we analyzed data from the GenTAC registry, the NIH-sponsored program that collects information about individuals with genetically triggered thoracic aortic aneurysms and cardiovascular conditions. We performed cross-sectional analyses in adults with Marfan syndrome (MFS), familial thoracic aortic aneurysm or dissection (FTAAD), bicuspid aortic valve (BAV) with thoracic aortic aneurysm or dissection, and subjects under 50 years of age with thoracic aortic aneurysm or dissection (TAAD <50 years). Women comprised 32% of 1,449 subjects and were 21% of subjects with BAV, 34% with FTAAD, 22% with TAAD <50 years, and 47% with MFS. Thoracic aortic dissections occurred with equal gender frequency yet women with BAV had more extensive dissections. Aortic size was smaller in women but was similar after controlling for BSA. Age at operation for aortic valve dysfunction, aneurysm or dissection did not differ by gender. Multivariate analysis (adjusting for age, BSA, hypertension, study site, diabetes, and subgroup diagnoses) showed that women had fewer total aortic surgeries (OR = 0.65, P < 0.01) and were less likely to receive angiotensin converting enzyme inhibitors (ACEi; OR = 0.68, P < 0.05). As in BAV, other genetically triggered aortic diseases such as FTAAD and TAAD <50 are more common in males. In women, decreased prevalence of aortic operations and less treatment with ACEi may be due to their smaller absolute aortic diameters. Longitudinal studies are needed to determine if women are at higher risk for adverse events.

The haploinsufficient Col3a1 mouse as a model for vascular Ehlers-Danlos syndrome.

Vascular Ehlers-Danlos syndrome is a rare genetic disorder resulting from mutations in the α-1 chain of type III collagen (COL3A1) and manifesting as tissue fragility with spontaneous rupture of the bowel, gravid uterus, or large or medium arteries. The heterozygous Col3a1 knockout mouse was investigated as a model for this disease. The collagen content in the abdominal aorta of heterozygotes was reduced, and functional testing revealed diminishing wall strength of the aorta in these mice. Colons were grossly and histologically normal, but reduced strength and increased compliance of the wall were found in heterozygotes via pressure testing. Although mice demonstrated no life-threatening clinical signs or gross lesions of vascular subtype Ehlers-Danlos syndrome type IV, thorough histological examination of the aorta of heterozygous mice revealed the presence of a spectrum of lesions similar to those observed in human patients. Lesions increased in number and severity with age (0/5 [0%] in 2-month-old males vs 9/9 [100%] in 14-month-old males, P < .05) and were more common in male than female mice (23/26 [88.5%] vs 14/30 [46.7%] in 9- to 21-month-old animals, P < .05). Haploinsufficiency for Col3a1 in mice recapitulates features of vascular Ehlers-Danlos syndrome in humans and can be used as an experimental model.

Mutations in fibrillin-1 cause congenital scleroderma: stiff skin syndrome.

The predisposition for scleroderma, defined as fibrosis and hardening of the skin, is poorly understood. We report that stiff skin syndrome (SSS), an autosomal dominant congenital form of scleroderma, is caused by mutations in the sole Arg-Gly-Asp sequence-encoding domain of fibrillin-1 that mediates integrin binding. Ordered polymers of fibrillin-1 (termed microfibrils) initiate elastic fiber assembly and bind to and regulate the activation of the profibrotic cytokine transforming growth factor-beta (TGFbeta). Altered cell-matrix interactions in SSS accompany excessive microfibrillar deposition, impaired elastogenesis, and increased TGFbeta concentration and signaling in the dermis. The observation of similar findings in systemic sclerosis, a more common acquired form of scleroderma, suggests broad pathogenic relevance.

Neuroradiologic manifestations of Loeys-Dietz syndrome type 1.

