Functional analysis of mRNA scavenger decapping enzymes

Liu, Shin Wu; Jiao, Xuan; Liu, Hudan; Gu, Meigang; Lima, Christopher D.; Kiledjian, Megerditch

doi:10.1261/rna.7660804

Cited by 76 publications

(126 citation statements)

References 37 publications

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“…The saturated amplitudes indicate that the DcpS-cap complex K d is Ͻ20 nM (Fig. 1E), consistent with the K d of 75 nM we previously reported (29).…”

Section: Hydrolysis Of Cap Structure By Dcps Is Rate-limiting At the supporting

confidence: 91%

Mechanistic and Kinetic Analysis of the DcpS Scavenger Decapping Enzyme

Liu

Rajagopal²,

Patel³

et al. 2008

Journal of Biological Chemistry

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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...

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“…The saturated amplitudes indicate that the DcpS-cap complex K d is Ͻ20 nM (Fig. 1E), consistent with the K d of 75 nM we previously reported (29).…”

Section: Hydrolysis Of Cap Structure By Dcps Is Rate-limiting At the supporting

confidence: 91%

Mechanistic and Kinetic Analysis of the DcpS Scavenger Decapping Enzyme

Liu

Rajagopal²,

Patel³

et al. 2008

Journal of Biological Chemistry

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“…Findings presented by Mike Kiledjian (Rutgers) provide an explanation for the observa-tion that intact mRNAs are not substrates for DcpS. Kiledjian showed that the affinity and subsequent activity of mammalian DcpS for 5Ј cap structures is sensitive to the length of the RNA in which the cap is attached Liu et al 2004). DcpS demonstrates 2500-fold greater activity on a capped dinucleotide versus a full-length capped mRNA substrate, and, moreover, the robust activity of DcpS on capped decay intermediates effectively competes for, and hydrolyzes, cap structure in the presence of the cap-binding protein, eIF4E.…”

Section: Mrna Stabilization By Cis-acting Elements: Parallels Betweenmentioning

confidence: 99%

Under the Tucson sun: A meeting in the desert on mRNA decay

Baker¹,

Condon²

2004

RNA

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“…DcpS utilizes an evolutionarily conserved Histidine Triad (HIT) motif (Liu et al 2002) to hydrolyze the triphosphate linkage of the cap structure and is the only known cap-binding protein that hydrolyzes its substrate (Liu et al 2002). Despite its purification from cytoplasmic extract (Liu et al 2002) and its well-characterized cytoplasmic decapping function (Liu et al 2002;van Dijk et al 2003;Cohen et al 2004), DcpS is predominantly if not exclusively a nuclear protein by immunocytochemistry (Salehi et al 2002;Cougot et al 2004;Liu et al 2004).…”

Section: Introductionmentioning

confidence: 99%

“…Recent experiments reveal an auto-regulatory mechanism whereby DcpS positively feeds back onto the 59-39 directional mRNA degradation pathway (Liu and Kiledjian 2005). Despite these studies that emphasize cytoplasmic functions, the unexpected DcpS nuclear localization (Cougot et al 2004;Liu et al 2004) strongly indicates an unknown nucleus-associated function.…”

Section: Introductionmentioning

confidence: 99%

DcpS scavenger decapping enzyme can modulate pre-mRNA splicing

Shen¹,

Liu²,

Liu³

et al. 2008

RNA

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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.

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Functional analysis of mRNA scavenger decapping enzymes

Cited by 76 publications

References 37 publications

Mechanistic and Kinetic Analysis of the DcpS Scavenger Decapping Enzyme

Mechanistic and Kinetic Analysis of the DcpS Scavenger Decapping Enzyme

Under the Tucson sun: A meeting in the desert on mRNA decay

DcpS scavenger decapping enzyme can modulate pre-mRNA splicing

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