Darwin 200

The 200th anniversary of the birth of Charles Robert Darwin falls on 12 February 2009. No single researcher has since matched his collective impact on the natural and social sciences; on politics, religions, and philosophy; on art and cultural relations. In this landmark year, our Nature news special provides continuously updated news, research and analysis on Darwin's life, his science and his legacy.

http://www.nature.com/news/specials/darwin/index.html

Toward minimal bacterial cells: evolution vs. design.

Andrés Moya 1,2,3 , Rosario Gil 1,2,3 , Amparo Latorre 1,2,3 , Juli Peretó 1,2,4 , Maria Pilar Garcillán-Barcia 5 & Fernando de la Cruz 5
1 Institut Cavanilles de Biodiversitat i Biologia Evolutiva, Universitat de València, València, Spain; 2 CIBER de Epidemiología y Salud Pública, Madrid, Spain; 3 Departament de Genètica, Universitat de València, València, Spain; 4 Departament de Bioquímica i Biologia Molecular, Universitat de València, València, Spain; and 5 Departamento de Biología Molecular, Universidad de Cantabria, Santander, Spain

Recent technical and conceptual advances in the biological sciences opened the possibility of the construction of newly designed cells. In this paper we review the state of the art of cell engineering in the context of genome research, paying particular attention to what we can learn on naturally reduced genomes from either symbiotic or free living bacteria. Different minimal hypothetically viable cells can be defined on the basis of several computational and experimental approaches. Projects aiming at simplifying living cells converge with efforts to make synthetic genomes for minimal cells. The panorama of this particular view of synthetic biology lead us to consider the use of defined minimal cells to be applied in biomedical, bioremediation, or bioenergy application by taking advantage of existing naturally minimized cells.

http://dx.doi.org/10.1111/j.1574-6976.2008.00151.x

Coprinus cinereus rad50 Mutants Reveal an Essential Structural Role for Rad50 in Axial Element and Synaptonemal Complex Formation, Homolog Pairing and

Sonia N. Acharya*,1, Alexander M. Many*,1,2, Andrew P. Schroeder*, Felicia M. Kennedy*, Oleksandr P. Savytskyy*, Jennifer T. Grubb*, Jack A. Vincent*, Elizabeth A. Friedle*, Martina Celerin*, Daniel S. Maillet*, Heather J. Palmerini*, Megan A. Greischar*, Gabriel Moncalian,3, R. Scott Williams, John A. Tainer and Miriam E. Zolan*,4
* Department of Biology, Indiana University, Bloomington, Indiana 47405 and The Scripps Research Institute, La Jolla, California 92037

The Mre11/Rad50/Nbs1 (MRN) complex is required for eukaryotic DNA double-strand break (DSB) repair and meiotic recombination. We cloned the Coprinus cinereus rad50 gene and showed that it corresponds to the complementation group previously named rad12, identified mutations in 15 rad50 alleles, and mapped two of the mutations onto molecular models of Rad50 structure. We found that C. cinereus rad50 and mre11 mutants arrest in meiosis and that this arrest is Spo11 dependent. In addition, some rad50 alleles form inducible, Spo11-dependent Rad51 foci and therefore must be forming meiotic DSBs. Thus, we think it likely that arrest in both mre11-1 and the collection of rad50 mutants is the result of unrepaired or improperly processed DSBs in the genome and that Rad50 and Mre11 are dispensable in C. cinereus for DSB formation, but required for appropriate DSB processing. We found that the ability of rad50 mutant strains to form Rad51 foci correlates with their ability to promote synaptonemal complex formation and with levels of stable meiotic pairing and that partial pairing, recombination initiation, and synapsis occur in the absence of wild-type Rad50 catalytic domains. Examination of single- and double-mutant strains showed that a spo11 mutation that prevents DSB formation enhances axial element (AE) formation for rad50-4, an allele predicted to encode a protein with intact hook region and hook-proximal coiled coils, but not for rad50-1, an allele predicted to encode a severely truncated protein, or for rad50-5, which encodes a protein whose hook-proximal coiled-coil region is disrupted. Therefore, Rad50 has an essential structural role in the formation of AEs, separate from the DSB-processing activity of the MRN complex.

http://dx.doi.org/10.1534/genetics.108.092775%20

Analysis of ColE1 MbeC unveils an extended ribbon-helix-helix family of nicking-accessory proteins

Athanasia Varsaki, Gabriel Moncalián, Maria del Pilar Garcillán-Barcia, Constantin Drainas, and Fernando de la Cruz*
MbeC is a 13 kDa ColE1-encoded protein required for efficient mobilization of ColE1, a plasmid widely used in cloning vector technology. MbeC protein was purified and used for in vitro DNA-binding, which showed that it binds specifically dsDNA containing the ColE1 oriT. Amino-acid sequence comparison and secondary structure prediction imply that MbeC is related to the ribbon-helix-helix (RHH) protein family. Alignment with RHH members pointed a conserved arginine (R13 in MbeC) which was mutated to alanine. The mutant MbeC(R13A) was unable to bind either ssDNA or dsDNA. Limited proteolysis fragmented MbeC in two stable folding domains: the N-terminal domain, which contains the RHH motif and the C-terminal domain, which comprises a signature shared by nicking-accessory proteins. Results indicate that MbeC plays a similar role in conjugation as TraY and TrwA of plasmids F and R388, respectively. Thus, it appears that an extended, possibly universal mechanism of DNA conjugative processing exists, in which oriT-processing is carried out by relaxases assisted by homologous nicking-accessory proteins. This mechanism seems to be shared by all major conjugative systems analyzed so far.
http://dx.doi.org/10.1128/JB.01342-08