BACKGROUND AND PURPOSE: Loeys-Dietz syndrome (LDS) is a recently described entity that has the triad of arterial tortuosity and aneurysms, hypertelorism, and bifid uvula or cleft palate. Its neuroradiologic manifestations have not been well delineated. We sought to describe the neuroradiologic features of LDS and to assess the manifestations that would warrant follow-up imaging. MATERIALS AND METHODS: Two neuroradiologists retrospectively reviewed CT angiography (CTA), MR imaging, and plain film studies related to the head and neck in 25 patients ranging from 1 to 55 years of age, all of whom had positive genetic testing and clinical characteristics of LDS. Arterial tortuosity was evaluated by subjective assessment of 2D and 3D volumetric CTA and MR angiography data. Craniosynostosis and spinal manifestations were assessed by using plain films and CT images. MR images mostly of the head were reviewed for associated findings such as hydrocephalus, Chiari malformation, etc. Clinical manifestations were collated from the electronic patient record. RESULTS: All patients had extreme arterial tortuosity, which is characteristic of this syndrome. Thirteen patients had scoliosis, 12 had craniosynostosis, 8 had intracranial aneurysms, 6 had spinal instability, 3 had dissections of the carotid and vertebrobasilar arteries, 3 had hydrocephalus, 4 had dural ectasia, 2 had a Chiari malformation, and 1 had intracranial hemorrhage as a complication of vascular dissection. CONCLUSIONS: Significant neuroradiologic manifestations are associated with LDS, predominantly arterial tortuosity. Most of the patients in this series were young and, therefore, may require serial CTA monitoring for development of intra- and extracranial dissections and aneurysms, on the basis of the fact that most of the patients with pseudoaneurysms and dissection were older at the time of imaging. Other findings of LDS such as craniosynostosis, Chiari malformation, and spinal instability may also need to be addressed.

Effect of mutation type and location on clinical outcome in 1,013 probands with Marfan syndrome or related phenotypes and FBN1 mutations: an international study.

Mutations in the fibrillin-1 (FBN1) gene cause Marfan syndrome (MFS) and have been associated with a wide range of overlapping phenotypes. Clinical care is complicated by variable age at onset and the wide range of severity of aortic features. The factors that modulate phenotypical severity, both among and within families, remain to be determined. The availability of international FBN1 mutation Universal Mutation Database (UMD-FBN1) has allowed us to perform the largest collaborative study ever reported, to investigate the correlation between the FBN1 genotype and the nature and severity of the clinical phenotype. A range of qualitative and quantitative clinical parameters (skeletal, cardiovascular, ophthalmologic, skin, pulmonary, and dural) was compared for different classes of mutation (types and locations) in 1,013 probands with a pathogenic FBN1 mutation. A higher probability of ectopia lentis was found for patients with a missense mutation substituting or producing a cysteine, when compared with other missense mutations. Patients with an FBN1 premature termination codon had a more severe skeletal and skin phenotype than did patients with an inframe mutation. Mutations in exons 24-32 were associated with a more severe and complete phenotype, including younger age at diagnosis of type I fibrillinopathy and higher probability of developing ectopia lentis, ascending aortic dilatation, aortic surgery, mitral valve abnormalities, scoliosis, and shorter survival; the majority of these results were replicated even when cases of neonatal MFS were excluded. These correlations, found between different mutation types and clinical manifestations, might be explained by different underlying genetic mechanisms (dominant negative versus haploinsufficiency) and by consideration of the two main physiological functions of fibrillin-1 (structural versus mediator of TGF beta signalling). Exon 24-32 mutations define a high-risk group for cardiac manifestations associated with severe prognosis at all ages.

Extracellular microfibrils in development and disease.

Fibrillins are the structural components of extracellular microfibrils that impart physical properties to tissues, alone or together with elastin as elastic fibers. Genetic studies in mice have revealed that fibrillin-rich microfibrils are also involved in regulating developmental programs and homeostatic processes through the modulation of TGF-beta/BMP signaling events. A new paradigm has thus emerged whereby the spatiotemporal organization of microfibrils dictates both the cellular activities and physical properties of connective tissues. These observations have paved the way to novel therapeutic approaches aimed at counteracting the life-threatening complications in human conditions caused by dysfunctions of fibrillin-rich microfibrils.

