Nuclear pore component Nup98 is a potential tumor suppressor and regulates posttranscriptional expression of select p53 target genes.

The p53 tumor suppressor utilizes multiple mechanisms to selectively regulate its myriad target genes, which in turn mediate diverse cellular processes. Here, using conventional and single-molecule mRNA analyses, we demonstrate that the nucleoporin Nup98 is required for full expression of p21, a key effector of the p53 pathway, but not several other p53 target genes. Nup98 regulates p21 mRNA levels by a posttranscriptional mechanism in which a complex containing Nup98 and the p21 mRNA 3'UTR protects p21 mRNA from degradation by the exosome. An in silico approach revealed another p53 target (14-3-3σ) to be similarly regulated by Nup98. The expression of Nup98 is reduced in murine and human hepatocellular carcinomas (HCCs) and correlates with p21 expression in HCC patients. Our study elucidates a previously unrecognized function of wild-type Nup98 in regulating select p53 target genes that is distinct from the well-characterized oncogenic properties of Nup98 fusion proteins.

In vivo single-particle imaging of nuclear mRNA export in budding yeast demonstrates an essential role for Mex67p.

Many messenger RNA export proteins have been identified; yet the spatial and temporal activities of these proteins and how they determine directionality of messenger ribonucleoprotein (mRNP) complex export from the nucleus remain largely undefined. Here, the bacteriophage PP7 RNA-labeling system was used in Saccharomyces cerevisiae to follow single-particle mRNP export events with high spatial precision and temporal resolution. These data reveal that mRNP export, consisting of nuclear docking, transport, and cytoplasmic release from a nuclear pore complex (NPC), is fast (∼ 200 ms) and that upon arrival in the cytoplasm, mRNPs are frequently confined near the nuclear envelope. Mex67p functions as the principal mRNP export receptor in budding yeast. In a mex67-5 mutant, delayed cytoplasmic release from NPCs and retrograde transport of mRNPs was observed. This proves an essential role for Mex67p in cytoplasmic mRNP release and directionality of transport.

Positive charge is an important feature of the C-terminal transport signal of the VirB/D4-translocated proteins of Agrobacterium.

Several human pathogens and the plant pathogen Agrobacterium tumefaciens use a type IV secretion system for translocation of effector proteins into host cells. How effector proteins are selected for transport is unknown, but a C-terminal transport signal is present in the proteins translocated by the A. tumefaciens VirB/D4 type IV secretion system. We characterized this signal in the virulence protein VirF by alanine scanning and further site-directed mutagenesis. The Cre recombinase was used as a reporter to measure the translocation efficiency of Cre-Vir fusions from A. tumefaciens to Arabidopsis. The data unambiguously showed that positive charge is an essential characteristic of the C-terminal transport signal. We increased the sensitivity of this translocation assay by modifying the Cre-induced readout in host cells from kanamycin resistance to GFP expression. This improvement allowed us to detect translocation of the VirD2 relaxase protein in the absence of transferred DNA, showing that attachment to the transferred DNA is not essential for transport by the VirB/D4 system. We also found another translocated effector protein, namely the VirD5 protein encoded by the tumor-inducing plasmid. According to secondary structure predictions, the C termini of all VirB/D4-translocated proteins identified so far are unstructured; however, they contain a characteristic hydropathic profile. Based on sequence alignments and mutational analysis of VirF, we conclude that the C-terminal transport signal for recruitment and translocation of effector proteins by the A. tumefaciens VirB/D4 system is hydrophilic and has a net positive charge with a consensus motif of R-X(7)-R-X-R-X-R-X-X(n)>.