Hot off the presses! Jan 07 mol cell

The Jan 07 issue of the mol cell is now up on Pubget (About mol cell): if you're at a subscribing institution, just click the link in the latest link at the home page. (Note you'll only be able to get all the PDFs in the issue if your institution subscribes to Pubget.)

Latest Articles Include:

• New Partners for HP1 in Transcriptional Gene Silencing
  - mol cell 41(1):1-2 (2011)
  A new study in this issue of Molecular Cell (Yamane et al., 2010) demonstrates how chromatin assembly proteins HIRA/Asf1 help enforce transcriptional gene silencing in heterochromatin by bridging interactions between HP1 and histone deacetylase complexes.
• San1-Mediated Quality Control: Substrate Recognition "Sans" Chaperones
  - mol cell 41(1):2-3 (2011)
  In this issue of Molecular Cell, Rosenbaum et al. describe a mechanism that allows San1 to selectively detect misfolded proteins for nuclear protein quality control.
• Bacteria as Control Engineers
  - mol cell 41(1):4-5 (2011)
  Bacteria encounter fluctuations in both their external and internal environments, and to manage these conditions, they employ various control mechanisms. In this issue of Molecular Cell, Hart et al. (2011) investigate how E. coli robustly controls nitrogen assimilation.
• P-Rex1, a Guanine Exchange Factor that Is Overexpressed in Breast Cancer, Is a Convergence Node for ErbB and CXCR4 Signaling
  - mol cell 41(1):5-7 (2011)
  In a recent issue of Molecular Cell, Kazanietz and colleagues (Sosa et al., 2010) show that P-Rex1, a Rac1 GEF, is overexpressed in ER+ and/or ErbB2+ breast cancers, suggesting that P-Rex1 might be a convergence node downstream of these receptors and an attractive therapeutic target.
• Proteasome Activators
  - mol cell 41(1):8-19 (2011)
  Proteasomes degrade a multitude of protein substrates in the cytosol and nucleus, and thereby are essential for many aspects of cellular function. Because the proteolytic sites are sequestered in a closed barrel-shaped structure, activators are required to facilitate substrate access. Structural and biochemical studies of two activator families, 11S and Blm10, have provided insights to proteasome activation mechanisms, although the biological functions of these factors remain obscure. Recent advances have improved our understanding of the third activator family, including the 19S activator, which targets polyubiquitylated proteins for degradation. Here we present a structural perspective on how proteasomes are activated and how substrates are delivered to the proteolytic sites.
• Yeast Sen1 Helicase Protects the Genome from Transcription-Associated Instability
  - mol cell 41(1):21-32 (2011)
  Sen1 of S. cerevisiae is a known component of the NRD complex implicated in transcription termination of nonpolyadenylated as well as some polyadenylated RNA polymerase II transcripts. We now show that Sen1 helicase possesses a wider function by restricting the occurrence of RNA:DNA hybrids that may naturally form during transcription, when nascent RNA hybridizes to DNA prior to its packaging into RNA protein complexes. These hybrids displace the nontranscribed strand and create R loop structures. Loss of Sen1 results in transient R loop accumulation and so elicits transcription-associated recombination. SEN1 genetically interacts with DNA repair genes, suggesting that R loop resolution requires proteins involved in homologous recombination. Based on these findings, we propose that R loop formation is a frequent event during transcription and a key function of Sen1 is to prevent their accumulation and associated genome instability.
• PARP-3 and APLF Function Together to Accelerate Nonhomologous End-Joining
  - mol cell 41(1):33-45 (2011)
  PARP-3 is a member of the ADP-ribosyl transferase superfamily of unknown function. We show that PARP-3 is stimulated by DNA double-strand breaks (DSBs) in vitro and functions in the same pathway as the poly (ADP-ribose)-binding protein APLF to accelerate chromosomal DNA DSB repair. We implicate PARP-3 in the accumulation of APLF at DSBs and demonstrate that APLF promotes the retention of XRCC4/DNA ligase IV complex in chromatin, suggesting that PARP-3 and APLF accelerate DNA ligation during nonhomologous end-joining (NHEJ). Consistent with this, we show that class switch recombination in Aplf−/− B cells is biased toward microhomology-mediated end-joining, a pathway that operates in the absence of XRCC4/DNA ligase IV, and that the requirement for PARP-3 and APLF for NHEJ is circumvented by overexpression of XRCC4/DNA ligase IV. These data identify molecular roles for PARP-3 and APLF in chromosomal DNA double-strand break repair reactions.