Reply to The binding stoichiometry of CIN85 SH3 domain A and Cbl-b

Daniela Jozic, Nayra Cardenes, Yonathan Lissanu Deribe, Gabriel Moncalian, Daniela Hoeller, Yvonne Groemping, Ivan Dikic, Katrin Rittinger, Jeronimo Bravo
Nature Structural & Molecular Biology 15, 891 - 892 (01 Sep 2008), doi: http://dx.doi.org/10.1038/nsmb0908-891, Correspondence

Olimpiada de Biología - Departamento de Biología Molecular

El Departamento de Biología Molecular http://departamentos.unican.es/biomol/ (Tel. 942 20 19 46) se ocupa de la organización de la fase local de la IV Olimpiada de Biología dirigida a estudiantes de Bachillerato.

http://www.olimpiadadebiologia.edu.es/

Biotechnology Journal 2008: Changing the recognition site of a conjugative relaxase by rational design

Blanca González-Pérez, José Daniel Carballeira, Gabriel Moncalián, Fernando de la Cruz.
Abstract
TrwC is a relaxase protein, which starts and finishes DNA processing during bacterial conjugation in plasmid R388. TrwC recognizes a specific sequence of DNA (25 nucleotides) in the donor cell: the nic-site. As a model example, a single transversion C24G in nic avoids DNA processing by TrwC. Using this simple model, our objective was to obtain a proof of principle that TrwC specificity can be changed. Several structures of DNA-TrwC complexes were used as reference to design a focused saturation mutagenesis library (NNK) randomizing amino acid Lys262, since its side chain seems to sterically hinder the recognition of the C24G nic mutation by wild-type TrwC. Using bacterial conjugation as an in vivo selection system, several TrwC variants were found that show changes in substrate specificity. These variants were also tested in a competitive assay to evaluate their conjugation efficiency.

http://dx.doi.org/10.1002/biot.200800184

Un investigador de la UC publica en 'Cell'

Un investigador del Instituto de Biomedicina y Biotecnología (IBBTEC) de la Universidad de Cantabria (UC) ha publicado en la revista científica 'Cell' -la más prestigiosa del mundo en el campo de la biomedicina- un trabajo que aporta nuevos detalles sobre la reparación del ADN, un proceso que resulta fundamental para la supervivencia de las células. Gabriel Moncalián Montes, investigador del programa 'Ramón y Cajal', y los otros trece coautores del descubrimiento describen en su artículo la forma en la que una proteína llamada Mre11 actúa en el proceso de reparación de las moléculas de ADN. El trabajo (cuyo título original es 'Mre11 Dimers Coordinate DNA End Bridging and Nuclease Processing in Double-Strand-Break Repair') explica que la Mre11 se convierte en una proteína compuesta por dos unidades, y que con esa estructura doble la proteína replica los trozos de ADN para restaurar la cadena. Moncalián lidera el subgrupo de investigación 'Ingeniería de Proteínas' del IBBTEC, del que forman parte Yera Tena, Juan Villa, Blanca González y Matilde Cabezas.

http://www.eldiariomontanes.es/20081005/cantabria/noticias/investigador-publica-cell-20081005.html

Cell 2008: Mre11 Dimers Coordinate DNA End Bridging and Nuclease Processing in Double-Strand-Break Repair .

R . Williams* , G . Moncalian* , J . Williams , Y . Yamada , O . Limbo , D . Shin , L . Groocock , D . Cahill , C . Hitomi , G . Guenther. (* these authors contribute equally)

Mre11 forms the core of the multifunctional Mre11-Rad50-Nbs1 (MRN) complex that detects DNA double-strand breaks (DSBs), activates the ATM checkpoint kinase, and initiates homologous recombination (HR) repair of DSBs. To define the roles of Mre11 in both DNA bridging and nucleolytic processing during initiation of DSB repair, we combined small-angle X-ray scattering (SAXS) and crystal structures of Pyrococcus furiosus Mre11 dimers bound to DNA with mutational analyses of fission yeast Mre11. The Mre11 dimer adopts a four-lobed U-shaped structure that is critical for proper MRN complex assembly and for binding and aligning DNA ends. Further, mutations blocking Mre11 endonuclease activity impair cell survival after DSB induction without compromising MRN complex assembly or Mre11-dependant recruitment of Ctp1, an HR factor, to DSBs. These results show how Mre11 dimerization and nuclease activities initiate repair of DSBs and collapsed replication forks, as well as provide a molecular foundation for understanding cancer-causing Mre11 mutations in ataxia telangiectasia-like disorder (ATLD). http://dx.doi.org/10.1016/j.cell.2008.08.017

Diario Médico

Europa press

Journal Bacteriology, 2008 ATPase activity and oligomeric state of TrwK, the VirB4 homologue of plasmid R388 Type IV secretion system.

Ignacio Arechaga, Alejandro Peña, Sandra Zunzunegui, María del Carmen Fernández-Alonso, Germán Rivas and Fernando de la Cruz.

Type IV secretion systems (T4SS) mediate the transfer of DNA and protein substrates to target cells. TrwK, encoded by the conjugative plasmid R388, is a member of the VirB4 family, the largest and most conserved protein of T4SS. VirB4 was suggested to be an ATPase involved in energizing pilus assembly and substrate transport. However, conflicting experimental evidence concerning VirB4 ATP hydrolase activity was reported. Here, we demonstrate that TrwK is able to hydrolyze ATP in vitro in the absence of its potential macromolecular substrates and other T4SS components. The kinetic parameters of its ATPase activity have been characterized. TrwK oligomerization state was investigated by analytical ultracentrifugation and electron microscopy, and its effects on the ATPase activity were analyzed. The results suggest that the hexameric form of TrwK is the catalytically active state, much like the structurally related protein TrwB, the conjugative coupling protein. http://dx.doi.org/10.1128/JB.00321-08