Complete removal of residual N-7 guanine cap from degraded messenger RNA is necessary to prevent accumulation of intermediates that might interfere with RNA processing, export, and translation. The human scavenger decapping enzyme, DcpS, catalyzes residual cap hydrolysis following mRNA degradation, releasing N-7 methyl guanosine monophosphate and 5'-diphosphate terminated cap or mRNA products. DcpS structures bound to m(7)GpppG or m(7)GpppA reveal an asymmetric DcpS dimer that simultaneously creates an open nonproductive DcpS-cap complex and a closed productive DcpS-cap complex that alternate via 30 A domain movements. Structural and biochemical analysis suggests an autoregulatory mechanism whereby premature decapping mRNA is prevented by blocking the conformational changes that are required to form a closed productive active site capable of cap hydrolysis.

The human scavenger decapping enzyme, DcpS, functions to hydrolyze the resulting cap structure following cytoplasmic mRNA decay yet is, surprisingly, a nuclear protein by immunofluorescence. Here, we show that DcpS is a nucleocytoplasmic shuttling protein that contains separable nuclear import and Crm-1-dependent export signals. We postulated that the presence of DcpS in both cellular compartments and its ability to hydrolyze cap structure may impact other cellular events dependent on capbinding proteins. An shRNA-engineered cell line with markedly diminished DcpS levels led to a corresponding reduction in capproximal intron splicing of a reporter minigene and endogenous genes. The impaired cap catabolism and resultant imbalanced cap concentrations were postulated to sequester the cap-binding complex (CBC) from its normal splicing function. In support of this explanation, DcpS efficiently displaced the nuclear cap-binding protein Cbp20 from cap structure, and complementation with Cbp20 reversed the reduced splicing, indicating that modulation of splicing by DcpS is mediated through Cbp20. Our studies demonstrate that the significance of DcpS extends beyond its well-characterized role in mRNA decay and involves a broader range of functions in RNA processing including nuclear pre-mRNA splicing.

Decapping is an important process in the control of eukaryotic mRNA degradation. The control of mRNA degradation is a critical step in the posttranscriptional regulation of gene expression, and the steady state level of any mRNA species depends on both the rate of mRNA synthesis and its breakdown. In eukaryotic cells, cytoplasmic mRNA degradation proceeds predominantly through an initial removal of the polyadenylated tail (1, 2) followed by 5Ј to 3Ј or 3Ј to 5Ј decay (3, 4). In the 5Ј-3Ј decay pathway, the 5Ј cap structure is cleaved by the catalytic activity of the Dcp2 decapping enzyme to release the m 7 Gpp and monophosphorylated RNA (5-8). The resulting uncapped monophosphorylated RNA is digested by a 5Ј-3Ј exonuclease, XrnI (1, 9). In the 3Ј-5Ј pathway, subsequent to deadenylation, the capped RNA body is continuously degraded by an exosome complex (10, 11). The resulting capped oligonucleotide m 7 GpppN(pN) n (n Ͻ 9) is hydrolyzed by a second type of decapping enzyme, DcpS, to release the m 7 Gp and ppN(pN) n products (12, 13). These pathways need not be mutually exclusive and could occur simultaneously (14, 15), and an interplay between the two pathways could also exist. DcpS, which was originally characterized as the decapping enzyme in the 3Ј-5Ј pathway, is also able to hydrolyze the 5Ј-3Ј decapping product m 7 Gpp to release m 7 Gp (16,17). Furthermore, disruption of the yeast DcpS ortholog, Dcs1p, impeded the 5Ј-exonuclease activity (18), indicating that the Dcs1p decapping products might serve as signaling molecules for the 5Ј decay pathway.DcpS is a member of the histidine triad (HIT) 3 hydrolase family of proteins that contain a stretch of His-X-His-X-His-X residues, where X denotes hydrophobic amino acid residues. HIT proteins are dimeric nucleotide binding proteins that have hydrolase activities (19 -21). The central histidine residue is critical for the hydrolase activity, since it is thought to serve as the nucleophile attacking the phosphate most proximal to the methylated guanosine in m 7 GpppG (22) and is also critical for DcpS hydrolysis (13).Structural analysis of DcpS has revealed that it is a homodimer with a symmetric structure when in the ligand-free form (16, 23) or asymmetric homodimer in the ligand-bound form (16,24). Each DcpS monomer possesses a distinct N-terminal domain and a C-terminal domain containing the HIT motif linked by a hinge region. The N-terminal domain displays a domain-swapped form by exchanging an identical ␣-helix and two antiparallel ␤-strands with the second monomer. The DcpS homodimer contains two cap binding pockets, which serve as the active sites for cap hydrolysis. In the ligand-bound form, DcpS forms a closed conformation on one side and an open conformation on the other, with substrate bound at the C-terminal domain of each side (24). The structure suggests that the closed conformation constitutes the cap hydrolysis productive site, whereas the open site would be nonproductive. The N-terminal domains can both be in the open state, or one side can be in an...

RBM4 modulates alternative exon selection of Numb and up-regulates proneural Mash1 gene expression, possibly via specific Numb isoforms. RBM4 overexpression promotes neuronal cell differentiation. Moreover, RBM4 is essential for neurite outgrowth in primary cortical neurons by modulating specific Numb isoform expression.

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