One of the main mechanisms of messenger RNA degradation in eukaryotes occurs by deadenylation-dependent decapping which leads to 5'-to-3' decay. A family of Sm-like (Lsm) proteins has been identified, members of which contain the 'Sm' sequence motif, form a complex with U6 small nuclear RNA and are required for pre-mRNA splicing. Here we show that mutations in seven yeast Lsm proteins (Lsm1-Lsm7) also lead to inhibition of mRNA decapping. In addition, the Lsm1-Lsm7 proteins co-immunoprecipitate with the mRNA decapping enzyme (Dcp1), a decapping activator (Pat1/Mrt1) and with mRNA. This indicates that the Lsm proteins may promote decapping by interactions with the mRNA and the decapping machinery. In addition, the Lsm complex that functions in mRNA decay appears to be distinct from the U6-associated Lsm complex, indicating that Lsm proteins form specific complexes that affect different aspects of mRNA metabolism.

Decapping is a critical step in mRNA decay. In the 59-to-39 mRNA decay pathway conserved in all eukaryotes, decay is initiated by poly(A) shortening, and oligoadenylated mRNAs (but not polyadenylated mRNAs) are selectively decapped allowing their subsequent degradation by 59 to 39 exonucleolysis. The highly conserved heptameric Lsm1p-7p complex (made up of the seven Sm-like proteins, Lsm1p-Lsm7p) and its interacting partner Pat1p activate decapping by an unknown mechanism and localize with other decapping factors to the P-bodies in the cytoplasm. The Lsm1p-7p-Pat1p complex also protects the 39-ends of mRNAs in vivo from trimming, presumably by binding to the 39-ends. In order to determine the intrinsic RNA-binding properties of this complex, we have purified it from yeast and carried out in vitro analyses. Our studies revealed that it directly binds RNA at/near the 39-end. Importantly, it possesses the intrinsic ability to distinguish between oligoadenylated and polyadenylated RNAs such that the former are bound with much higher affinity than the latter. These results indicate that the intrinsic RNAbinding characteristics of this complex form a critical determinant of its in vivo interactions and functions.

The decapping of eukaryotic mRNA s is a key step in their degradation. The heteroheptameric L sm1p-7p complex is a general activator of decapping and also functions in protecting the 3Ј ends of deadenylated mRNA s from a 3Ј-trimming reaction. L sm1p is the unique member of the L sm1p-7p complex, distinguishing that complex from the functionally different L sm2p-8p complex. To understand the function of L sm1p, we constructed a series of deletion and point mutations of the LSM1 gene and examined their effects on phenotype. These studies revealed the following: (i) Mutations affecting the predicted RNAbinding and inter-subunit interaction residues of L sm1p led to impairment of mRNA decay, suggesting that the integrity of the L sm1p-7p complex and the ability of the L sm1p-7p complex to interact with mRNA are important for mRNA decay function; (ii) mutations affecting the predicted RNA contact residues did not affect the localization of the L sm1p-7p complex to the P-bodies; (iii) mRNA 3Ј-end protection could be indicative of the binding of the L sm1p-7p complex to the mRNA prior to activation of decapping, since all the mutants defective in mRNA 3Ј end protection were also blocked in mRNA decay; and (iv) in addition to the Sm domain, the C-terminal domain of L sm1p is also important for mRNA decay function.

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