Endothelial dysfunction and compromised eNOS/Akt signaling in the thoracic aorta during the progression of Marfan syndrome.

BACKGROUND AND PURPOSE: Aortic complications account for the major mortality in Marfan syndrome (MFS), a connective tissue disorder caused by mutations in FBN1 encoding fibrillin-1. We hypothesized that MFS impaired endothelial function and nitric oxide (NO) production in the aorta. EXPERIMENTAL APPROACH: Mice (at 3, 6, 9 and 12 months of age) heterozygous for the Fbn1 allele encoding a cysteine substitution (Fbn1 (C1039G/+), Marfan mice, n=75), the most common class of mutation in MFS, were compared with age-matched control littermates (n=75). Thoracic and abdominal aortas from the two groups were studied. KEY RESULTS: Isometric force measurements revealed that relaxation to ACh (but not to sodium nitroprusside) was diminished in the phenylephrine-precontracted Marfan thoracic aorta at 6 months of age (pEC(50)=6.12+/-0.22; maximal response, E(max)=52.7+/-6.8%; control: pEC(50)=7.34+/-0.19; E(max)=84.8+/-2.2%). At one year, both inhibition of NO production with N(omega)-nitro-L-arginine methyl ester, or denudation of endothelium increased the phenylephrine-stimulated contraction in the control thoracic aorta by 35%, but had no effect in the Marfan aorta, indicating a loss of basal NO production in the Marfan vessel. From 6 months, a reduced phosphorylation of endothelial NOS (eNOS)(Ser1177) and Akt(Thr308) detected by Western blotting was observed in the Marfan thoracic aorta, which was accompanied by decreased levels of cGMP. Expressions of Akt and eNOS in the abdominal aorta were not different between the two groups. CONCLUSIONS AND IMPLICATIONS: MFS impairs endothelial function and signaling of NO production in the thoracic aorta, suggesting the importance of NO in the age-related progression of thoracic aortic manifestations.

The molecular genetics of Marfan syndrome and related disorders.

Marfan syndrome (MFS), a relatively common autosomal dominant hereditary disorder of connective tissue with prominent manifestations in the skeletal, ocular, and cardiovascular systems, is caused by mutations in the gene for fibrillin-1 (FBN1). The leading cause of premature death in untreated individuals with MFS is acute aortic dissection, which often follows a period of progressive dilatation of the ascending aorta. Recent research on the molecular physiology of fibrillin and the pathophysiology of MFS and related disorders has changed our understanding of this disorder by demonstrating changes in growth factor signalling and in matrix-cell interactions. The purpose of this review is to provide a comprehensive overview of recent advances in the molecular biology of fibrillin and fibrillin-rich microfibrils. Mutations in FBN1 and other genes found in MFS and related disorders will be discussed, and novel concepts concerning the complex and multiple mechanisms of the pathogenesis of MFS will be explained.

Interaction between krit1 and icap1alpha infers perturbation of integrin beta1-mediated angiogenesis in the pathogenesis of cerebral cavernous malformation.

Cerebral cavernous malformation (CCM) is a common autosomal dominant disorder characterized by venous sinusoids that predispose to intracranial hemorrhage. CCM is genetically heterogeneous, with loci at 7q, 7p and 3q. Mutations in KRIT1 account for all cases linked to 7q (CCM1), but the pathogenesis of CCM is not understood. Krev Interaction Trapped 1 (krit1) was originally identified through its interaction with the Ras-family GTPase krev1/rap1a in a two-hybrid screen, inferring a role in GTPase signaling cascades. We demonstrated additional 5'-coding exons for krit1, extending the N-terminus by 207 amino acids compared to the previously reported protein. Remarkably, by two-hybrid analysis and co-immunoprecipitation, full-length krit1 fails to interact with krev1/rap1a but shows strong interaction with integrin cytoplasmic domain-associated protein-1 (icap1). Icap1 binds to a NPXY motif in the cytoplasmic domain of beta1 integrin and participates in beta1-mediated cell adhesion and migration. The novel N-terminus of krit1 contains a NPXY motif that it is required for icap1 interaction. Like beta1 integrin, krit1 interacts with the 200 amino acid isoform of icap1 (icap1alpha), but not a 150 amino acid form that results from alternative splicing (icap1beta). In a competition assay, induced expression of krit1 diminishes the interaction between icap1alpha and beta1 integrin. Taken together, these data suggest that beta1 integrin and krit1 compete for the same site on icap1alpha, perhaps constituting a regulatory mechanism. Loss-of-function KRIT1 mutations, as observed in CCM1, would shift the balance with predicted consequences for endothelial cell performance during integrin beta1-dependent angiogenesis.