• DNA Repair Factor APLF Is a Histone Chaperone
  - mol cell 41(1):46-55 (2011)
  Poly(ADP-ribosyl)ation plays a major role in DNA repair, where it regulates chromatin relaxation as one of the critical events in the repair process. However, the molecular mechanism by which poly(ADP-ribose) modulates chromatin remains poorly understood. Here we identify the poly(ADP-ribose)-regulated protein APLF as a DNA-damage-specific histone chaperone. APLF preferentially binds to the histone H3/H4 tetramer via its C-terminal acidic motif, which is homologous to the motif conserved in the histone chaperones of the NAP1L family (NAP1L motif). We further demonstrate that APLF exhibits histone chaperone activities in a manner that is dependent on its acidic domain and that the NAP1L motif is critical for the repair capacity of APLF in vivo. Finally, we identify structural analogs of APLF in lower eukaryotes with the ability to bind histones and localize to the sites of DNA-damage-induced poly(ADP-ribosyl)ation. Collectively, these findings define the involvement of ! histone chaperones in poly(ADP-ribose)-regulated DNA repair reactions.
• Asf1/HIRA Facilitate Global Histone Deacetylation and Associate with HP1 to Promote Nucleosome Occupancy at Heterochromatic Loci
  - mol cell 41(1):56-66 (2011)
  Heterochromatin impacts various nuclear processes by providing a recruiting platform for diverse chromosomal proteins. In fission yeast, HP1 proteins Chp2 and Swi6, which bind to methylated histone H3 lysine 9, associate with SHREC (Snf2/HDAC repressor complex) and Clr6 histone deacetylases (HDACs) involved in heterochromatic silencing. However, heterochromatic silencing machinery is not fully defined. We describe a histone chaperone complex containing Asf1 and HIRA that spreads across silenced domains via its association with Swi6 to enforce transcriptional silencing. Asf1 functions in concert with a Clr6 HDAC complex to silence heterochromatic repeats, and it suppresses antisense transcription by promoting histone deacetylation. Furthermore, we demonstrate that Asf1 and SHREC facilitate nucleosome occupancy at heterochromatic regions but TFIIIC transcription factor binding sites within boundary elements are refractory to these factors. These analyses uncover a role f! or Asf1 in global histone deacetylation and suggest that HP1-associated histone chaperone promotes nucleosome occupancy to assemble repressive heterochromatin.
• Chromodomain-Mediated Oligomerization of HP1 Suggests a Nucleosome-Bridging Mechanism for Heterochromatin Assembly
  - mol cell 41(1):67-81 (2011)
  HP1 proteins are central to the assembly and spread of heterochromatin containing histone H3K9 methylation. The chromodomain (CD) of HP1 proteins specifically recognizes the methyl mark on H3 peptides, but the same extent of specificity is not observed within chromatin. The chromoshadow domain of HP1 proteins promotes homodimerization, but this alone cannot explain heterochromatin spread. Using the S. pombe HP1 protein, Swi6, we show that recognition of H3K9-methylated chromatin in vitro relies on an interface between two CDs. This interaction causes Swi6 to tetramerize on a nucleosome, generating two vacant CD sticky ends. On nucleosomal arrays, methyl mark recognition is highly sensitive to internucleosomal distance, suggesting that the CD sticky ends bridge nearby methylated nucleosomes. Strengthening the CD-CD interaction enhances silencing and heterochromatin spread in vivo. Our findings suggest that recognition of methylated nucleosomes and HP1 spread on chromati! n are structurally coupled and imply that methylation and nucleosome arrangement synergistically regulate HP1 function.
