* 164760

RAF1 PROTOONCOGENE, SERINE/THREONINE KINASE ; RAF1


Alternative titles; symbols

V-RAF-1 MURINE LEUKEMIA VIRAL ONCOGENE HOMOLOG 1
ONCOGENE RAF1
TRANSFORMING REPLICATION-DEFECTIVE MURINE RETROVIRUS 3611-MSV
ONCOGENE MIL
CRAF


Other entities represented in this entry:

RAF1/SRGAP3 FUSION GENE, INCLUDED

HGNC Approved Gene Symbol: RAF1

Cytogenetic location: 3p25.2     Genomic coordinates (GRCh38): 3:12,583,601-12,664,117 (from NCBI)


Gene-Phenotype Relationships
Location Phenotype Phenotype
MIM number
Inheritance Phenotype
mapping key
3p25.2 Cardiomyopathy, dilated, 1NN 615916 AD 3
LEOPARD syndrome 2 611554 AD 3
Noonan syndrome 5 611553 AD 3

TEXT

Cloning and Expression

Rapp et al. (1983) cloned a unique acutely transforming replication-defective mouse type C virus and characterized its acquired oncogene, called v-raf. The viral genome bears close similarities to the Moloney murine leukemia virus (see MOS, 190060). The cellular homolog, c-raf, is present in 1 or 2 copies per haploid genome in mouse and human DNA. The MIL oncogene, a second oncogene in the avian retrovirus MH2, which contains the MYC oncogene, is the avian equivalent of the murine RAF oncogene, i.e., they are identical.


Mapping

Bonner et al. (1984) assigned the RAF1 gene to chromosome 3p25 by in situ hybridization. This suggested that RAF1 may be involved in mixed parotid gland tumors with the t(3;8)(p25;q21) translocation (Mark et al. (1980, 1982)).

Tory et al. (1992) constructed a genetic linkage map of 96 loci on 3p, extending from the terminal band to the centromere. Multipoint linkage analysis indicated that the male, female, and sex-averaged maps extend for 102, 147, and 116 cM, respectively. RAF1 and 16 DNA markers were localized by fluorescence in situ hybridization. RAF1 was regionalized to 3p25.

Pseudogenes

Bonner et al. (1984) showed that RAF2, a processed pseudogene, is on chromosome 4.

Hiroshige et al. (1986) assigned the RAF2 pseudogene to region 4pter-p15 by the study of hybrid cells containing various chromosome 4 regions.


Gene Function

The function of RAF1 was reviewed by Li et al. (1991).

Wang et al. (1996) showed that RAF1 can be targeted to the mitochondria by BCL2 (151430), a regulator of apoptotic cell death. Active RAF1 improved BCL2-mediated resistance to apoptosis. They also showed that RAF1 phosphorylates BAD (603167).

Alavi et al. (2003) showed that basic fibroblast growth factor (FGFB; 134920) and vascular endothelial growth factor (VEGF; 192240) differentially activate Raf1, resulting in protection from distinct pathways of apoptosis in human endothelial cells and chick embryo vasculature. BFGF activated Raf1 via p21-activated protein kinase-1 (PAK1; 602590) phosphorylation of serines 338 and 339, resulting in Raf1 mitochondrial translocation and endothelial cell protection from the intrinsic pathway of apoptosis, independent of the mitogen-activated protein kinase kinase-1 (MEK1; 176872). In contrast, VEGF activated Raf1 via Src kinase (CSK; 124095), leading to phosphorylation of tyrosines 340 and 341 and MEK1-dependent protection from extrinsic-mediated apoptosis. Alavi et al. (2003) concluded that RAF1 may be a pivotal regulator of endothelial cell survival during angiogenesis.

Lorenz et al. (2003) demonstrated that the RAF kinase inhibitor protein (RKIP; 604591) is a physiologic inhibitor of GRK2 (109635). After stimulation of G protein-coupled receptors, RKIP dissociates from its known target, RAF1, to associate with GRK2 and block its activity. This switch is triggered by a protein kinase C (PKC; see 176960)-dependent phosphorylation of RKIP on serine-153. Lorenz et al. (2003) concluded that their data delineate a new principle in signal transduction: by activating PKC, the incoming receptor signal is enhanced both by removing an inhibitor from RAF1 and by blocking receptor internalization. A physiologic role for this mechanism is shown in cardiomyocytes in which the downregulation of RKIP restrains beta-adrenergic signaling and contractile activity.

O'Neill et al. (2004) used proteomic analysis of RAF1 signaling complexes to show that RAF1 counteracts apoptosis by suppressing the activation of mammalian sterile 20-like kinase (MST2; 605030). RAF1 prevents dimerization and phosphorylation of the activation loop of MST2 independently of its protein kinase activity. Depletion of MST2 from Raf1-null mouse or human cells abrogated sensitivity to apoptosis, whereas overexpression of MST2 induced apoptosis. Conversely, depletion of Raf1 from Raf1 +/+ mouse or human cells led to MST2 activation and apoptosis. The concomitant depletion of both RAF1 and MST2 prevented apoptosis.

By creating a kinase-defective version of Raf1 in mice or by using a Raf1 inhibitor, Noble et al. (2008) showed that Raf1 autophosphorylation on ser621 prevented its degradation by the proteasome.

Using Drosophila Schneider S2 cells, Rajakulendran et al. (2009) demonstrated that RAF catalytic function is regulated in response to a specific mode of dimerization of its kinase domain, which they termed the side-to-side dimer. Moreover, they found that the RAF-related pseudokinase KSR (601132) also participates in forming side-to-side heterodimers with RAF and can thereby trigger RAF activation. This mechanism provides an elegant explanation for the longstanding conundrum about RAF catalytic activation, and also provides an explanation for the capacity of KSR, despite lacking catalytic function, to directly mediate RAF activation.