When the message goes awry: disease-producing mutations that influence mRNA content and performance.

Mutations that cause disease commonly occur in the coding sequence and directly influence protein structure and function. However, many diseases result from mutations that influence various aspects of mRNA metabolism, including processing, export, stability, and translational control.

Familial thoracic aortic aneurysms and dissections: genetic heterogeneity with a major locus mapping to 5q13-14.

BACKGROUND: Aneurysms and dissections affecting the ascending aorta are associated primarily with degeneration of the aortic media, called medial necrosis. Families identified with dominant inheritance of thoracic aortic aneurysms and dissections (TAA/dissections) indicate that single gene mutations can cause medial necrosis in the absence of an associated syndrome. METHODS AND RESULTS: Fifteen families were identified with multiple members with TAAs/dissections. DNA from affected members from 2 of the families was used for a genome-wide search for the location of the defective gene by use of random polymorphic markers. The data were analyzed by the affected-pedigree-member method of linkage analysis. This analysis revealed 3 chromosomal loci with multiple markers demonstrating evidence of linkage to the phenotype. Linkage analysis using further markers in these regions and DNA from 15 families confirmed linkage of some of the families to 5q13-14. Genetic heterogeneity for the condition was confirmed by a heterogeneity test. Data from 9 families with the highest conditional probability of being linked to 5q were used to calculate the pairwise and multipoint logarithm of the odds (LOD) scores, with a maximum LOD of 4.74, with no recombination being obtained for the marker D5S2029. In 6 families, the phenotype was not linked to the 5q locus. CONCLUSIONS: A major locus for familial TAAs and dissections maps to 5q13-14, with the majority (9 of 15) of the families identified demonstrating evidence of linkage to this locus. The condition is genetically heterogeneous, with 6 families not demonstrating evidence of linkage to any loci previously associated with aneurysm formation.

A strategy for disease gene identification through nonsense-mediated mRNA decay inhibition.

Premature termination codons (PTCs) have been shown to initiate degradation of mutant transcripts through the nonsense-mediated messenger RNA (mRNA) decay (NMD) pathway. We report a strategy, termed gene identification by NMD inhibition (GINI), to identify genes harboring nonsense codons that underlie human diseases. In this strategy, the NMD pathway is pharmacologically inhibited in cultured patient cells, resulting in stabilization of nonsense transcripts. To distinguish stabilized nonsense transcripts from background transcripts upregulated by drug treatment, drug-induced expression changes are measured in control and disease cell lines with complementary DNA (cDNA) microarrays. Transcripts are ranked by a nonsense enrichment index (NEI), which relates expression changes for a given transcript in NMD-inhibited control and patient cell lines. The most promising candidates can be selected using information such as map location or biological function; however, an important advantage of the GINI strategy is that a priori information is not essential for disease gene identification. GINI was tested on colon cancer and Sandhoff disease cell lines, which contained previously characterized nonsense mutations in the MutL homolog 1 (MLH1) and hexosaminidase B (HEXB) genes, respectively. A list of genes was produced in which the MLH1 and HEXB genes were among the top 1% of candidates, thus validating the strategy.

Characterization of microsatellite markers flanking FBN1: utility in the diagnostic evaluation for Marfan syndrome.

