Small RNAs of 20-30 nucleotides can target both chromatin and transcripts, and thereby keep both the genome and the transcriptome under extensive surveillance. Recent progress in high-throughput sequencing has uncovered an astounding landscape of small RNAs in eukaryotic cells. Various small RNAs of distinctive characteristics have been found and can be classified into three classes based on their biogenesis mechanism and the type of Argonaute protein that they are associated with: microRNAs (miRNAs), endogenous small interfering RNAs (endo-siRNAs or esiRNAs) and Piwi-interacting RNAs (piRNAs). This Review summarizes our current knowledge of how these intriguing molecules are generated in animal cells.

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Biogenesis of small RNAs in animals.

Since the discovery in 1993 of the first small silencing RNA, a dizzying number of small RNA classes have been identified, including microRNAs (miRNAs), small interfering RNAs (siRNAs) and Piwi-interacting RNAs (piRNAs). These classes differ in their biogenesis, their modes of target regulation and in the biological pathways they regulate. There is a growing realization that, despite their differences, these distinct small RNA pathways are interconnected, and that small RNA pathways compete and collaborate as they regulate genes and protect the genome from external and internal threats.

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Small silencing RNAs: an expanding universe

The canonical role of messenger RNA (mRNA) is to deliver protein-coding information to sites of protein synthesis. However, given that microRNAs bind to RNAs, we hypothesized that RNAs could possess a regulatory role that relies on their ability to compete for microRNA binding, independently of their protein-coding function. As a model for the protein-coding-independent role of RNAs, we describe the functional relationship between the mRNAs produced by the PTEN tumour suppressor gene and its pseudogene PTENP1 and the critical consequences of this interaction. We find that PTENP1 is biologically active as it can regulate cellular levels of PTEN and exert a growth-suppressive role. We also show that the PTENP1 locus is selectively lost in human cancer. We extended our analysis to other cancer-related genes that possess pseudogenes, such as oncogenic KRAS. We also demonstrate that the transcripts of protein-coding genes such as PTEN are biologically active. These findings attribute a novel biological role to expressed pseudogenes, as they can regulate coding gene expression, and reveal a non-coding function for mRNAs.

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A coding-independent function of gene and pseudogene mRNAs regulates tumour biology

PIWI-interacting RNAs (piRNAs) are a distinct class of small non-coding RNAs that form the piRNA-induced silencing complex (piRISC) in the germ line of many animal species. The piRISC protects the integrity of the genome from invasion by 'genomic parasites' — transposable elements — by silencing them. Owing to their limited expression in gonads and their sequence diversity, piRNAs have been the most mysterious class of small non-coding RNAs regulating RNA silencing. Now, much progress is being made into our understanding of their biogenesis and molecular functions, including the specific subcellular compartmentalization of the piRNA pathway in granular cytoplasmic bodies.

PIWI-interacting small RNAs: the vanguard of genome defence.

To gain insight into how genomic information is translated into cellular and developmental programs, the Drosophila model organism Encyclopedia of DNA Elements (modENCODE) project is comprehensively mapping transcripts, histone modifications, chromosomal proteins, transcription factors, replication proteins and intermediates, and nucleosome properties across a developmental time course and in multiple cell lines. We have generated more than 700 data sets and discovered protein-coding, noncoding, RNA regulatory, replication, and chromatin elements, more than tripling the annotated portion of the Drosophila genome. Correlated activity patterns of these elements reveal a functional regulatory network, which predicts putative new functions for genes, reveals stage- and tissue-specific regulators, and enables gene-expression prediction. Our results provide a foundation for directed experimental and computational studies in Drosophila and related species and also a model for systematic data integration toward comprehensive genomic and functional annotation.

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Identification of Functional Elements and Regulatory Circuits by Drosophila modENCODE

A broadly conserved pathway generates 3'UTR-directed primary piRNAs.

Two distinct mechanisms generate endogenous siRNAs from bidirectional transcription in Drosophila melanogaster

Two distinct mechanisms generate endogenous siRNAs from bidirectional transcription in Drosophila melanogaster.

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Article A Broadly Conserved Pathway Generates 3 0 UTR-Directed Primary piRNAs

The multiplex polymerase chain reaction (MP-PCR) is a quick and inexpensive technique in molecular biology for amplifying multiple DNA loci in a single Polymerase Chain Reaction (PCR). One of the criteria to achieve highly specific reaction products is to keep the concentration of the amplification primers low. In research, the dilemma associated with primer minimization for MP-PCR reactions has been formulated as the Multiple Degenerate Primer Selection Problem (MDPSP). MDPSP is related to the earlier Degenerate Primer Design (DPD) problem that has proven to be NP-complete. This paper formulates a new, so far, unexplored variant, the Multiple Degenerate Primer Selection Problem with Errors (MDPSPE) and introduces new algorithms for solving this new version. Furthermore, we implement an exact algorithm, DPS-HDR for solving the earlier MDPSP and compare the algorithm's performance on randomly generated data sets with DPS-HD, thus far the most efficient algorithm for solving MDPSP introduced in [2]. We expect to reduce the execution time of the algorithm in comparison to DPS-HD.1

Sudha Balla

Papers

A broadly conserved pathway generates 3'UTR-directed primary piRNAs.

Two distinct mechanisms generate endogenous siRNAs from bidirectional transcription in Drosophila melanogaster.

Article A Broadly Conserved Pathway Generates 3 0 UTR-Directed Primary piRNAs

Degenerate primer selection algorithms