Hollander et al. (2010) found that microRNA-212 (MIR212; 613487) was upregulated in the dorsal striatum of rats with a history of extended access to cocaine. Striatal miR212 decreased responsiveness to the motivational properties of cocaine by markedly amplifying the stimulatory effects of the drug on Creb (123810) signaling. Studies in rats and HEK cells showed that amplification of CREB signaling occurred through miR212-enhanced RAF1 activity, resulting in adenylyl cyclase sensitization and increased expression of the essential Creb coactivator TORC (see CRTC1; 607536). miR212 activated RAF1, at least in part, through repression of SPRED1 (609291). Hollander et al. (2010) concluded that striatal miR212 signaling has a key role in determining vulnerability to cocaine addiction.

Using immunoprecipitation of endogenous LZTR1 (600574) followed by Western blotting, Umeki et al. (2019) showed that LZTR1 bound to the RAF1-SHOC2 (602775)-PPP1CB (600590) complex. Mutations in all these genes cause Noonan syndrome or Noonan-like phenotypes. Cells transfected with siRNA against LZTR1 exhibited decreased levels of RAF1 phosphorylated at ser259.

Oncogenic Function

Shimizu et al. (1985) identified the activated RAF gene in the stomach cancer of a Japanese patient. Stomach cancer is the most common cancer in Japan. Fukui et al. (1985) found that transforming DNA in a human glioblastoma line was apparently the RAF gene. Teyssier et al. (1986) presented evidence for a relationship of RAF1 to renal cell carcinoma (144700).

Kasid et al. (1987) transfected tumor cell DNA into NIH/3T3 cells to demonstrate that a radiation-resistant laryngeal carcinoma cell line contained altered RAF1 sequences. The karyotype of the tumor cells showed absence of chromosome 3, and transformed cells had double-minute chromosomes.

Poulikakos et al. (2010) used chemical genetic methods to show that drug-mediated transactivation of RAF dimers is responsible for the paradoxical activation of the enzyme by inhibitors. Induction of ERK signaling requires direct binding of the drug to the ATP-binding site of one kinase of the dimer and is dependent on RAS activity. Drug binding to one member of RAF homodimers (CRAF-CRAF) or heterodimers (CRAF-BRAF) inhibits one promoter, but results in transactivation of the drug-free protomer. In BRAF(V600E) (164757.0001) tumors, RAS is not activated, thus transactivation is minimal and ERK signaling is inhibited in cells exposed to RAF inhibitors. These results indicated that RAF inhibitors will be effective in tumors in which BRAF is mutated. Furthermore, because RAF inhibitors do not inhibit ERK signaling in other cells, the model predicted that they would have a higher therapeutic index and greater antitumor activity than mitogen-activated protein kinase kinase (MEK) inhibitors, but could also cause toxicity due to the MEK/ERK activation. Poulikakos et al. (2010) noted that these predictions were borne out in a clinical trial of the RAF inhibitor PLX4032, as reported by Chapman et al. (2009) and Flaherty et al. (2009). The model indicated that promotion of RAF dimerization by elevation of wildtype RAF expression or RAS activity could lead to drug resistance in mutant BRAF tumors. In agreement with this prediction, RAF inhibitors do not inhibit ERK signaling in cells that coexpress BRAF(V600E) and mutant RAS.

Hatzivassiliou et al. (2010) demonstrated that ATP-competitive RAF inhibitors have 2 opposing mechanisms of action depending on the cellular context. In BRAF(V600E) tumors, RAF inhibitors effectively block the mitogen-activated protein kinase (MAPK) signaling pathway and decrease tumor growth. Notably, in KRAS mutant and RAS/RAF wildtype tumors, RAF inhibitors activate the RAF-MEK-ERK pathway in a RAS-dependent manner, thus enhancing tumor growth in some xenograft models. Inhibitor binding activates wildtype RAF isoforms by inducing dimerization, membrane localization, and interaction with RAS-GTP. These events occur independently of kinase inhibition and are, instead, linked to direct conformational effects of inhibitors on the RAF kinase domain. On the basis of these findings, Hatzivassiliou et al. (2010) demonstrated that ATP-competitive kinase inhibitors can have opposing functions as inhibitors or activators of signaling pathways, depending on the cellular context. The authors stated that their work provided new insights into the therapeutic use of ATP-competitive RAF inhibitors.


Molecular Genetics

Noonan Syndrome 5 or LEOPARD syndrome 2

Pandit et al. (2007) analyzed the RAF1 gene in 231 individuals with Noonan syndrome who did not have mutations in the PTPN11 (176876), KRAS (190070), or SOS1 (182530) genes, and in 6 persons with LEOPARD syndrome who did not have mutations in PTPN11. They identified 13 different missense mutations (see, e.g., 164760.0001-164760.0003) in 18 unrelated patients with NS (NS5; 611553) and 2 missense mutations (164760.0001 and 164760.0004) in 2 patients with LEOPARD syndrome (LPRD2; 611554), respectively. Most mutations altered a motif flanking ser259 located in the CR2 domain, critical for autoinhibition of RAF1 through 14-3-3 (see 113508) binding. Of 17 NS patients with a RAF1 mutation in either of 2 hotspots (clustering around ser259 or ser612), 16 (94%) had hypertrophic cardiomyopathy (CMH; see 192600), compared with an 18% prevalence of CMH among NS patients in general. Pandit et al. (2007) also scanned RAF1 exons mutated in NS and LEOPARD patients in 241 unrelated individuals with nonsyndromic CMH who did not have mutations in 8 myofilament genes known to cause CMH, and the authors identified a thr260-to-ile mutation in the RAF1 gene in 1 patient. Ectopically expressed RAF1 mutants from the 2 CMH hotspots had increased kinase activity and enhanced ERK (see 176948) activation, whereas non-CMH-associated mutants were kinase impaired.

Razzaque et al. (2007) analyzed the RAF1 gene in 30 individuals clinically diagnosed with Noonan syndrome who were negative for mutations in the PTPN11, KRAS, HRAS (190020), or SOS1 genes, and identified 5 different missense mutations in RAF1 in 10 (33%) individuals. The authors noted that 8 of the 10 mutation-positive patients with 1 of 4 mutations causing changes in the CR2 domain of RAF1 (see, e.g., 164760.0001 and 164760.0003) had hypertrophic cardiomyopathy, whereas the 2 affected individuals with a mutation leading to changes in the CR3 domain did not (164760.0004). Transfection studies in HEK293 cells demonstrated that all 5 mutations in RAF1 behaved as gain-of-function mutants with increased kinase and ERK activation compared with wildtype RAF1.

