Volume 20 Issue 12, December 2013

An unresolved question in mammalian epigenetic regulation is how ubiquitously expressed chromatin-modifying complexes such as Polycomb group complex 2 (PRC2) find their specific target sites across an intricate choreography of localization events in time and space. Two recent studies now provide critical new insights into an intriguing genome-wide role for RNA in PRC2 regulation.

Mediator of DNA-damage checkpoint 1 (MDC1) has a central role in repair of DNA double-strand breaks (DSBs) by both homologous recombination and nonhomologous end joining, and its function is regulated by post-translational phosphorylation, ubiquitylation and SUMOylation. In this issue, a new study by Watanabe et al. reveals that methylation of MDC1 is also critical for its function in DSB repair and specifically affects repair through BRCA1-dependent homologous recombination.

Faithful bypass of replication forks encountering obstructive DNA lesions is essential to prevent fork collapse and cell death. PrimPol is a new human primase and translesion polymerase that is able to bypass fork-blocking UV-induced lesions and to restart replication by origin-independent repriming.

HIV-1 Env glycoprotein binds receptors on the host cells, triggering a conformational change from a closed to an open state. Now single-particle cryo-EM analysis of a soluble, trimeric Env construct reveals the structure of the closed state at ∼6-Å resolution. The structure features three gp41 helices at the center of the trimer, thus indicating that HIV-1 and influenza viruses use similar mechanisms to enter the cell.

The nuclear cap–binding complex (CBC) stimulates RNA maturation, but the mechanistic basis is not well understood. In vitro reconstitution experiments combined with functional analyses have revealed a new CBC complex containing ARS2, a major effector of CBC. ARS2 links the cap to 3'-end maturation for several RNA families, thus favoring the production of short RNAs.

How the nuclear exosome is targeted to nuclear RNA substrates is poorly understood. An affinity-capture MS approach and functional analyses now demonstrate a physical and functional connection between the human exosome and the cap-binding complex (CBC). A CBC-containing complex was found to promote transcription termination of several RNA types, thus suggesting a direct link to exosomal RNA degradation.

Pentatricopeptide repeat (PPR) proteins are involved in different RNA processes, but how they recognize their target RNAs has been unclear. Now crystal structures of plant PPR protein–THA8 in complex with RNA, along with functional analyses, reveal an asymmetric THA8 dimer with RNA bound at the dimeric interface.

DNA damage that causes replication forks to stall can be bypassed via translesion synthesis (TLS). New work has identified a bifunctional human primase and TLS polymerase, PrimPol, that reinitiates DNA synthesis beyond the damage site by virtue of its unique primase activity, revealing a novel pathway of DNA-damage tolerance.

Using 13 intermediate-translocation-state models derived from X-ray and cryo-EM structures of Escherichia coli ribosomes to guide large-scale molecular dynamics simulations, a new study now models the path taken by tRNAs during spontaneous translocation to uncover the mechanisms that facilitate tRNA movement through the ribosome.

Centromere identity is conferred epigenetically by incorporation of the histone H3 variant CENP-A. A new study shows that incorporation of histone H2A variant H2A.Z prevents ectopic stabilization of neocentromeres in Schizosaccharomyces pombe, suggesting that de novo establishment of centromeres is also epigenetically controlled.

Through its association with Cdk–cyclin complexes, Cks has been implicated in the multisite phosphorylation of numerous cell cycle–regulatory proteins. A structural analysis of Cks1 bound to a target phosphopeptide, combined with binding-specificity studies, now reveals a Cks-binding consensus sequence and how Cks1 confers phosphodependent substrate specificity to Cdk1.

Selective protein phosphorylation in a time-resolved manner is essential to an ordered progression through the cell cycle. A new study now shows how the phopshoadaptor protein Cks1 acts as a signal processor for Cdk1 by regulating the differential amplification of multiple phosphorylation events to generate a wide range of thresholds during the cell cycle.

Although the DNA damage–induced ubiquitylation by RNF8 and/or RNF168 ubiquitin ligases is crucial for the DNA-damage response (DDR), the precise mechanisms of ubiquitylation-mediated chromatin modulation and recruitment of DNA-repair proteins 53BP1 and RAP80–BRCA1 are not fully understood. A new study now indicates that human demethylase JMJD1C regulates the RAP80–BRCA1 branch of this DDR pathway.

The Rbfox family of developmentally important splicing factors controls alternative splicing in a tissue-specific manner. Genome-wide analyses now show that more than half of Rbfox-binding sites are located distally from exons, that these distal sites are conserved and functionally important, and that long-range RNA-RNA secondary structures mediate distal splicing regulation by Rbfox.

TDP-43 regulates alternative splicing of the cystic fibrosis transmembrane regulator (CFTR) and is found in cytosolic granules associated with several neurodegenerative disorders. A new solution structure of the tandem RNA-recognition motifs (RRMs) that mediate interactions with its UG-rich RNA targets reveals a new RRM arrangement critical for TDP-43 function.

Single-molecule FRET studies of pre-mRNA dynamics within the spliceosome offer new insights into the mechanism of splicing by showing that Prp2 juxtaposes the 5' splice site and branch point elements, and that splicing factor Cwc25 enhances first step catalysis by stabilizing this proximal conformation.

A transcription activator–like effector nuclease (TALEN)-mediated knockout approach to delete human microRNA (miRNA) genes was used to generate a library of 540 TALEN pairs for 274 miRNA loci. As a case study, single and double knockouts for two related miRNAs, miR-141 and miR-200c, revealed intriguing functional differences.