Marfan syndrome (MFS) is an autosomal dominant disorder of connective tissue with marked interfamilial and intrafamilial variation in phenotype. The primary defect in affected patients resides in the gene for fibrillin-1 (FBN1) on 15q21. Linkage analysis has shown no locus heterogeneity in the classic phenotype, although substantial allelic heterogeneity exists. Recently it has been shown that the size of the gene is approximately 200 kb. These and other factors have precluded routine mutation screening for presymptomatic and prenatal diagnosis. Previously we described four intragenic microsatellite polymorphisms that can be used for haplotype segregation analysis. The utility of this approach is limited because the markers do not fully span the gene and show incomplete informativeness, with 16% homozygosity for the most common haplotype. We have now identified and localized highly polymorphic microsatellite markers that fall within 1 Mb of FBN1. Complete haplotype heterozygosity was observed in a population of 50 unrelated control individuals when the flanking markers and existing intragenic polymorphisms were used in combination. We demonstrate the utility of haplotype segregation analysis in the presymptomatic diagnosis and counseling of families showing atypical or equivocal manifestations of MFS.

Phenotypic alteration of vascular smooth muscle cells precedes elastolysis in a mouse model of Marfan syndrome.

Marfan syndrome is associated with early death due to aortic aneurysm. The condition is caused by mutations in the gene (FBN1) encoding fibrillin-1, a major constituent of extracellular microfibrils. Prior observations suggested that a deficiency of microfibrils causes failure of elastic fiber assembly during late fetal development. Mice homozygous for a targeted hypomorphic allele (mgR) of Fbn1 revealed a predictable sequence of abnormalities in the vessel wall including elastic fiber calcification, excessive deposition of matrix elements, elastolysis, and intimal hyperplasia. Here we describe previously unrecognized concordant findings in elastic vessels from patients with Marfan syndrome. Furthermore, ultrastructural analysis of mgR mice revealed cellular events that initiate destructive changes. The first detectable abnormality was an unusually smooth surface of elastic laminae, manifesting the loss of cell attachments that are normally mediated by fibrillin-1. Adjacent cells adopted alteration in their expression profile accompanied by morphological changes but retained expression of vascular smooth muscle cell markers. The abnormal synthetic repertoire of these morphologically abnormal smooth muscle cells in early vascular lesions included elastin, among other matrix elements, and matrix metalloproteinase 9, a known mediator of elastolysis. Ultimately, cell processes associated with zones of elastic fiber thinning and fragmentation. These data suggest that the loss of cell attachments signals a nonproductive program to synthesize and remodel an elastic matrix. This refined understanding of the pathogenesis of vascular disease in Marfan syndrome will facilitate the development of therapeutic strategies.

Rent1, a trans-effector of nonsense-mediated mRNA decay, is essential for mammalian embryonic viability.

The ability to detect and degrade transcripts that lack full coding potential is ubiquitous but non-essential in lower eukaryotes, leaving in question the evolutionary basis for complete maintenance of this function. One hypothesis holds that nonsense-mediated RNA decay (NMD) protects the organism by preventing the translation of truncated peptides with dominant negative or deleterious gain-of-function potential. All organisms studied to date that are competent for NMD express a structural homolog of Saccharomyces cerevisiae Upf1p. We have now explored the consequences of loss of NMD function in vertebrates through targeted disruption of the Rent1 gene in murine embryonic stem cells which encodes a mammalian ortholog of Upf1p. Mice heterozygous for the targeted allele showed no apparent phenotypic abnormalities but homozygosity was never observed, demonstrating that Rent1 is essential for embryonic viability. Homozygous targeted embryos show complete loss of NMD and are viable in the pre-implantation period, but resorb shortly after implantation. Furthermore, Rent1(-/-) blastocysts isolated at 3.5 days post-coitum undergo apoptosis in culture following a brief phase of cellular expansion. These data suggest that NMD is essential for mammalian cellular viability and support a critical role for the pathway in the regulated expression of selected physiologic transcripts.

Familial Tetralogy of Fallot caused by mutation in the jagged1 gene.