Dilated Cardiomyopathy 1NN

In 8 patients from South and North India and 2 patients from Japan with nonsyndromic dilated cardiomyopathy (CMD1NN; 615916), Dhandapany et al. (2014) identified heterozygous mutations in the RAF1 gene (see, e.g., 164760.0005-164760.0007) that segregated with disease in the families and were not found in ancestry-matched controls. Biochemical studies showed that the CMD-associated RAF1 mutants had altered kinase activity, resulting in largely unchanged ERK (see 601795) activation but in AKT (164730) that was hyperactivated in a BRAF (164757)-dependent manner. Constitutive expression of these mutants in zebrafish embryos resulted in a heart failure phenotype with AKT hyperactivation that was rescued by treatment with rapamycin.


Cytogenetics

Jones et al. (2009) found a tandem duplication at chromosome 3p25 in a pilocytic astrocytoma (see 137800) that resulted in fusion of exons 1 through 12 of the SRGAP3 gene (606525) to exons 10 through 17 of the RAF1 gene. The fusion transcript encodes a deduced protein containing the first 513 N-terminal amino acids of SRGAP3, including the FES (190030)/CIP4 (TRIP10; 604504) homology domain, fused to 318 C-terminal amino acids of RAF1, including the entire RAF1 kinase domain. The fusion protein showed higher activity than wildtype RAF1 in phosphorylation of endogenous Mek1 in mouse fibroblasts, and it conferred anchorage-independent cell growth.


Animal Model

To examine the in vivo role of RAF1 in the heart, Yamaguchi et al. (2004) generated cardiac muscle-specific Raf1 conditionally deleted mice. The mice demonstrated left ventricular systolic dysfunction and heart dilation without cardiac hypertrophy or lethality, and showed a significant increase in the number of apoptotic cardiomyocytes. The expression level and activation of MEK1/2 (see 176872 and 601263, respectively) and ERK (see 601795) showed no difference, but the kinase activity of apoptosis signal-regulating kinase-1 (ASK1; 602448), JNK (see 601158), and p38 (600289) increased significantly compared to that of controls. The ablation of ASK1 rescued heart dysfunction and dilatation as well as cardiac fibrosis. Yamaguchi et al. (2004) concluded that RAF1 promotes cardiomyocyte survival through a MEK/ERK-independent mechanism.


ALLELIC VARIANTS ( 7 Selected Examples):

.0001 NOONAN SYNDROME 5

LEOPARD SYNDROME 2, INCLUDED
RAF1, SER257LEU
  
RCV000014985...

In 7 unrelated patients with Noonan syndrome (NS5; 611553) and 1 patient with LEOPARD syndrome-2 (LPRD2; 611554), Pandit et al. (2007) identified heterozygosity for a 770C-T transition in exon 7 of the RAF1 gene, resulting in a ser257-to-leu (S257L) substitution at a conserved residue in the CR2 domain. All patients had hypertrophic cardiomyopathy (CMH), including a 3.6-year-old girl with CMH at birth and a 35-year-old woman with LEOPARD syndrome. Ectopically expressed S257L mutants demonstrated increased kinase activity and enhanced ERK (see 176948) activation.

Razzaque et al. (2007) identified the S257L mutation of the RAF1 gene in 4 unrelated patients with Noonan syndrome, 3 with obstructive and 1 with nonobstructive CMH. The mutation was not found in 100 control individuals or in 100 patients with CMH without Noonan syndrome.


.0002 NOONAN SYNDROME 5

RAF1, PRO261SER
  
RCV000014987...

In 5 affected individuals of 2 unrelated families with Noonan syndrome (NS5; 611553), Pandit et al. (2007) identified heterozygosity for a 781C-T transition in exon 7 of the RAF1 gene, resulting in a pro261-to-ser (P261S) substitution at a conserved residue in the CR2 domain. Four of the 5 patients had hypertrophic cardiomyopathy (CMH); the 1 individual with a P261S change but without CMH was a 6-year-old girl whose 38-year-old mother had the same mutation and had been diagnosed with CMH at 23 years of age. The mutation was not found in 210 control individuals.

Razzaque et al. (2007) identified the P261S mutation in 3 Noonan syndrome patients, a 1-year-old boy and his 33-year-old father and an unrelated 16-year-old boy. All 3 displayed CMH. The mutation was not found in 100 control individuals or in 100 individuals with CMH without Noonan syndrome. Transfection studies in HEK293 cells demonstrated that P261S behaved as a gain-of-function mutant with increased kinase and ERK (see 176948) activation compared with wildtype RAF1.


.0003 NOONAN SYNDROME 5

RAF1, THR491ARG
  
RCV000014988...

In a sister and brother with Noonan syndrome (NS5; 611553), Pandit et al. (2007) identified heterozygosity for a 1472C-G transversion in exon 14 of the RAF1 gene, resulting in a thr491-to-arg (T491R) substitution in the CR3 domain. Neither sib had hypertrophic cardiomyopathy. The mutation was not found in 210 control individuals.


.0004 LEOPARD SYNDROME 2

NOONAN SYNDROME 5, INCLUDED
RAF1, LEU613VAL
  
RCV000014989...

In a 43-year-old woman with LEOPARD syndrome-2 (LPRD2; 611554), Pandit et al. (2007) identified an 1837C-G transversion in exon 17 of the RAF1 gene, resulting in a leu613-to-val (L613V) substitution at a conserved residue in the C terminus. The patient had hypertrophic cardiomyopathy.

Razzaque et al. (2007) identified the L613V mutation, which they designated as being located in the CR3 domain of RAF1, in 2 unrelated boys with Noonan syndrome (NS5; 611553), neither of whom had hypertrophic cardiomyopathy. The mutation was not found in 100 control individuals or in 100 patients with hypertrophic cardiomyopathy without Noonan syndrome. Transfection studies in HEK293 cells demonstrated that L613V behaved as a gain-of-function mutant with increased kinase and ERK (see 176948) activation compared with wildtype RAF1.


