Perspective

BRAF and MEK Mutations Make a Late Entrance

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Science's STKE  28 Mar 2006:
Vol. 2006, Issue 328, pp. pe15
DOI: 10.1126/stke.3282006pe15

Abstract

The small guanosine triphosphatase KRAS and the protein kinases BRAF, which is a mitogen-activated protein kinase kinase kinase (MAPKKK), and mitogen-activated protein kinase kinase 1 and 2 (MAPKK1/2, also known as MKK1/2 or MEK1/2) are signaling partners in the MAPK signal transduction pathway. They are involved in many biological processes and play crucial roles during embryonic development. When inappropriately expressed, KRAS, BRAF, and MEK1/2 are also frequently implicated in tumor progression. Hence, it might reasonably have been predicted that either loss- or gain-of-function germline mutations in the genes that encode them would cause embryonic death. However, in a surprising development, two articles report that germline mutations in the KRAS, BRAF, and MEK1/2 genes are associated with cardio-facio-cutaneous (CFC) syndrome. This unexpected discovery demonstrates that mutations in KRAS, BRAF, and MEK can pass through the germline to cause specific developmental syndromes. This finding will undoubtedly stimulate further research into the function of these proteins in development and in both inherited and sporadic cancers.

Two kinases of the mitogen-activated protein kinase kinase [MAPKK, also known as the mitogen-activated protein kinase (MAPK)/extracellular signal–regulated kinase (ERK) kinase (MKK or MEK)] family, MEK1 and MEK2, are evolutionarily conserved, regulatory protein kinases that play pivotal roles in a wide variety of developmental cellular processes, including growth, division, and differentiation. Instructions are relayed to these proteins through a signaling cascade (Fig. 1). Extracellular growth factors bind specific cell surface receptors, causing changes in their conformation and in some cases, such as with tyrosine kinase receptors, inducing intrinsic kinase activity. This activity leads to the recruitment of adaptor proteins, like Grb2 and SHC, and the activation of proteins, like Ras and Raf, at the inner surface of the cell membrane. Raf is a mitogen-activated protein kinase kinase kinase (MAPKKK) that phosphorylates and activates MEK 1 or 2, which in turn phosphorylate extracellular signal–regulated, mitogen-activated protein kinase 1 or 2 (ERK1/2 or MAPK1/2). Activated MAPK1/2 then phosphorylates and activates effector molecules that control cellular processes at the transcriptional, translational, or posttranslational levels (1).

Fig. 1.

Patterned expression of phosphorylated, active MAPK in mouse embryos. (Left) Outline of MEK-to-MAPK signaling pathway. (Right) Pattern of phosphorylated ERK expression in a 10.5-day postcoitus mouse embryo. ba, branchial arches; fnp, frontonasal processes; lb, limb buds; lp, liver primordia; np, nasal pits; TSI, tissue-specific inhibitors. [Mouse image courtesy of J. Rossant, Toronto Sick Kids Hospital]

MEK-to-MAPK signaling is crucial at multiple stages of embryonic development. For example, increased MAPK activity triggers meiotic maturation of oocytes and maintains cell cycle arrest at metaphase II in eggs before fertilization (2, 3). In addition, MAPK signaling is required for mesoderm induction during gastrulation (4). At later stages of embryogenesis, the spatial and temporal control of MAPK activity is exquisitely regulated in Drosophila, Xenopus, and mice (57). In mice, sustained MAPK activity is detected in the ectoplacental cone, extraembryonic ectoderm, limb buds, branchial arches, and frontonasal process, as well as in the tailbud, forebrain, midbrain-hindbrain boundary, foregut, and liver primordia; transient MAPK activity is seen in the neural crest, peripheral nervous system, nascent blood vessels, and precursors of the eye, ear, and heart (Fig. 1) (7). Further, as might be expected, disruption of MEK-to-MAPK signaling has deadly consequences: Gene-targeted deletion of KRAS or BRAF in mice results in death at mid-gestation (8, 9), and MEK1-null mice die in utero with extensive defects in placental vascularization (10).

