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Gain-of-function RAF1 mutations cause Noonan and LEOPARD syndromes with hypertrophic cardiomyopathy


Noonan and LEOPARD syndromes are developmental disorders with overlapping features, including cardiac abnormalities, short stature and facial dysmorphia. Increased RAS signaling owing to PTPN11, SOS1 and KRAS mutations causes 60% of Noonan syndrome cases1,2,3,4,5,6, and PTPN11 mutations cause 90% of LEOPARD syndrome cases7. Here, we report that 18 of 231 individuals with Noonan syndrome without known mutations (corresponding to 3% of all affected individuals) and two of six individuals with LEOPARD syndrome without PTPN11 mutations have missense mutations in RAF1, which encodes a serine-threonine kinase that activates MEK1 and MEK2. Most mutations altered a motif flanking Ser259, a residue critical for autoinhibition of RAF1 through 14-3-3 binding. Of 19 subjects with a RAF1 mutation in two hotspots, 18 (or 95%) showed hypertrophic cardiomyopathy (HCM), compared with the 18% prevalence of HCM among individuals with Noonan syndrome in general. Ectopically expressed RAF1 mutants from the two HCM hotspots had increased kinase activity and enhanced ERK activation, whereas non–HCM-associated mutants were kinase impaired. Our findings further implicate increased RAS signaling in pathological cardiomyocyte hypertrophy.

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Figure 1: RAF1 domain structure and location of residues altered in Noonan and LEOPARD syndromes.
Figure 2: RAF1 kinase assays.
Figure 3: ERK activation assays.
Figure 4: 14-3-3 binding and phosphorylation status of Ser259 of RAF1.


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We are indebted to the affected individuals and families who participated in the study, the physicians who referred the subjects and the Joint Genome Institute's production sequencing group. We thank D.K. Morrison (National Cancer Institute, US National Institutes of Health) for critical reagents and suggestions. We also thank G. Crisponi (Clinica Sant'Anna, Cagliari, Italy), F. Faravelli (Ospedale Galliera, Genova, Italy), L. Memo (ULLS9, Treviso, Italy) and F. Stanzial (Regional Hospital, Bolzano, Italy) for their valuable clinical assistance. This work was supported by Telethon-Italy grant GGP07115 (M.T.) and Programma di Collaborazione Italia-USA/malattie rare 2007 grants (M.T. and A.S.); US National Institutes of Health Grants HL71207, HD01294, HL074728 (B.D.G.), HD042569 (M.J.A.) and DK57683 (P.M.); the American Heart Association (M.J.A.); the Dr. Scholl Foundation (M.J.A.); the CJ Foundation for SIDS and the Mayo Foundation (M.J.A.) and Italian Ministry of Health Grant RC 2006-2007, Italian Ministry of University and Research C26A06KTTN and RBIP06PMF2_005 (B.D.). Research conducted at the E.O. Lawrence Berkeley National Laboratory and the Joint Genome Institute was performed under the Berkeley Program for Genomic Applications, funded by the US National Heart, Lung and Blood Institute (HL066681) and Department of Energy Contract DE-AC02-05CH11231 (University of California). We regret our inability to cite certain references describing work similar to references that were cited due to the limits of this journal.

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Correspondence to Marco Tartaglia or Bruce D Gelb.

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Pandit, B., Sarkozy, A., Pennacchio, L. et al. Gain-of-function RAF1 mutations cause Noonan and LEOPARD syndromes with hypertrophic cardiomyopathy. Nat Genet 39, 1007–1012 (2007).

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