BAF57 governs androgen receptor action and androgen-dependent proliferation through SWI/SNF

Androgen receptor (AR) activity is required for prostate cancer development and progression. Thus, there is a major impetus to understand the regulation of AR action. We and others have previously shown that AR transactivation potential is dependent on the presence of an active SWI/SNF chromatin remodeling complex. However, the mechanisms underlying SWI/SNF regulation of the AR remained unsolved. We show here that the BAF57 subunit, an accessory component of the remodeling complex, is a critical regulator of AR function. We show that BAF57 is expressed in the luminal epithelia of the prostate and is required for AR-dependent transactivation in prostatic adenocarcinoma cells. Our data reveal that BAF57 can directly bind to the AR and is recruited to endogenous AR targets upon ligand activation. Loss of BAF57 or inhibition of BAF57 function severely compromised AR activity, as observed with both exogenous and endogenous AR targets. Rescue of BAF57 function restored AR activity, thus demonstrating a specific requirement of BAF57 for AR activity. This action of BAF57 proved to be dependent on SWI/SNF ATPase function. BAF57 has previously been implicated in nuclear receptor coactivator function, and we show that, although BAF57 facilitated coactivator activity, only a selected subset required BAF57 for coactivator function. Lastly, we demonstrate that both BAF57 and BRM are required for the proliferation of AR-dependent prostatic adenocarcinoma cells. In summary, these findings identify BAF57 as a critical modulator of the AR that is capable of altering AR activity, coactivator function, and AR-dependent proliferation.     

N-acyl-N-alkyl-sulfonamide anchors derived from Kenner's safety-catch linker: powerful tools in bioorganic and medicinal chemistry

In 1971 Kenner et al. introduced the safety-catch principle into solid phase peptide synthesis. Thus two contradicting needs were addressed. On the one hand, sufficient stability of the linker substrate bond to impede hydrolysis or similar side reactions, on the other hand mild chemical conditions allowing for unscathed liberation of the precious products. Over the years this linker type emerged in several different chemical disciplines and nowadays it presents a useful and broadly applicable tool. Recent advancements and applications based on Kenner's safety-catch linker are reviewed.     

Parallel synthesis and dopamine D3/D2 receptor screening of novel {4-[4-(2-methoxyphenyl)piperazin-1-yl]butyl}carboxamides

We have applied a fast and high-yielding method for the parallel amidation of 4-[4-(2-methoxyphenyl)piperazin-1-yl]-butylamine yielding analogs of the partial dopamine receptor agonist BP 897. Using this amino scaffold prepared in solution and polymer-bound carboxylic acid equivalents, we have synthesized a series of high affinity dopamine D(3) receptor ligands. The novel compounds were obtained in good to excellent yield and purity. Biological evaluation included determination of binding affinities at hD(2S) and hD(3) receptor subtypes. From the 22 novel structures presented here, compound 4v showed high affinity (K(i) (hD(3)) 1.6nM) and a 136-fold preference for the D(3) receptor versus that for the D(2) receptor subtype. Our results suggest that this derivatization technique is a useful method to speed up structure-activity relationships studies on dopamine receptor subtype modulators.     

Reassignment of sense codons in vivo

The genetic code maps one or more of the 61 sense codons onto a set of 20 canonical amino acids. Reassignment of sense codons to non-canonical amino acids in model organisms such as Escherichia coli has been achieved through manipulation of the cellular protein synthesis machinery. Specifically, control of amino acid pools, coupled with engineering of the aminoacyl-tRNA synthetase activity of the host, has enabled assignment of sense codons to a wide variety of non-canonical amino acids under conditions routinely used for expression of recombinant proteins. Codon reassignment is leading to important advances in protein engineering and bioorganic chemistry. Here we summarize some of those advances, and provide detailed protocols for codon reassignment.     