.0005 CARDIOMYOPATHY, DILATED, 1NN

RAF1, LEU603PRO
  
RCV000131334

In 3 patients with nonsyndromic dilated cardiomyopathy (CMD1NN; 615916), a 40-year-old South Indian mother and her 4-year-old son and an unrelated 21-year-old South Indian woman, Dhandapany et al. (2014) identified heterozygosity for a c.1808T-C transition in the RAF1 gene, resulting in a leu603-to-pro (L603P) substitution at a highly conserved residue in the CR3 domain. Age at onset of disease in the 3 patients was 24 years, 3 years, and 10 years, respectively. The mutation was not found in the 4-year-old boy's unaffected father, in 500 ancestry-matched South Indian controls, or in 13,600 European and African American alleles in the Exome Sequencing Project database. Functional analysis in HEK293 cells showed impaired kinase activity and reduced ERK (see 601795) activation with the L603P mutant. Constitutive expression of the L603P mutant in zebrafish embryos resulted in a heart failure phenotype, including elongated ventricular and atrial chambers, profound pericardial edema, blood congestion at the cardiac inflow tract, and impaired cardiac contractions. Immunoblotting showed AKT hyperactivation, and the heart defects were partially rescued by treatment with rapamycin, an AKT-mTOR (601231) inhibitor.


.0006 CARDIOMYOPATHY, DILATED, 1NN

RAF1, THR641MET
  
RCV000131336...

In a 15-year-old South Indian girl and an unrelated 21-year-old North Indian man with nonsyndromic dilated cardiomyopathy (CMD1NN; 615916), Dhandapany et al. (2014) identified heterozygosity for a c.1922T-C transition in the RAF1 gene, resulting in a thr641-to-met (T641M) substitution at a highly conserved residue. Age at onset of disease in the 2 patients was 7 years and 16 years, respectively. The mutation, which segregated with disease in the available family, was not found in 500 ancestry-matched South Indian controls or in 350 ancestry-matched North Indian controls. Functional analysis in HEK293 cells showed a mild increase in kinase activity with the T641M mutant that was less augmented than that of the CMH-associated RAF1 mutants tested, L613V (164760.0004) and S257L (164760.0001). ERK (see 601795) activation with the T641M mutant was similar to that observed with wildtype RAF1 and was significantly less than with the CMH-associated mutants.


.0007 CARDIOMYOPATHY, DILATED, 1NN

RAF1, ALA237THR
  
RCV000131337...

In a 44-year-old Japanese man who had onset of nonsyndromic dilated cardiomyopathy (CMD1NN; 615916) at 40 years of age, Dhandapany et al. (2014) identified heterozygosity for a c.709G-A transition in the RAF1 gene, resulting in an ala237-to-thr (A237T) substitution at a highly conserved residue. The mutation was not found in 300 Japanese controls. Functional analysis in HEK293 cells showed a mild increase in kinase activity with the A237T mutant that was less augmented than that of the CMH-associated RAF1 mutants tested, L613V (164760.0004) and S257L (164760.0001). ERK (see 601795) activation with the A237T mutant was similar to that observed with wildtype RAF1 and was significantly less than with the CMH-associated mutants, whereas activation of AKT and tuberin (TSC2; 191092) was excessive compared to CMH-associated mutants or wildtype RAF1.


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Ada Hamosh - updated : 01/10/2019
Marla J. F. O'Neill - updated : 7/30/2014
Ada Hamosh - updated : 8/24/2010
Patricia A. Hartz - updated : 5/19/2010
Ada Hamosh - updated : 4/15/2010
Ada Hamosh - updated : 10/19/2009
Patricia A. Hartz - updated : 5/29/2009
Marla J. F. O'Neill - updated : 10/24/2007
Ada Hamosh - updated : 1/14/2005
Marla J. F. O'Neill - updated : 12/2/2004
Ada Hamosh - updated : 12/30/2003
Ada Hamosh - updated : 7/24/2003
Jennifer P. Macke - updated : 10/20/1998
Creation Date:
Victor A. McKusick : 6/2/1986
carol : 12/30/2019
alopez : 05/28/2019
alopez : 01/10/2019
carol : 10/20/2016
carol : 08/23/2016
carol : 11/14/2014
carol : 8/1/2014
mcolton : 7/30/2014
mgross : 8/25/2010
terry : 8/24/2010
carol : 7/16/2010
mgross : 5/20/2010
mgross : 5/20/2010
terry : 5/19/2010
alopez : 4/20/2010
terry : 4/15/2010
alopez : 10/26/2009
alopez : 10/26/2009
terry : 10/19/2009
mgross : 6/2/2009
terry : 5/29/2009
wwang : 10/25/2007
wwang : 10/25/2007
terry : 10/24/2007
alopez : 1/20/2005
alopez : 1/20/2005
terry : 1/14/2005
carol : 12/2/2004
mgross : 4/13/2004
mgross : 3/17/2004
alopez : 12/31/2003
terry : 12/30/2003
carol : 7/24/2003
terry : 7/24/2003
alopez : 10/20/1998
mark : 6/4/1996
carol : 4/7/1993
carol : 6/1/1992
supermim : 3/16/1992
carol : 3/2/1992
carol : 2/20/1991
supermim : 3/20/1990

* 164760

RAF1 PROTOONCOGENE, SERINE/THREONINE KINASE ; RAF1


Alternative titles; symbols

V-RAF-1 MURINE LEUKEMIA VIRAL ONCOGENE HOMOLOG 1
ONCOGENE RAF1
TRANSFORMING REPLICATION-DEFECTIVE MURINE RETROVIRUS 3611-MSV
ONCOGENE MIL
CRAF


Other entities represented in this entry:

RAF1/SRGAP3 FUSION GENE, INCLUDED

HGNC Approved Gene Symbol: RAF1

Cytogenetic location: 3p25.2     Genomic coordinates (GRCh38): 3:12,583,601-12,664,117 (from NCBI)


