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A significant challenge in the post-genomic era is how to prioritize differentially expressed and uncharacterized novel genes found in hepatocellular carcinoma (HCC) microarray profiling. One such category is cell cycle regulated genes that have only evolved in higher organisms but not in lower eukaryotic cells. Characterization of these genes may reveal some novel human cancer-specific abnormalities. A novel transcript, FLJ10540 was identified. FLJ10540 is overexpressed in HCC as examined by quantitative reverse transcription-polymerase chain reaction and immunohistochemistry. The patients with higher FLJ10540 expression had a poor survival than those with lower FLJ10540 expression. Functional characterization indicates that FLJ10540 displays a number of characteristics associated with an oncogene, including anchorage-independent growth, enhanced cell growth at low serum levels and induction of tumorigenesis in nude mice. FLJ10540-elicited cell transformation is mediated by activation of the phosphatidylinositol 3'-kinase (PI3K)/AKT pathway. Moreover, FLJ10540 forms a complex with PI3K and can activate PI3K activity, which provides a mechanistic basis for FLJ10540-mediated oncogenesis. Together, using a combination of bioinformatics searches and empirical data, we have identified a novel oncogene, FLJ10540, which is conserved only in higher organisms. The finding raises the possibility that FLJ10540 is a potential new therapeutic target for HCC treatment. These findings may contribute to the development of new therapeutic strategies that are able to block the PI3K/AKT pathway in cancer cells.

Protein methylation is one of the most important post-translational modifications that contribute to the diversity and complexity of proteome. Here we report the study of in vitro methylation of heterogeneous nuclear ribonucleoprotein K (hnRNP K) with protein arginine methyltransferase 1 (PRMT1), as an enzyme, and S-adenosyl-L-methionine (SAM), as a methyl donor. The mass analysis of tryptic peptides of hnRNP K before and after methylation reveals the addition of four methyl groups in the residues 288-303. Tandem mass-spectrometric analysis of this peptide shows that both Arg296 and Arg299 are dimethylated. In addition, fragmentation analysis of such methylated arginines illustrate that they are both asymmetric dimethylarginines. Since Arg296 and Arg299 are located near the SH3-binding domains of hnRNP K, such methylation has the potential in regulating the interaction of hnRNP K with Src protein family. Our results provide crucial information for further functional study of hnRNP K methylation.

Casein kinase II is composed of two catalytic (alpha) and two regulatory (beta) subunits, the amino acid sequences of the alpha and beta subunits are highly conserved between species. To examine whether heterologous casein kinase II could be formed, recombinant alpha and beta subunits from human and Drosophila were reconstituted from inclusion bodies. Casein kinase II containing either human alpha and Drosophila beta or Drosophila alpha and human beta subunits exhibited enzymatic properties similar to those of the homologous holoenzymes with regard to specific activity, salt optima, and autophosphorylation. However, renaturation and reconstitution of casein kinase II was dependent on the type of beta subunits and the redox conditions, with the Drosophila beta subunits requiring more reduced conditions. Chimeric beta subunits prepared from human and Drosophila cDNA revealed that the N-terminal region was responsible for the requirement for the reduced redox state during renaturation. The N-terminal region also affected solubility and electrophoretic mobility of the beta subunit.