Tyrosine phosphorylation is important in signaling pathways underlying tumorigenesis. A mutational analysis of the Protein Tyrosine Kinase (PTK) gene family in cutaneous metastatic melanoma identified 30 somatic mutations in the kinase domain of 19 PTKs. The whole of the coding region of these 19 PTKs was further evaluated for somatic mutations in a total of 79 melanoma samples. This analysis revealed novel ERBB4 mutations in 19% of melanoma patients and that an additional two kinases (FLT1 and PTK2B) are mutated in 10% of melanomas. Seven missense mutations in the most commonly altered PTK (ERBB4) were examined and found to increase kinase activity and transformation ability. Melanoma cells expressing mutant ERBB4 had reduced cell growth after shRNA-mediated knockdown of ERBB4 or treatment with the ERBB inhibitor lapatinib. These studies might lead to personalized therapeutics specifically targeting the kinases that are mutationally altered in individual melanomas.Malignant melanoma is the most fatal skin cancer 1,2 . To develop personalized treatments for advanced disease, it is important to identify genetic alterations leading to melanoma. Protein tyrosine kinases (PTKs) are frequently mutated in cancer (http://www.sanger.ac.uk/genetics/CGP/Census/), and since they are amenable to pharmacologic inhibition 3,4 , further analysis of the PTK gene family may identify new therapeutic strategies. In this study, we used high-throughput gene sequencing to analyze the entire PTK gene family in melanoma, and have identified many novel somatic alterations.We initially sequenced the coding exons comprising the kinase domains of all 86 members of this gene superfamily in 29 melanomas (Supplementary Table 1). A total of 593 exons were extracted from genomic databases and amplified by polymerase chain reaction (PCR) * To whom correspondence should be addressed: National Human Genome Research Institute, 50 South Drive, MSC 8000, Building 50, Room 5140, Bethesda MD 20892-8000, Phone: 301-451-2628, Fax: 301-480-9864, samuelsy@mail.nih.gov. Author contributions T.DP. and Y.S. designed the study; J.R.W. and S.A.R. collected and analyzed the melanoma samples, N.S.A., J.C.C., K.E.Y., J.C.L., NISC., P.C and Y.S. analyzed the genetic data; T.D.P., X.W. and K.E.Y., performed and analyzed the functional data. All authors contributed to the final version of the paper. NIH Public Access Author ManuscriptNat Genet. Author manuscript; available in PMC 2010 July 6. Table 2) and directly sequenced with dye-terminator chemistry. We determined whether a mutation was somatic (i.e., tumor-specific) by examining the sequence of the gene in genomic DNA from normal tissue of the relevant patient. From the ~12 Mb of sequence information obtained, we identified 19 genes containing a total of 30 somatic mutations within their kinase domains. All coding exons of these 19 genes were then analyzed for mutations in a total of 79 melanoma samples using specific primers (Supplementary Table 3).We identified 99 non-synonymous, somatic mutations in ...

Structural equation modeling was used to test a model of test anxiety. Variables in the model included gender, number of years since one's last math course, attributions for failure and success, math self-concept, perceived self-efficacy, achievement, general test anxiety, and statistical test anxiety. Failure and success attributions were found to influence general test anxiety and statistical test anxiety for both male and female students. Women who attributed success to behavioral causes were found to have higher levels of math self-concept than women attributing success to external causes. For men, those attributing failure to external causes were found to have higher levels of the worry component of statistical test anxiety. Math self-concept was negatively related to both general test anxiety and statistics test anxiety, whereas perceived self-efficacy had a negative relationship with the worry component of statistics anxiety.The possible influences of constructs such as attributions for success and failure (

The leukemia-associated protein EVI1 possesses seven zinc fingers within an N-terminal domain (amino acids 1-250) that binds to GACAAGATA. Single amino acid missense mutants of EVI1 were developed that failed to bind DNA either in vitro, as assessed by gel shift assay, or in vivo, as shown by transactivation studies. Specifically, mutation R205N lacks high affinity binding to the GACAAGATA motif. Putative EVI1 target genes were identified by using an EVI1-(1-250)-VP16 fusion protein that acts as a transcriptional activator with the binding specificity of EVI1. Sixteen genes induced in NIH 3T3 cells by wild type EVI1-VP16 but not by mutant forms were identified. Sequence analysis revealed evolutionarily conserved GACAA-GATA-like motifs within 10 kb of their transcription start sites, and by chromatin immunoprecipitation in fibroblasts, we showed occupancy of many of these sites by EVI1-VP16. To assess whether native EVI1 binds to these sites in EVI1-transformed myeloid cells, we performed chromatin immunoprecipitation in 32Dcl3 and NFS58 cells, using anti-EVI1 antisera, and we showed that the majority of these sites is bound by wild type EVI1. These putative target genes include Gadd45g, Gata2, Zfpm2/Fog2, Skil (SnoN), Klf5 (BTEB2), Dcn, and Map3k14 (Nik). In this study we demonstrated for the first time that the N-terminal DNA binding domain of EVI1 has the capacity to bind to endogenous genes. We hypothesized that these genes play a critical role in EVI1-induced transformation.