Gene-Phenotype Relationships

Location Phenotype Phenotype
MIM number
Inheritance Phenotype
mapping key
3p25.2 Cardiomyopathy, dilated, 1NN 615916 Autosomal dominant 3
LEOPARD syndrome 2 611554 Autosomal dominant 3
Noonan syndrome 5 611553 Autosomal dominant 3

TEXT

Cloning and Expression

Rapp et al. (1983) cloned a unique acutely transforming replication-defective mouse type C virus and characterized its acquired oncogene, called v-raf. The viral genome bears close similarities to the Moloney murine leukemia virus (see MOS, 190060). The cellular homolog, c-raf, is present in 1 or 2 copies per haploid genome in mouse and human DNA. The MIL oncogene, a second oncogene in the avian retrovirus MH2, which contains the MYC oncogene, is the avian equivalent of the murine RAF oncogene, i.e., they are identical.


Mapping

Bonner et al. (1984) assigned the RAF1 gene to chromosome 3p25 by in situ hybridization. This suggested that RAF1 may be involved in mixed parotid gland tumors with the t(3;8)(p25;q21) translocation (Mark et al. (1980, 1982)).

Tory et al. (1992) constructed a genetic linkage map of 96 loci on 3p, extending from the terminal band to the centromere. Multipoint linkage analysis indicated that the male, female, and sex-averaged maps extend for 102, 147, and 116 cM, respectively. RAF1 and 16 DNA markers were localized by fluorescence in situ hybridization. RAF1 was regionalized to 3p25.

Pseudogenes

Bonner et al. (1984) showed that RAF2, a processed pseudogene, is on chromosome 4.

Hiroshige et al. (1986) assigned the RAF2 pseudogene to region 4pter-p15 by the study of hybrid cells containing various chromosome 4 regions.


Gene Function

The function of RAF1 was reviewed by Li et al. (1991).

Wang et al. (1996) showed that RAF1 can be targeted to the mitochondria by BCL2 (151430), a regulator of apoptotic cell death. Active RAF1 improved BCL2-mediated resistance to apoptosis. They also showed that RAF1 phosphorylates BAD (603167).

Alavi et al. (2003) showed that basic fibroblast growth factor (FGFB; 134920) and vascular endothelial growth factor (VEGF; 192240) differentially activate Raf1, resulting in protection from distinct pathways of apoptosis in human endothelial cells and chick embryo vasculature. BFGF activated Raf1 via p21-activated protein kinase-1 (PAK1; 602590) phosphorylation of serines 338 and 339, resulting in Raf1 mitochondrial translocation and endothelial cell protection from the intrinsic pathway of apoptosis, independent of the mitogen-activated protein kinase kinase-1 (MEK1; 176872). In contrast, VEGF activated Raf1 via Src kinase (CSK; 124095), leading to phosphorylation of tyrosines 340 and 341 and MEK1-dependent protection from extrinsic-mediated apoptosis. Alavi et al. (2003) concluded that RAF1 may be a pivotal regulator of endothelial cell survival during angiogenesis.

Lorenz et al. (2003) demonstrated that the RAF kinase inhibitor protein (RKIP; 604591) is a physiologic inhibitor of GRK2 (109635). After stimulation of G protein-coupled receptors, RKIP dissociates from its known target, RAF1, to associate with GRK2 and block its activity. This switch is triggered by a protein kinase C (PKC; see 176960)-dependent phosphorylation of RKIP on serine-153. Lorenz et al. (2003) concluded that their data delineate a new principle in signal transduction: by activating PKC, the incoming receptor signal is enhanced both by removing an inhibitor from RAF1 and by blocking receptor internalization. A physiologic role for this mechanism is shown in cardiomyocytes in which the downregulation of RKIP restrains beta-adrenergic signaling and contractile activity.

O'Neill et al. (2004) used proteomic analysis of RAF1 signaling complexes to show that RAF1 counteracts apoptosis by suppressing the activation of mammalian sterile 20-like kinase (MST2; 605030). RAF1 prevents dimerization and phosphorylation of the activation loop of MST2 independently of its protein kinase activity. Depletion of MST2 from Raf1-null mouse or human cells abrogated sensitivity to apoptosis, whereas overexpression of MST2 induced apoptosis. Conversely, depletion of Raf1 from Raf1 +/+ mouse or human cells led to MST2 activation and apoptosis. The concomitant depletion of both RAF1 and MST2 prevented apoptosis.

By creating a kinase-defective version of Raf1 in mice or by using a Raf1 inhibitor, Noble et al. (2008) showed that Raf1 autophosphorylation on ser621 prevented its degradation by the proteasome.

Using Drosophila Schneider S2 cells, Rajakulendran et al. (2009) demonstrated that RAF catalytic function is regulated in response to a specific mode of dimerization of its kinase domain, which they termed the side-to-side dimer. Moreover, they found that the RAF-related pseudokinase KSR (601132) also participates in forming side-to-side heterodimers with RAF and can thereby trigger RAF activation. This mechanism provides an elegant explanation for the longstanding conundrum about RAF catalytic activation, and also provides an explanation for the capacity of KSR, despite lacking catalytic function, to directly mediate RAF activation.

Hollander et al. (2010) found that microRNA-212 (MIR212; 613487) was upregulated in the dorsal striatum of rats with a history of extended access to cocaine. Striatal miR212 decreased responsiveness to the motivational properties of cocaine by markedly amplifying the stimulatory effects of the drug on Creb (123810) signaling. Studies in rats and HEK cells showed that amplification of CREB signaling occurred through miR212-enhanced RAF1 activity, resulting in adenylyl cyclase sensitization and increased expression of the essential Creb coactivator TORC (see CRTC1; 607536). miR212 activated RAF1, at least in part, through repression of SPRED1 (609291). Hollander et al. (2010) concluded that striatal miR212 signaling has a key role in determining vulnerability to cocaine addiction.