Tetralogy of Fallot (ToF) is the most common form of complex congenital heart disease, occurring in approximately 1 in 3000 live births. Evaluation of candidate loci in a large kindred segregating autosomal dominant ToF with reduced penetrance culminated in identification of a missense mutation (G274D) in JAG1, the gene encoding jagged1, a Notch ligand expressed in the developing right heart. Nine of eleven mutation carriers manifested cardiac disease, including classic ToF, ventricular septal defect with aortic dextroposition and isolated peripheral pulmonic stenosis (PPS). All forms of ToF were represented, including variants with pulmonic stenosis, pulmonic atresia and absent pulmonary valve. No individual within this family met diagnostic criteria for any previously described clinical syndrome, including Alagille syndrome (AGS), caused by haploinsufficiency for jagged1. All mutation carriers had characteristic but variable facial features, including long, narrow and upslanting palpebral fissures, prominent nasal bridge, square dental arch and broad, prominent chin. This appearance was distinct from that of unaffected family members and typical AGS patients. The glycine corresponding to position 274 is highly conserved in other epidermal growth factor-like domains of jagged1 and in those of other proteins. Its substitution in other proteins has been associated with mild or atypical variants of disease. These data support either a relative loss-of-function or a gain-of-function pathogenetic mechanism in this family and suggest that JAG1 mutations may contribute significantly to common variants of right heart obstructive disease.

Mouse models of genetic diseases resulting from mutations in elastic fiber proteins.

The inability to study appropriate human tissues at various stages of development has precluded the elaboration of a thorough understanding of the pathogenic mechanisms leading to diseases linked to mutations in genes for elastic fiber proteins. Recently, new insights have been gained by studying mice harboring targeted mutations in the genes that encode fibrillin-1 and elastin. These genes have been linked to Marfan syndrome (MFS) and supravalvular aortic stenosis (SVAS), respectively. For fibrillin-1, mouse models have revealed that phenotype is determined by the degree of functional impairment. The haploinsufficiency state or the expression of low levels of a product with dominant-negative potential from one allele is associated with mild phenotypes with a predominance of skeletal features. Exuberant expression of a dominant-negative-acting protein leads to the more severe MFS phenotype. Mice harboring targeted deletion of the elastin gene (ELN) show many of the features of SVAS in humans, including abnormalities in the vascular wall and altered hemodynamics associated with changes in wall compliance. The genetically altered mice suggest that SVAS is predominantly a disease of haploinsufficiency. These studies have underscored the prominent role of the elastic matrix in the morphogenesis and homeostasis of the vessel wall.

Novel Upf2p orthologues suggest a functional link between translation initiation and nonsense surveillance complexes.

Transcripts harboring premature signals for translation termination are recognized and rapidly degraded by eukaryotic cells through a pathway known as nonsense-mediated mRNA decay (NMD). In addition to protecting cells by preventing the translation of potentially deleterious truncated peptides, studies have suggested that NMD plays a broader role in the regulation of the steady-state levels of physiologic transcripts. In Saccharomyces cerevisiae, three trans-acting factors (Upf1p to Upf3p) are required for NMD. Orthologues of Upf1p have been identified in numerous species, showing that the NMD machinery, at least in part, is conserved through evolution. In this study, we demonstrate additional functional conservation of the NMD pathway through the identification of Upf2p homologues in Schizosaccharomyces pombe and humans (rent2). Disruption of S. pombe UPF2 established that this gene is required for NMD in fission yeast. rent2 was demonstrated to interact directly with rent1, a known trans-effector of NMD in mammalian cells. Additionally, fragments of rent2 were shown to possess nuclear targeting activity, although the native protein localizes to the cytoplasmic compartment. Finally, novel functional domains of Upf2p and rent2 with homology to eukaryotic initiation factor 4G (eIF4G) and other translational regulatory proteins were identified. Directed mutations within these so-called eIF4G homology (4GH) domains were sufficient to abolish the function of S. pombe Upf2p. Furthermore, using the two-hybrid system, we obtained evidence for direct interaction between rent2 and human eIF4AI and Sui1, both components of the translation initiation complex. Based on these findings, a novel model in which Upf2p and rent2 effects decreased translation and accelerated decay of nonsense transcripts through competitive interactions with eIF4G-binding partners is proposed.