Parallel tandem: a program for parallel processing of tandem mass spectra using PVM or MPI and X!Tandem

A method for the rapid correlation of tandem mass spectra to a list of protein sequences in a database has been developed. The combination of the fast and accurate computational search algorithm, X!Tandem, and a Linux cluster parallel computing environment with PVM or MPI, significantly reduces the time required to perform the correlation of tandem mass spectra to protein sequences in a database. A file of tandem mass spectra is divided into a specified number of files, each containing an equal number of the spectra from the larger file. These files are then searched in parallel against a protein sequence database. The results of each parallel output file are collated into one file for viewing through a web interface. Thousands of spectra can be searched in an accurate, practical, and time effective manner. The source code for running Parallel Tandem utilizing either PVM or MPI on Linux operating system is available from http://www.thegpm.org. This source code is made available under Artistic License from the authors.     

Comparison of genetic algorithms for experimental multi-objective optimization on the example of medium design for cyanobacteria

In this work, two different genetic algorithms were applied to improve culture media composition for the autotrophic cyanobacteria Synechococcus PCC 7942. Biomass yield and conversion of the asymmetric reduction of 2', 3', 4', 5', 6'-pentafluoroacetophenone were considered as simultaneous objectives, resulting in a multi-objective optimization problem. Even when similar performances of both algorithms were observed, it could be shown that a novel strength pareto approach was able to achieve remarkable results with a reduced number of experiments (160 instead of 320). Handling a high number of media components (13), their concentrations were adjusted, delivering high improvements in comparison to the standard BG 11 culture media. The quality of the Synechococcus biocatalyst could be increased up to fivefold compared to the initial state of the optimization.     

Analyzing proteomes and protein function using graphical comparative analysis of tandem mass spectrometry results

Although generating large amounts of proteomic data using tandem mass spectrometry has become routine, there is currently no single set of comprehensive tools for the rigorous analysis of tandem mass spectrometry results given the large variety of possible experimental aims. Currently available applications are typically designed for displaying proteins and posttranslational modifications from the point of view of the mass spectrometrist and are not versatile enough to allow investigators to develop biological models of protein function, protein structure, or cell state. In addition, storage and dissemination of mass spectrometry-based proteomic data are problems facing the scientific community. To address these issues, we have developed a relational database model that efficiently stores and manages large amounts of tandem mass spectrometry results. We have developed an integrated suite of multifunctional analysis software for interpreting, comparing, and displaying these results. Our system, Bioinformatic Graphical Comparative Analysis Tools (BIGCAT), allows sophisticated analysis of tandem mass spectrometry results in a biologically intuitive format and provides a solution to many data storage and dissemination issues.     

Occurrence of tetraploidy in Nicotiana attenuata plants after Agrobacterium-mediated transformation is genotype specific but independent of polysomaty of explant tissue

Genotypes of Nicotiana attenuata collected from Utah and Arizona were transformed with 17 different vectors (14 unpublished vectors based on 3 new backbone vectors) using an Agrobacterium-mediated procedure to functionally analyze genes important for plant–insect interactions. None of the 51 T1–T3 transgenic Utah lines analyzed by the flow cytometry were tetraploid, as opposed to 18 of 33 transgenic Arizona lines (55%). Analysis of T0 regenerants transformed with the same vector carrying an inverted repeat (IR) N. attenuata pro-systemin construct confirmed the genotype dependency of tetraploidization: none of the 23 transgenic Utah lines were tetraploid but 31 (72%) of 43 transgenic Arizonas were tetraploid. We tested the hypothesis that the differences in polysomaty of the explant tissues accounted for genotype dependency of tetraploid formation by measuring polysomaty levels in different seedling tissues. Hypocotyls, cotyledons, and roots of Utah and Arizona genotypes contained similar percentages of 4C nuclei (61 and 60; 7 and 5; and 58 and 61%, respectively). Since we used hypocotyls as explant sources and the nonoccurrence of tetraploid Utah transformants does not correspond to the high percentage of 4C nuclei in Utah hypocotyls, we can rule out a direct relationship between tetraploid formation and polysomaty level. We hypothesize that the difference between the Utah and Arizona genotypes results from the failure of polyploid Utah callus to regenerate into fully competent plants. We propose that future work on post-transformation polyploidy concentrate on the processes that occur during callus formation and plant regeneration from callus.Electronic Supplementary Material Supplementary material is available for this article at