Using immunoprecipitation of endogenous LZTR1 (600574) followed by Western blotting, Umeki et al. (2019) showed that LZTR1 bound to the RAF1-SHOC2 (602775)-PPP1CB (600590) complex. Mutations in all these genes cause Noonan syndrome or Noonan-like phenotypes. Cells transfected with siRNA against LZTR1 exhibited decreased levels of RAF1 phosphorylated at ser259.

Oncogenic Function

Shimizu et al. (1985) identified the activated RAF gene in the stomach cancer of a Japanese patient. Stomach cancer is the most common cancer in Japan. Fukui et al. (1985) found that transforming DNA in a human glioblastoma line was apparently the RAF gene. Teyssier et al. (1986) presented evidence for a relationship of RAF1 to renal cell carcinoma (144700).

Kasid et al. (1987) transfected tumor cell DNA into NIH/3T3 cells to demonstrate that a radiation-resistant laryngeal carcinoma cell line contained altered RAF1 sequences. The karyotype of the tumor cells showed absence of chromosome 3, and transformed cells had double-minute chromosomes.

Poulikakos et al. (2010) used chemical genetic methods to show that drug-mediated transactivation of RAF dimers is responsible for the paradoxical activation of the enzyme by inhibitors. Induction of ERK signaling requires direct binding of the drug to the ATP-binding site of one kinase of the dimer and is dependent on RAS activity. Drug binding to one member of RAF homodimers (CRAF-CRAF) or heterodimers (CRAF-BRAF) inhibits one promoter, but results in transactivation of the drug-free protomer. In BRAF(V600E) (164757.0001) tumors, RAS is not activated, thus transactivation is minimal and ERK signaling is inhibited in cells exposed to RAF inhibitors. These results indicated that RAF inhibitors will be effective in tumors in which BRAF is mutated. Furthermore, because RAF inhibitors do not inhibit ERK signaling in other cells, the model predicted that they would have a higher therapeutic index and greater antitumor activity than mitogen-activated protein kinase kinase (MEK) inhibitors, but could also cause toxicity due to the MEK/ERK activation. Poulikakos et al. (2010) noted that these predictions were borne out in a clinical trial of the RAF inhibitor PLX4032, as reported by Chapman et al. (2009) and Flaherty et al. (2009). The model indicated that promotion of RAF dimerization by elevation of wildtype RAF expression or RAS activity could lead to drug resistance in mutant BRAF tumors. In agreement with this prediction, RAF inhibitors do not inhibit ERK signaling in cells that coexpress BRAF(V600E) and mutant RAS.

Hatzivassiliou et al. (2010) demonstrated that ATP-competitive RAF inhibitors have 2 opposing mechanisms of action depending on the cellular context. In BRAF(V600E) tumors, RAF inhibitors effectively block the mitogen-activated protein kinase (MAPK) signaling pathway and decrease tumor growth. Notably, in KRAS mutant and RAS/RAF wildtype tumors, RAF inhibitors activate the RAF-MEK-ERK pathway in a RAS-dependent manner, thus enhancing tumor growth in some xenograft models. Inhibitor binding activates wildtype RAF isoforms by inducing dimerization, membrane localization, and interaction with RAS-GTP. These events occur independently of kinase inhibition and are, instead, linked to direct conformational effects of inhibitors on the RAF kinase domain. On the basis of these findings, Hatzivassiliou et al. (2010) demonstrated that ATP-competitive kinase inhibitors can have opposing functions as inhibitors or activators of signaling pathways, depending on the cellular context. The authors stated that their work provided new insights into the therapeutic use of ATP-competitive RAF inhibitors.


Molecular Genetics

Noonan Syndrome 5 or LEOPARD syndrome 2

Pandit et al. (2007) analyzed the RAF1 gene in 231 individuals with Noonan syndrome who did not have mutations in the PTPN11 (176876), KRAS (190070), or SOS1 (182530) genes, and in 6 persons with LEOPARD syndrome who did not have mutations in PTPN11. They identified 13 different missense mutations (see, e.g., 164760.0001-164760.0003) in 18 unrelated patients with NS (NS5; 611553) and 2 missense mutations (164760.0001 and 164760.0004) in 2 patients with LEOPARD syndrome (LPRD2; 611554), respectively. Most mutations altered a motif flanking ser259 located in the CR2 domain, critical for autoinhibition of RAF1 through 14-3-3 (see 113508) binding. Of 17 NS patients with a RAF1 mutation in either of 2 hotspots (clustering around ser259 or ser612), 16 (94%) had hypertrophic cardiomyopathy (CMH; see 192600), compared with an 18% prevalence of CMH among NS patients in general. Pandit et al. (2007) also scanned RAF1 exons mutated in NS and LEOPARD patients in 241 unrelated individuals with nonsyndromic CMH who did not have mutations in 8 myofilament genes known to cause CMH, and the authors identified a thr260-to-ile mutation in the RAF1 gene in 1 patient. Ectopically expressed RAF1 mutants from the 2 CMH hotspots had increased kinase activity and enhanced ERK (see 176948) activation, whereas non-CMH-associated mutants were kinase impaired.

Razzaque et al. (2007) analyzed the RAF1 gene in 30 individuals clinically diagnosed with Noonan syndrome who were negative for mutations in the PTPN11, KRAS, HRAS (190020), or SOS1 genes, and identified 5 different missense mutations in RAF1 in 10 (33%) individuals. The authors noted that 8 of the 10 mutation-positive patients with 1 of 4 mutations causing changes in the CR2 domain of RAF1 (see, e.g., 164760.0001 and 164760.0003) had hypertrophic cardiomyopathy, whereas the 2 affected individuals with a mutation leading to changes in the CR3 domain did not (164760.0004). Transfection studies in HEK293 cells demonstrated that all 5 mutations in RAF1 behaved as gain-of-function mutants with increased kinase and ERK activation compared with wildtype RAF1.

