MicroRNAs (miRNAs) constitute a large class of regulatory RNAs that repress target messenger RNAs to control various biological processes. Accordingly, miRNA biogenesis is highly regulated, controlled at both transcriptional and post-transcriptional levels, and overexpression and underexpression of miRNAs are linked to various human diseases, particularly cancers. As RNA concentrations are generally a function of biogenesis and turnover, active miRNA degradation might also modulate miRNA accumulation, and the plant 3'-->5' exonuclease SDN1 has been implicated in miRNA turnover. Here we report that degradation of mature miRNAs in the nematode Caenorhabditis elegans, mediated by the 5'-->3' exoribonuclease XRN-2, affects functional miRNA homeostasis in vivo. We recapitulate XRN-2-dependent miRNA turnover in larval lysates, where processing of precursor-miRNA (pre-miRNA) by Dicer, unannealing of the miRNA duplex and loading of the mature miRNA into the Argonaute protein of the miRNA-induced silencing complex (miRISC) are coupled processes that precede degradation of the mature miRNA. Although Argonaute:miRNA complexes are highly resistant to salt, larval lysate promotes efficient release of the miRNA, exposing it to degradation by XRN-2. Release and degradation can both be blocked by the addition of miRNA target RNA. Our results therefore suggest the presence of an additional layer of regulation of animal miRNA activity that might be important for rapid changes of miRNA expression profiles during developmental transitions and for the maintenance of steady-state concentrations of miRNAs. This pathway might represent a potential target for therapeutic intervention on miRNA expression.
MicroRNAs (miRNAs) are tightly regulated through transcriptional and posttranscriptional mechanisms, including degradation by nucleases. Here, we report that in C. elegans, target mRNAs can protect their cognate miRNAs from degradation in vivo. We show that the let-7(n2853) mutation destabilizes the mature let-7 miRNA by impairing this protection. Moreover, presence of a cognate target or depletion of the xrn-1 (XRN1) or xrn-2 (XRN2/Rat1p) exoribonucleases enforces accumulation of certain miRNA passenger (miR(∗)) strands. Thus, following biased miRNA strand loading into Argonaute, elimination of nonfunctional RNAs can further refine miRNA strand selection. Conversely, by aligning the levels of miRNAs with those of their targets, the opposing activities of mature miRNA degradation and target-mediated miRNA protection (TMMP) may enable dynamic expression of either mature strand of a pre-miRNA, and evolution of miRNAs. Thus, it seems that mRNAs are more than inert targets and function with miRNAs in a network of mutual regulation.
In kinetoplastid protozoa, import of cytosolic tRNAs into mitochondria occurs through tRNAs interacting with membrane-bound proteins, the identities of which are unknown. The inner membrane RNA import complex of Leishmania tropica contains multiple proteins and is active for import in vitro . RIC1, the largest subunit of this complex, is structurally homologous to the conserved α subunit of F1 ATP synthase. The RIC1 gene complemented an atpA mutation in Escherichia coli . Antisense-mediated knockdown of RIC1/F1α in Leishmania resulted in depletion of several mitochondrial tRNAs belonging to distinct subsets (types I and II) that interact cooperatively or antagonistically within the import complex. The knockdown-induced defect in import of type I tRNAs was rectified in a reconstituted system by purified RIC1/F1α alone, but recovery of type II tRNA import additionally required a type I tRNA. RIC1/F1α formed stable complexes with type I, but not type II, tRNAs through the cooperation of its nucleotide binding and C-terminal domains. Thus, RIC1/F1α is a type I tRNA import receptor. As expected of a bifunctional protein, RIC1/F1α is shared by both the import complex and by respiratory complex V. Alternative use of ancient respiratory proteins may have been an important step in the evolution of tRNA import.