• Cdc48/p97 Mediates UV-Dependent Turnover of RNA Pol II
  - mol cell 41(1):82-92 (2011)
  Cdc48/p97 is an essential ATPase whose role in targeting substrates to the ubiquitin-proteasome system (UPS) remains unclear. Existing models posit that Cdc48 acts upstream of UPS receptors. To address this hypothesis, we examined the association of ubiquitin (Ub) conjugates with 26S proteasomes. Unexpectedly, proteasomes isolated from cdc48 mutants contain high levels of Ub conjugates, and mass spectrometry identified numerous nonproteasomal proteins, including Rpb1, the largest subunit of RNA Pol II. UV-induced turnover of Rpb1 depends upon Cdc48-Ufd1-Npl4, Ubx4, and the uncharacterized adaptor Ubx5. Ubiquitinated Rpb1, proteasomes, and Cdc48 accumulate on chromatin in UV-treated wild-type cells, and the former two accumulate to higher levels in mutant cells, suggesting that degradation of Rpb1 is facilitated by Cdc48 at sites of stalled transcription. These data reveal an intimate coupling of function between proteasomes and Cdc48 that we suggest is necessary to sus! tain processive degradation of unstable subunits of some macromolecular protein complexes.
• Disorder Targets Misorder in Nuclear Quality Control Degradation: A Disordered Ubiquitin Ligase Directly Recognizes Its Misfolded Substrates
  - mol cell 41(1):93-106 (2011)
  Protein quality control (PQC) degradation systems protect the cell from the toxic accumulation of misfolded proteins. Because any protein can become misfolded, these systems must be able to distinguish abnormal proteins from normal ones, yet be capable of recognizing the wide variety of distinctly shaped misfolded proteins they are likely to encounter. How individual PQC degradation systems accomplish this remains an open question. Here we show that the yeast nuclear PQC ubiquitin ligase San1 directly recognizes its misfolded substrates via intrinsically disordered N- and C-terminal domains. These disordered domains are punctuated with small segments of order and high sequence conservation that serve as substrate-recognition sites San1 uses to target its different substrates. We propose that these substrate-recognition sites, interspersed among flexible, disordered regions, provide San1 an inherent plasticity which allows it to bind its many, differently shaped misfold! ed substrates.
• ARTS and Siah Collaborate in a Pathway for XIAP Degradation
  - mol cell 41(1):107-116 (2011)
  ARTS (apoptosis-related protein in the TGF-β signaling pathway) is a mitochondrial protein that binds XIAP (X-linked inhibitor of apoptosis protein) upon entering the cytosol, thus promoting cell death. Expression of ARTS is lost in some malignancies. Here, we show that ARTS binds to XIAP at BIR1, a domain distinct from the caspase-binding sites. Furthermore, ARTS interacts with the E3 ligase Siah-1 (seven in absentia homolog 1) to induce ubiquitination and degradation of XIAP. Cells lacking either Siah or ARTS contain higher steady-state levels of XIAP. Thus, ARTS serves as an adaptor to bridge Siah-1 to XIAP, targeting it for destruction.
• Robust Control of Nitrogen Assimilation by a Bifunctional Enzyme in E. coli
  - mol cell 41(1):117-127 (2011)
  Bacteria regulate the assimilation of multiple nutrients to enable growth. How is balanced utilization achieved, despite fluctuations in the concentrations of the enzymes that make up the regulatory circuitry? Here we address this question by studying the nitrogen system of E. coli. A mechanism based on the avidity of a bifunctional enzyme, adenylyltransferase (AT/AR), to its multimeric substrate, glutamine synthetase, is proposed to maintain a robust ratio between two key metabolites, glutamine and α-ketoglutarate. This ratio is predicted to be insensitive to variations in protein levels of the core circuit and to the rate of nitrogen utilization. We find using mass spectrometry that the metabolite ratio is robust to variations in protein levels and that this robustness depends on the bifunctional enzyme. Moreover, robustness carries through to the bacteria growth rate. Interrupting avidity by adding a monofunctional AT/AR mutant to the native system abolishes robust! ness, as predicted by the proposed mechanism.