Dilated Cardiomyopathy 1NN

In 8 patients from South and North India and 2 patients from Japan with nonsyndromic dilated cardiomyopathy (CMD1NN; 615916), Dhandapany et al. (2014) identified heterozygous mutations in the RAF1 gene (see, e.g., 164760.0005-164760.0007) that segregated with disease in the families and were not found in ancestry-matched controls. Biochemical studies showed that the CMD-associated RAF1 mutants had altered kinase activity, resulting in largely unchanged ERK (see 601795) activation but in AKT (164730) that was hyperactivated in a BRAF (164757)-dependent manner. Constitutive expression of these mutants in zebrafish embryos resulted in a heart failure phenotype with AKT hyperactivation that was rescued by treatment with rapamycin.


Cytogenetics

Jones et al. (2009) found a tandem duplication at chromosome 3p25 in a pilocytic astrocytoma (see 137800) that resulted in fusion of exons 1 through 12 of the SRGAP3 gene (606525) to exons 10 through 17 of the RAF1 gene. The fusion transcript encodes a deduced protein containing the first 513 N-terminal amino acids of SRGAP3, including the FES (190030)/CIP4 (TRIP10; 604504) homology domain, fused to 318 C-terminal amino acids of RAF1, including the entire RAF1 kinase domain. The fusion protein showed higher activity than wildtype RAF1 in phosphorylation of endogenous Mek1 in mouse fibroblasts, and it conferred anchorage-independent cell growth.


Animal Model

To examine the in vivo role of RAF1 in the heart, Yamaguchi et al. (2004) generated cardiac muscle-specific Raf1 conditionally deleted mice. The mice demonstrated left ventricular systolic dysfunction and heart dilation without cardiac hypertrophy or lethality, and showed a significant increase in the number of apoptotic cardiomyocytes. The expression level and activation of MEK1/2 (see 176872 and 601263, respectively) and ERK (see 601795) showed no difference, but the kinase activity of apoptosis signal-regulating kinase-1 (ASK1; 602448), JNK (see 601158), and p38 (600289) increased significantly compared to that of controls. The ablation of ASK1 rescued heart dysfunction and dilatation as well as cardiac fibrosis. Yamaguchi et al. (2004) concluded that RAF1 promotes cardiomyocyte survival through a MEK/ERK-independent mechanism.


ALLELIC VARIANTS 7 Selected Examples):

.0001   NOONAN SYNDROME 5

LEOPARD SYNDROME 2, INCLUDED
RAF1, SER257LEU
SNP: rs80338796, gnomAD: rs80338796, ClinVar: RCV000014985, RCV000014986, RCV000020509, RCV000149826, RCV000157426, RCV000157685, RCV000418940, RCV000428775, RCV000435984, RCV000436233, RCV000515222, RCV000624417, RCV000824754, RCV000856803, RCV001731288, RCV001813205, RCV002399323, RCV003231105

In 7 unrelated patients with Noonan syndrome (NS5; 611553) and 1 patient with LEOPARD syndrome-2 (LPRD2; 611554), Pandit et al. (2007) identified heterozygosity for a 770C-T transition in exon 7 of the RAF1 gene, resulting in a ser257-to-leu (S257L) substitution at a conserved residue in the CR2 domain. All patients had hypertrophic cardiomyopathy (CMH), including a 3.6-year-old girl with CMH at birth and a 35-year-old woman with LEOPARD syndrome. Ectopically expressed S257L mutants demonstrated increased kinase activity and enhanced ERK (see 176948) activation.

Razzaque et al. (2007) identified the S257L mutation of the RAF1 gene in 4 unrelated patients with Noonan syndrome, 3 with obstructive and 1 with nonobstructive CMH. The mutation was not found in 100 control individuals or in 100 patients with CMH without Noonan syndrome.


.0002   NOONAN SYNDROME 5

RAF1, PRO261SER
SNP: rs121434594, ClinVar: RCV000014987, RCV000159076, RCV000208421, RCV000211849, RCV000468714, RCV000618568, RCV000622893, RCV001813206, RCV003450640

In 5 affected individuals of 2 unrelated families with Noonan syndrome (NS5; 611553), Pandit et al. (2007) identified heterozygosity for a 781C-T transition in exon 7 of the RAF1 gene, resulting in a pro261-to-ser (P261S) substitution at a conserved residue in the CR2 domain. Four of the 5 patients had hypertrophic cardiomyopathy (CMH); the 1 individual with a P261S change but without CMH was a 6-year-old girl whose 38-year-old mother had the same mutation and had been diagnosed with CMH at 23 years of age. The mutation was not found in 210 control individuals.

Razzaque et al. (2007) identified the P261S mutation in 3 Noonan syndrome patients, a 1-year-old boy and his 33-year-old father and an unrelated 16-year-old boy. All 3 displayed CMH. The mutation was not found in 100 control individuals or in 100 individuals with CMH without Noonan syndrome. Transfection studies in HEK293 cells demonstrated that P261S behaved as a gain-of-function mutant with increased kinase and ERK (see 176948) activation compared with wildtype RAF1.


.0003   NOONAN SYNDROME 5

RAF1, THR491ARG
SNP: rs80338799, ClinVar: RCV000014988, RCV000680803, RCV001229313

In a sister and brother with Noonan syndrome (NS5; 611553), Pandit et al. (2007) identified heterozygosity for a 1472C-G transversion in exon 14 of the RAF1 gene, resulting in a thr491-to-arg (T491R) substitution in the CR3 domain. Neither sib had hypertrophic cardiomyopathy. The mutation was not found in 210 control individuals.


.0004   LEOPARD SYNDROME 2

NOONAN SYNDROME 5, INCLUDED
RAF1, LEU613VAL
SNP: rs80338797, ClinVar: RCV000014989, RCV000014990, RCV000020508, RCV000159089, RCV000254689, RCV000440827, RCV000824753, RCV001256891, RCV003415704

In a 43-year-old woman with LEOPARD syndrome-2 (LPRD2; 611554), Pandit et al. (2007) identified an 1837C-G transversion in exon 17 of the RAF1 gene, resulting in a leu613-to-val (L613V) substitution at a conserved residue in the C terminus. The patient had hypertrophic cardiomyopathy.

