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Chromosomal instability and the subsequent genetic mutations are considered to be critical factors in the development of the majority of solid tumors. Here, we describe how the nucleoside diphosphate kinase Nm23-H1, a protein with a known link to cancer progression, regulates a critical step during cytokinesis. Nm23-H1 acts to provide a local source of GTP for the GTPase dynamin. Loss of Nm23-H1 in diploid cells leads to cytokinetic furrow regression, followed by cytokinesis failure and generation of tetraploid cells. Loss of dynamin phenocopies loss of Nm23-H1, and ectopic overexpression of WT dynamin complements the loss of Nm23-H1. In the absence of p53 signaling, the tetraploid cells resulting from loss of Nm23-H1 continue cycling and develop classic hallmarks of tumor cells. We thus provide evidence that the loss of Nm23-H1, an event suspected to promote metastasis, may additionally function at an earlier stage of tumor development to drive the acquisition of chromosomal instability. aneuploidy | RNAi screen | cell cycle | tetraploidy | p53 doi/ 10.1073/pnas.1010633107

Sequence-specific interactions between aminoacyl-tRNA synthetases and their cognate tRNAs both ensure accurate RNA recognition and prevent the binding of noncognate substrates. Here we show for Escherichia coli glutaminyl-tRNA synthetase (GlnRS; EC 6.1.1.18) that the accuracy of tRNA recognition also determines the efficiency of cognate amino acid recognition. Steady-state kinetics revealed that interactions between tRNA identity nucleotides and their recognition sites in the enzyme modulate the amino acid affinity of GInRS. Perturbation of any of the protein-RNA interactions through mutation of either component led to considerable changes in glutamine affinity with the most marked effects seen at the discriminator base, the 10:25 base pair, and the anticodon. Reexamination of the identity set of [tRNA.sup.Gln] in the light of these results indicates that its constituents can be differentiated based upon biochemical function and their contribution to the apparent Gibbs' free energy of tRNA binding. Interactions with the acceptor stem act as strong determinants of tRNA specificity, with the discriminator base positioning the 3[prime] end. The 10:25 base pair and U35 are apparently the major binding sites to GlnRS, with G36 contributing both to binding and recognition. Furthermore, we show that E. coli tryptophanyl-tRNA synthetase also displays tRNA-dependent changes in tryptophan affinity when charging a noncognate tRNA. The ability of tRNA to optimize amino acid recognition reveals a novel mechanism for maintaining translational fidelity and also provides a strong basis for the coevolution of tRNAs and their cognate synthetases.