Given the multiple, key roles for MAPK signaling in embryonic development, it might reasonably have been predicted that germline mutations in this pathway would be embryonic lethal and so not be associated with specific developmental disorders in humans. However, nature has chosen to remind us how little we truly understand its complexities. Rodriguez-Viciana et al. (11) and Niihori et al. (12) show that genetic mutations in KRAS, BRAF, MEK1, and MEK2 are associated with cardio-facio-cutaneous (CFC) syndrome. CFC is a rare, autosomal, and presumably dominant syndrome characterized by distinctive facial features, cardiac anomalies, hair and skin abnormalities, postnatal growth deficiency, and hypotonia (low muscle tone) (13). Rodriguez-Viciana’s group screened a panel of blood samples from CFC patients for germline mutations in this pathway and found 11 different missense mutations in BRAF (9 of which had not been previously identified) in 18 out of 23 patients. Likewise, Niihori’s group screened for mutations in 43 samples and found germline KRAS and BRAF mutations in 3 and 19 patients, respectively. No mutations were detected in the remaining 24 patient samples. Rodriguez-Viciana’s group also found that three of their remaining five patients harbored novel mutations in MEK1 and MEK2. This is particularly noteworthy because it is the first identified instance of naturally occurring mutations in these genes. Their observations also offer interesting insight into the biochemistry of MEK 1 and 2 because none of the mutated residues had previously been shown to influence MEK’s biologic activities.

Whereas most of the mutations that Rodriguez-Viciana et al. identified caused constitutive activation of the MEK pathway, at least one (G596V BRAF) appeared to be deficient in its ability to activate MEK. This seems paradoxical because it suggests that a similar phenotype may be caused by both activating and inactivating mutations in BRAF. A similar situation exists for the related autosomal dominant Noonan (NS) and LEOPARD syndromes (LS), which are caused by gain-of-function mutations and inactivating mutations in the protein tyrosine phosphatase PTP11 (SHP2), leading to constitutive (14, 15) or impaired MAPK activation (16), respectively. If the developmental processes underlying these syndromes require transient activation of MAPK, then the dampened or sustained elevation of MAPK activity may be sufficient to disrupt it. Alternatively, loss of MAPK signaling may trigger activation of a compensatory mechanism, such as an alternative RAF signaling pathway that mimics sustained activation of this pathway and elicits a similar phenotypic result.

CFC, NS, and LS—as well as Costello syndrome (CS), an autosomal dominant disease that has been linked to activating mutations in HRAS (17, 18)—each give rise to a similar array of phenotypic consequences, including facial dysmorphia, cardiomyopathy, and abnormal growth. Each of these syndromes also carries with it an elevated risk of malignancy. Patients with CS have an increased incidence of rhabdomyosarcoma, transitional cell carcinoma, and neuroblastoma (19); NS patients have an increased incidence of rhabdomyosarcoma, juvenile myelomonocytic leukemia, and acute lymphoblastic leukemia (20); and LS patients have an increased incidence of acute myelogenous leukemia and neuroblastoma (21, 22). CFC has not been definitively linked with malignancies though this may be a reflection of the limited number of patients with this disease. In one respect, this is not surprising given that somatic mutations in KRAS and BRAF have been identified in many cancers and that activated MAPK or elevated MAPK expression has been detected in various human tumors. What is perplexing, though, is that each of these syndromes should be associated with such a narrow, yet distinct, range of malignancies. The results of these latest studies indicate that there must exist tissue-specific inhibitors that repress elevated BRAF-to-MEK signaling and that these must function at the level of MAPK signaling or at a later stage in the signaling cascade.

MAPK and MEK were first identified almost 20 and 15 years ago, respectively (23, 24), and their ability to transform cells was established more than 10 years ago (25). Their roles in cancer and development have been extensively studied. Why, then, has it taken so long to identify germline mutations in these genes? The answer may be simply that no one thought to look. Fortunately, Rodriguez-Viciana and colleagues felt unencumbered by popular preconceptions. Their novel finding that germline mutations in KRAS, BRAF, MEK1, and MEK2 are associated with a specific developmental syndrome represents an important next step in our understanding of these proteins. BRAF and MEK mutations have made a late, albeit grand, entrance.

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