The mitochondrial genomes of a wide variety of species contain an insufficient number of functional tRNA genes, and translation of mitochondrial mRNAs is sustained by import of nucleus-encoded tRNAs. In Leishmania, transfer of tRNAs across the inner membrane can be regulated by positive and negative interactions between them. To define the factors involved in such interactions, a large multisubunit complex (molecular mass, ϳ640 kDa) from the inner mitochondrial membrane of the kinetoplastid protozoon Leishmania, consisting of ϳ130-Å particles, was isolated. The complex, when incorporated into phospholipid vesicles, induced specific, ATP-and proton motive force-dependent transfer of Leishmania There is remarkable diversity in the scope and mechanism of mitochondrial tRNA import (reviewed in reference 18). Human mitochondria do not import tRNA, but a number of neuromuscular degenerative and metabolic diseases are caused by mutations in mitochondrial tRNA genes (21). In yeast, a single tRNA is imported, apparently through protein import channels and requiring at least two soluble factors, including the mitochondrial form of the cognate aminoacyl-tRNA synthetase (8). By contrast, in kinetoplastid protozoa (Leishmania and trypanosomes), import of a whole spectrum of tRNAs is necessitated by the complete lack of mitochondrial tRNA genes (5,19). In this system, membrane-bound tRNA binding proteins recognize specific structural motifs (import signals) on tRNA, soluble factors are not required, and the translocation pathway appears to be distinct from that for protein import (11,14,17). Moreover, the sequence and bioenergetic requirements for outer and inner membrane transfer are nonidentical (2), indicating the presence of a distinct transport machinery (the RNA import complex [RIC]) at the inner membrane, a situation similar to the TOM and TIM complexes for protein import (15). A 15-kDa polypeptide has been shown to be required for import into Leishmania mitochondria (1); otherwise, the import machinery remains undefined.Using an in vitro evolution protocol, it was recently shown that Leishmania mitochondria recognize a number of short sequence motifs homologous to multiple domains in tRNAs, suggesting the presence of several import signals (3). Moreover, novel positive and negative allosteric interactions between these aptamers, as well as between intact tRNAs, at the inner membrane were described (3). The RNAs could be classified into two types: type I RNAs are efficiently transferred through the inner membrane but are inhibited by type II. In contrast, type II RNAs have poor inner membrane transfer efficiencies and are stimulated by type I. For example, tRNA Tyr (GUA) is a type I RNA containing the conserved motif UA GAGC in the D domain, while tRNA Ile (UAU) is type II with the sequence UCGCGGGUU in the variable loop-T domain (V-T) region (3). The mechanism of these allosteric interactions is unknown, but there are several possibilities. A single conformationally flexible dimeric or multimeric receptor could bind t...
Abstract-Performance of regularized least-squares estimation in noisy compressed sensing is analyzed in the limit when the dimensions of the measurement matrix grow large. The sensing matrix is considered to be from a class of random ensembles that encloses as special cases standard Gaussian, roworthogonal, geometric and so-called T -orthogonal constructions. Source vectors that have non-uniform sparsity are included in the system model. Regularization based on 1-norm and leading to LASSO estimation, or basis pursuit denoising, is given the main emphasis in the analysis. Extensions to 2-norm and "zero-norm" regularization are also briefly discussed. The analysis is carried out using the replica method in conjunction with some novel matrix integration results. Numerical experiments for LASSO are provided to verify the accuracy of the analytical results.The numerical experiments show that for noisy compressed sensing, the standard Gaussian ensemble is a suboptimal choice for the measurement matrix. Orthogonal constructions provide a superior performance in all considered scenarios and are easier to implement in practical applications. It is also discovered that for non-uniform sparsity patterns the T -orthogonal matrices can further improve the mean square error behavior of the reconstruction when the noise level is not too high. However, as the additive noise becomes more prominent in the system, the simple row-orthogonal measurement matrix appears to be the best choice out of the considered ensembles.
A large number of cytoplasmic tRNAs are imported into the kinetoplast-mitochondrion of Leishmania by a receptor-mediated process. To identify the sequences recognized by import receptors, mitochondria were incubated with a combinatorial RNA library. Repeated cycles of amplification of the imported sequences (SELEX) resulted in rapid selection of several import aptamers containing sequence motifs present in the anticodon arm, the D arm, the V-T region, and acceptor stem of known tRNAs, confirming or suggesting the presence of import signals in these domains. As predicted, truncated derivatives of tRNA Ile (UAU) containing the D arm or the V-T region were imported in vitro. Four aptamers were studied in detail. All were imported in vitro as well as in transiently transfected cells, using the same pathway as tRNA, but their individual import efficiencies were different. Two types of aptamers were discernible: the A arm and D arm homologues (type I), which were efficiently transferred across the inner mitochondrial membrane, and the V-T homologues (type II), which were not. Remarkably, subnanomolar concentrations of type I RNAs stimulated the entry of type II RNAs into the matrix, whereas type II RNAs inhibited inner membrane transfer of type I RNAs. Moreover, tRNA Tyr (GUA) and tRNA Ile (UAU) interacted with one another as type I and type II, respectively. Such cooperative and antagonistic interactions may allow the use of a limited number of receptors to recognize a large number of tRNAs of variable affinity and enable the maintenance of a properly balanced tRNA pool for mitochondrial translation.
Abstract-In a Bayesian linear model, suppose observation y = Hx+n stems from independent inputs x and n which are Gaussian mixture (GM) distributed. With known matrix H, the minimum mean square error (MMSE) estimator for x, has analytical form. However, its performance measure, the MMSE itself, has no such closed form. Because existing Bayesian MMSE bounds prove to have limited practical value under these settings, we instead seek analytical bounds for the MMSE, both upper and lower. This paper provides such bounds, and relates them to the signal-to-noise-ratio (SNR).
For reconstruction of low-rank matrices from undersampled measurements, we develop an iterative algorithm based on least-squares estimation. While the algorithm can be used for any low-rank matrix, it is also capable of exploiting a-priori knowledge of matrix structure. In particular, we consider linearly structured matrices, such as Hankel and Toeplitz, as well as positive semidefinite matrices. The performance of the algorithm, referred to as alternating least-squares (ALS), is evaluated by simulations and compared to the Cramer-Rao bounds. QC 20120411
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