Biomedical subjects
Publications and source records attributed to Kaoru Sakabe.
Estrogen receptor beta is differentially regulated by alternative O-GlcNAcylation/O-phosphorylation at Ser(16). NMR, CD, and molecular dynamics analyses of model peptides show that these alternative modifications induce different peptide conformations, providing a molecular basis for their differential regulation of protein function.
Farnesyltransferase inhibitors (FTIs) block Ras farnesylation, subcellular localization and activity, and inhibit the growth of Ras-transformed cells. Although FTIs are ineffective against K-Ras4B, the Ras isoform most commonly mutated in human cancers, they can inhibit the growth of tumors containing oncogenic K-Ras4B, implicating other farnesylated proteins or suggesting distinct functions for farnesylated and for geranylgeranylated K-Ras, which is generated when farnesyltransferase is inhibited. In addition to bypassing FTI blockade through geranylgeranylation, K-Ras4B resistance to FTIs may also result from its higher affinity for farnesyltransferase. Using chimeric Ras proteins containing all combinations of Ras background, CAAX motif, and K-Ras polybasic domain, we show that either a polybasic domain or an alternatively prenylated CAAX renders Ras prenylation, Ras-induced Elk-1 activation, and anchorage-independent cell growth FTI-resistant. The polybasic domain alone increases the affinity of Ras for farnesyltransferase, implying independent roles for each K-Ras4B sequence element in FTI resistance. Using microarray analysis and colony formation assays, we confirm that K-Ras function is independent of the identity of the prenyl group and, therefore, that FTI inhibition of K-Ras transformed cells is likely to be independent of K-Ras inhibition. Our results imply that relevant FTI targets will lack both polybasic and potentially geranylgeranylated methionine-CAAX motifs.
A novel branch of the Ras family, Rit, was recently identified. Rit exhibits a distinct C-terminus and effector domain, and does not activate mitogen-activated protein kinase (MAPK) but can cooperate with Raf to transform fibroblasts. Here, we found that when overexpressed, activated mutants of Rit transform NIH 3T3 cells efficiently, and stimulate p38gamma but not MAPK, p38alpha, p38gamma, p38delta, or ERK5. Furthermore, we provide evidence that p38gamma activation is required for the ability of Rit to stimulate gene expression and cellular transformation. These findings suggest that this unique GTPase stimulates proliferative pathways distinct from those regulated by other Ras family members.
N-Acetylglucosamine O-linked to serines and threonines of cytosolic and nuclear proteins (O-GlcNAc) is an abundant reversible post-translational modification found in all higher eukaryotes. Evidence for functional regulation of proteins by this dynamic saccharide is rapidly accumulating. Deletion of the gene encoding the enzyme that attaches O-GlcNAc (OGT) is lethal at the single cell level, indicating the fundamental requirement for this modification. Recent studies demonstrate a role for O-GlcNAcylation in processes as diverse as transcription in the nucleus and signaling in the cytoplasm, suggesting that O-GlcNAc has both protein and site-specific influences on biochemistry and metabolism throughout the cell.