In this study, we developed a new pharmacophore-based interaction fingerprint (Pharm-IF) and examined its usefulness for in silico screening using machine learning techniques such as support vector machine (SVM) and random forest (RF) instead of similarity-based ranking. Using the docking results of PKA, SRC, cathepsin K, carbonic anhydrase II, and HIV-1 protease, the screening efficiencies of the Pharm-IF models were compared to GLIDE score and the residue-based IF (PLIF) models. The combination of SVM and Pharm-IF demonstrated a higher enrichment factor at 10% (5.7 on average) than those of GLIDE score (4.2) and PLIF (4.3). In terms of the size of the training sets, learning more than five crystal structures enabled the machine learning models to stably achieve better efficiencies than GLIDE score. We also employed the docking poses of known active compounds, in addition to the crystal structures, as positive samples of training sets. The enrichment factors of the RF models at 10% using the docking poses for SRC and cathepsin K showed significantly higher values (6.5 and 6.3) than those using only the crystal structures (3.9 and 3.2), respectively.

The development of cancer is driven not only by genetic mutations but also by epigenetic alterations. Here, we show that TET1-mediated production of 5-hydroxymethylcytosine (5hmC) is required for the tumorigenicity of glioblastoma cells. Furthermore, we demonstrate that chromatin target of PRMT1 (CHTOP) binds to 5hmC. We found that CHTOP is associated with an arginine methyltransferase complex, termed the methylosome, and that this promotes the PRMT1-mediated methylation of arginine 3 of histone H4 (H4R3) in genes involved in glioblastomagenesis, including EGFR, AKT3, CDK6, CCND2, and BRAF. Moreover, we found that CHTOP and PRMT1 are essential for the expression of these genes and that CHTOP is required for the tumorigenicity of glioblastoma cells. These results suggest that 5hmC plays a critical role in glioblastomagenesis by recruiting the CHTOP-methylosome complex to selective sites on the chromosome, where it methylates H4R3 and activates the transcription of cancer-related genes.

In addition to their pleiotropic functions under physiological conditions, transcription factors STAT3 and STAT5 also have oncogenic activities, but how activated STATs are transported to the nucleus has not been fully understood. Here we show that an MgcRacGAP mutant lacking its nuclear localizing signal (NLS) blocks nuclear translocation of p-STATs both in vitro and in vivo. Unlike wild-type MgcRacGAP, this mutant did not promote complex formation of phosphorylated STATs (p-STATs) with importin ␣ in the presence of GTPbound Rac1, suggesting that MgcRacGAP functions as an NLS-containing nuclear chaperone. We also demonstrate that mutants of STATs lacking the MgcRacGAP binding site (the strand ␤b) are hardly tyrosine phosphorylated after cytokine stimulation. Intriguingly, mutants harboring small deletions in the C-adjacent region (␤b-␤c loop region) of the strand ␤b became constitutively active with the enhanced binding to MgcRacGAP. The molecular basis of this phenomenon will be discussed, based on the computer-assisted tertiary structure models of STAT3. Thus, MgcRacGAP functions as both a critical mediator of STAT's tyrosine phosphorylation and an NLS-containing nuclear chaperone of p-STATs.The STAT (signal transducers and activators of transcription) family proteins (STAT1 to -4, -5A, -5B, and -6) are phosphorylated by cytokine stimulation, form homo-or heterodimers, and enter the nucleus, where they regulate expression of their target genes (6, 13). STATs have a variety of functions, including antiapoptosis, proliferation, differentiation, inflammation, and development under physiological conditions, and of note, the oncogenic activities of STAT3 and STAT5 have also been demonstrated under pathological conditions (5).A small GTPase Rac1 is implicated in cytoskeletal organization, membrane ruffling, production of superoxide, phagocytosis, and chemotaxis as well as regulation of the cell cycle (15, 39). Recent studies revealed its distinct roles in nuclear translocation of phosphorylated STATs (p-STATs) and ␤-catenin and also its nuclear accumulation in the G 2 phase, promoting cell division (17,31,47). MgcRacGAP is an evolutionarily conserved GTPase-activating protein (GAP) for Rho family GTPases. We and others previously showed that MgcRacGAP controls the mitotic spindle through associating ␣-, ␤-, and ␥-tubulin, Rho family GTPases, and a kinesin protein, MKLP1, and plays essential roles in the completion of cytokinesis, accumulating to the midbody (12, 16, 32). Very recently, Yamada et al. reported that conditional knockout of MgcRac-GAP results in acute apoptosis even before the failure of cytokinesis in interleukin-7 (IL-7)-expanded B220 ϩ cells (48), indicating that MgcRacGAP is not simply involved in cell division but also in cell survival, at least in IL-7-expanded B220 ϩ cells.Molecular trafficking between the nucleus and cytoplasm occurs via nuclear pore complexes. To enter the nucleus, nuclear proteins larger than ϳ50 kDa usually harbor a functional nuclear localization signal (NLS) or bind NLS-co...