Stark MS, Woods SL, Gartside MG et al.Frequent somatic mutations in MAP3K5 and MAP3K9 in metastatic melanoma identified by exome sequencing. Nat Genet 44:165-169

Queensland Institute of Medical Research, Brisbane, Queensland, Australia.
Nature Genetics (Impact Factor: 29.35). 12/2011; 44(2):165-9. DOI: 10.1038/ng.1041
Source: PubMed


We sequenced eight melanoma exomes to identify new somatic mutations in metastatic melanoma. Focusing on the mitogen-activated protein (MAP) kinase kinase kinase (MAP3K) family, we found that 24% of melanoma cell lines have mutations in the protein-coding regions of either MAP3K5 or MAP3K9. Structural modeling predicted that mutations in the kinase domain may affect the activity and regulation of these protein kinases. The position of the mutations and the loss of heterozygosity of MAP3K5 and MAP3K9 in 85% and 67% of melanoma samples, respectively, together suggest that the mutations are likely to be inactivating. In in vitro kinase assays, MAP3K5 I780F and MAP3K9 W333* variants had reduced kinase activity. Overexpression of MAP3K5 or MAP3K9 mutants in HEK293T cells reduced the phosphorylation of downstream MAP kinases. Attenuation of MAP3K9 function in melanoma cells using siRNA led to increased cell viability after temozolomide treatment, suggesting that decreased MAP3K pathway activity can lead to chemoresistance in melanoma.

Download full-text


Available from: Irene Newsham
  • Source
    • "We also observed three recurrent alterations at S278F, E371K, and E1175K as well as five nonsense mutations. Recently, another group recently published a whole-exome screen of eight melanoma samples and found two additional somatic mutations in GRIN2A, suggesting that genetic alteration of this gene is important (Stark et al., 2011). Furthermore, seven somatic mutations found in our screen have been discovered by others as well as some listed in the COSMIC database (D252N, S278F, W343X, G449E, M653I, R920K, S929F, and E1073K; Berger et al., 2012; Hodis et al., 2012) (http://cancer.sanger. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The ionotropic glutamate receptors (NMDAR) are composed of large complexes of multi-protein subunits creating ion channels in the cell plasma membranes that allow for influx or efflux of mono- or divalent cations (e.g., Ca(2+)) important for synaptic transmissions, cellular migration and survival. Recently, we discovered the high prevalence of somatic mutations within one of the ionotropic glutamate receptors, GRIN2A, in malignant melanoma. Functional characterization of a subset of GRIN2A mutants demonstrated a loss of NMDAR complex formation between GRIN1 and GRIN2A, increased anchorage-independent growth in soft agar, and increased migration. Somatic mutation of GRIN2A results in a dominant negative effect inhibiting the tumor suppressive phenotype of wild type GRIN2A in melanoma. Depletion of endogenous GRIN2A in melanoma cells expressing wild-type GRIN2A resulted in increased proliferation compared to control. In contrast, shRNA depletion of GRIN2A in mutant cell lines slightly reduced proliferation. Our data shows that somatic mutation of GRIN2A results in increased survival and is the first such report to demonstrate the functional importance of GRIN2A mutations in melanoma and the significance ionotropic glutamate receptor signaling plays in malignant melanoma.Journal of Investigative Dermatology accepted article peview online, 16 April 2014. doi:10.1038/jid.2014.190.
    Full-text · Article · Apr 2014 · Journal of Investigative Dermatology
  • Source
    • "Kinases have been a frequent target for targeted therapies and therefore identification of mutation in these genes will be critical for future treatment regimens24. A number of exome studies have identified recurrent mutations in the mitogen-activated protein kinase genes MAP3K9, MAP3K5 and MAP2K12526. Mutations in these genes were identified in three of our unfixed/formalin-fixed pairs (A2058, HT144 and MeWo) (Table 2). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Massively parallel sequencing offers the ability to interrogate a tumour biopsy for multiple mutational changes. For clinical samples, methodologies must enable maximal extraction of available sequence information from formalin-fixed and paraffin-embedded (FFPE) material. We assessed the use of targeted capture for mutation detection in FFPE DNA. The capture probes targeted the coding region of all known kinase genes and selected oncogenes and tumour suppressor genes. Seven melanoma cell lines and matching FFPE xenograft DNAs were sequenced. An informatics pipeline was developed to identify variants and contaminating mouse reads. Concordance of 100% was observed between unfixed and formalin-fixed for reported COSMIC variants including BRAF V600E. mutations in genes not conventionally screened including ERBB4, ATM, STK11 and CDKN2A were readily detected. All regions were adequately covered with independent reads regardless of GC content. This study indicates that hybridisation capture is a robust approach for massively parallel sequencing of FFPE samples.
    Full-text · Article · Dec 2013 · Scientific Reports
  • Source
    • "No recurrent substitutions were found in known melanoma genes or genes in the Cancer Gene Census [24]. Over 60% of genes mutated in this study were found to be mutated at least once in human melanoma [9-11,25-27], which was unsurprising given the extensive mutation load in the human disease. Substitutions with predicted coding changes in known census cancer genes included a nonsense mutation in ikzf and missense mutations in nup214 and pik3cd, while a homozygous missense substitution in the anaphase promoting complex gene, anapc1, was identified in a BRAF, p53+/- tumor (ZD8a). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Melanoma is the most deadly form of skin cancer. Expression of oncogenic BRAF or NRAS, which are frequently mutated in human melanomas, promote the formation of nevi but are not sufficient for tumorigenesis. Even with germline mutated p53, these engineered melanomas present with variable onset and pathology, implicating additional somatic mutations in a multi-hit tumorigenic process. To decipher the genetics of these melanomas, we sequence the protein coding exons of 53 primary melanomas generated from several BRAFV600E or NRASQ61K driven transgenic zebrafish lines. We find that engineered zebrafish melanomas show an overall low mutation burden, which has a strong, inverse association with the number of initiating germline drivers. Although tumors reveal distinct mutation spectrums, they show mostly C > T transitions without UV light exposure, and enrichment of mutations in melanogenesis, p53 and MAPK signaling. Importantly, a recurrent amplification occurring with pre-configured drivers BRAFV600E and p53-/- suggests a novel path of BRAF cooperativity through the protein kinase A pathway. This is the first analysis of a melanoma mutational landscape in the absence of UV light, where tumors manifest with remarkably low mutation burden and high heterogeneity. Genotype specific amplification of protein kinase A in cooperation with BRAF and p53 mutation suggests the involvement of melanogenesis in these tumors. This work is important for defining the spectrum of events in BRAF or NRAS driven melanoma in the absence of UV light, and for informed exploitation of models such as transgenic zebrafish to better understand mechanisms leading to human melanoma formation.
    Full-text · Article · Oct 2013 · Genome biology
Show more