Razzaque et al. (2007) identified the L613V mutation, which they designated as being located in the CR3 domain of RAF1, in 2 unrelated boys with Noonan syndrome (NS5; 611553), neither of whom had hypertrophic cardiomyopathy. The mutation was not found in 100 control individuals or in 100 patients with hypertrophic cardiomyopathy without Noonan syndrome. Transfection studies in HEK293 cells demonstrated that L613V behaved as a gain-of-function mutant with increased kinase and ERK (see 176948) activation compared with wildtype RAF1.


.0005   CARDIOMYOPATHY, DILATED, 1NN

RAF1, LEU603PRO
SNP: rs587777586, ClinVar: RCV000131334

In 3 patients with nonsyndromic dilated cardiomyopathy (CMD1NN; 615916), a 40-year-old South Indian mother and her 4-year-old son and an unrelated 21-year-old South Indian woman, Dhandapany et al. (2014) identified heterozygosity for a c.1808T-C transition in the RAF1 gene, resulting in a leu603-to-pro (L603P) substitution at a highly conserved residue in the CR3 domain. Age at onset of disease in the 3 patients was 24 years, 3 years, and 10 years, respectively. The mutation was not found in the 4-year-old boy's unaffected father, in 500 ancestry-matched South Indian controls, or in 13,600 European and African American alleles in the Exome Sequencing Project database. Functional analysis in HEK293 cells showed impaired kinase activity and reduced ERK (see 601795) activation with the L603P mutant. Constitutive expression of the L603P mutant in zebrafish embryos resulted in a heart failure phenotype, including elongated ventricular and atrial chambers, profound pericardial edema, blood congestion at the cardiac inflow tract, and impaired cardiac contractions. Immunoblotting showed AKT hyperactivation, and the heart defects were partially rescued by treatment with rapamycin, an AKT-mTOR (601231) inhibitor.


.0006   CARDIOMYOPATHY, DILATED, 1NN

RAF1, THR641MET
SNP: rs587777587, gnomAD: rs587777587, ClinVar: RCV000131336, RCV000852545, RCV001657807, RCV001849941

In a 15-year-old South Indian girl and an unrelated 21-year-old North Indian man with nonsyndromic dilated cardiomyopathy (CMD1NN; 615916), Dhandapany et al. (2014) identified heterozygosity for a c.1922T-C transition in the RAF1 gene, resulting in a thr641-to-met (T641M) substitution at a highly conserved residue. Age at onset of disease in the 2 patients was 7 years and 16 years, respectively. The mutation, which segregated with disease in the available family, was not found in 500 ancestry-matched South Indian controls or in 350 ancestry-matched North Indian controls. Functional analysis in HEK293 cells showed a mild increase in kinase activity with the T641M mutant that was less augmented than that of the CMH-associated RAF1 mutants tested, L613V (164760.0004) and S257L (164760.0001). ERK (see 601795) activation with the T641M mutant was similar to that observed with wildtype RAF1 and was significantly less than with the CMH-associated mutants.


.0007   CARDIOMYOPATHY, DILATED, 1NN

RAF1, ALA237THR
SNP: rs587777588, gnomAD: rs587777588, ClinVar: RCV000131337, RCV001588987, RCV001775085, RCV001857459

In a 44-year-old Japanese man who had onset of nonsyndromic dilated cardiomyopathy (CMD1NN; 615916) at 40 years of age, Dhandapany et al. (2014) identified heterozygosity for a c.709G-A transition in the RAF1 gene, resulting in an ala237-to-thr (A237T) substitution at a highly conserved residue. The mutation was not found in 300 Japanese controls. Functional analysis in HEK293 cells showed a mild increase in kinase activity with the A237T mutant that was less augmented than that of the CMH-associated RAF1 mutants tested, L613V (164760.0004) and S257L (164760.0001). ERK (see 601795) activation with the A237T mutant was similar to that observed with wildtype RAF1 and was significantly less than with the CMH-associated mutants, whereas activation of AKT and tuberin (TSC2; 191092) was excessive compared to CMH-associated mutants or wildtype RAF1.


See Also:

Bonner et al. (1985); Bonner et al. (1986)

REFERENCES

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Contributors:
Ada Hamosh - updated : 01/10/2019
Marla J. F. O'Neill - updated : 7/30/2014
Ada Hamosh - updated : 8/24/2010
Patricia A. Hartz - updated : 5/19/2010
Ada Hamosh - updated : 4/15/2010
Ada Hamosh - updated : 10/19/2009
Patricia A. Hartz - updated : 5/29/2009
Marla J. F. O'Neill - updated : 10/24/2007
Ada Hamosh - updated : 1/14/2005
Marla J. F. O'Neill - updated : 12/2/2004
Ada Hamosh - updated : 12/30/2003
Ada Hamosh - updated : 7/24/2003
Jennifer P. Macke - updated : 10/20/1998

Creation Date:
Victor A. McKusick : 6/2/1986

Edit History:
carol : 12/30/2019
alopez : 05/28/2019
alopez : 01/10/2019
carol : 10/20/2016
carol : 08/23/2016
carol : 11/14/2014
carol : 8/1/2014
mcolton : 7/30/2014
mgross : 8/25/2010
terry : 8/24/2010
carol : 7/16/2010
mgross : 5/20/2010
mgross : 5/20/2010
terry : 5/19/2010
alopez : 4/20/2010
terry : 4/15/2010
alopez : 10/26/2009
alopez : 10/26/2009
terry : 10/19/2009
mgross : 6/2/2009
terry : 5/29/2009
wwang : 10/25/2007
wwang : 10/25/2007
terry : 10/24/2007
alopez : 1/20/2005
alopez : 1/20/2005
terry : 1/14/2005
carol : 12/2/2004
mgross : 4/13/2004
mgross : 3/17/2004
alopez : 12/31/2003
terry : 12/30/2003
carol : 7/24/2003
terry : 7/24/2003
alopez : 10/20/1998
mark : 6/4/1996
carol : 4/7/1993
carol : 6/1/1992
supermim : 3/16/1992
carol : 3/2/1992
carol : 2/20/1991
supermim : 3/20/1990