Molecular and clinical analysis ofRAF1in Noonan syndrome and related disorders: dephosphorylation of serine 259 as the essential mechanism for mutant activation

Abstract

Noonan syndrome (NS) and related disorders are autosomal dominant disorders characterized by heart defects, facial dysmorphism, ectodermal abnormalities, and mental retardation. The dysregulation of the RAS/MAPK pathway appears to be a common molecular pathogenesis of these disorders: mutations in PTPN11, KRAS, and SOS1 have been identified in patients with NS, those in KRAS, BRAF, MAP2K1, and MAP2K2 in patients with CFC syndrome, and those in HRAS mutations in Costello syndrome patients. Recently, mutations in RAF1 have been also identified in patients with NS and two patients with LEOPARD (multiple lentigines, electrocardiographic conduction abnormalities, ocular hypertelorism, pulmonary stenosis, abnormal genitalia, retardation of growth, and sensorineural deafness) syndrome. In the current study, we identified eight RAF1 mutations in 18 of 119 patients with NS and related conditions without mutations in known genes. We summarized clinical manifestations in patients with RAF1 mutations as well as those in NS patients withPTPN11, SOS1, or KRAS mutations previously reported. Hypertrophic cardiomyopathy and short stature were found to be more frequently observed in patients with RAF1 mutations. Mutations in RAF1 were clustered in the conserved region 2 (CR2) domain, which carries an inhibitory phosphorylation site (serine at position 259; S259). Functional studies revealed that the RAF1 mutants located in the CR2 domain resulted in the decreased phosphorylation of S259, and that mutant RAF1 then dissociated from 14-3-3, leading to a partial ERK activation. Our results suggest that the dephosphorylation of S259 is the primary pathogenic mechanism in the activation of RAF1 mutants located in the CR2 domain as well as of downstream ERK.

Growth hormone and noonan syndrome: update in dysfunctional signaling aspects and in therapy for short stature

“…In this study, the authors also observed that individuals with BRAF mutations, similarly to those mutated in SOS1, have better preserved postnatal growth compared with patients harboring other genotypes [14]. In contrast, short stature seems to be more frequently associated with mutations of RAF1 (mean height SDS of -0.5±0.9 SDS compared to NS standards) or SHOC2 (-1.3±0.7) [14,20,21]. Nonetheless, it must be emphasized that these reports did not take into account a large number of patients, notably in the case of subjects with rare genotypes (e.g., SHOC2 mutations).…”

Clinical, pathological, and molecular analyses of cardiovascular abnormalities in Costello syndrome: A Ras/MAPK pathway syndrome

“…Among Costello syndrome patients with a cardiovascular problem, the frequent occurrence of arrhythmia (65% this study, 45% in literature review) is striking. Only patients with a RAF1 mutation approach this in overall frequency (13% among all patients), or with any atrial tachycardia (31%), one of whom had EAT [Kobayashi et al, 2010]. The finding of an abnormal electrocardiogram with conduction abnormalities in Noonan syndrome with multiple lentigines syndrome is well-described (45% reporting patients) and is not compared to patients with non-reentrant atrial tachycardia.…”

“…suggests that severe subaortic obstruction requiring surgical treatment is more frequent in Costello syndrome than in Noonan or CFC. HCM due to RAF1 mutations [Kobayashi et al, 2010] and in Noonan syndrome with multiple lentigines [Limongelli et al, 2007] is often severe, and longitudinal studies are needed to delineate a comparison to Costello syndrome. The resolution by echocardiographic criteria of HCM in five patients with Costello syndrome is not unique among RASopathies, and requires monitoring across the spectrum of RASopathies in longitudinal studies.…”

Signaling to Cardiac Hypertrophy: Insights from Human and Mouse RASopathies

“…RAS activation results in the stimulation of RAF family members, which have been implicated in the development of hypertrophy (56,57) and of HCM in RASopathies (9,13,14). RAF1 mutations have been more frequently found to be associated with HCM, and nearly all NS patients with RAF1 mutations exhibit HCM (58). Like NS patients, mice heterozygous for the NS-associated RAF1 mutation exhibit eccentric cardiac hypertrophy (59).…”