A sum-frequency generation (SFG) microscope that is sensitive toward molecular chirality was demonstrated for the first time. Optically active images of chiral 1,1’-bi-2-naphthol solutions were obtained with submicron spatial resolution. Three-dimensional sectioning capability of our microscope was also demonstrated. This optically active SFG microscopy can potentially become a powerful imaging technique for biological samples.

We pursue the hypothesis that neuronal placement in animals minimizes wiring costs for given functional constraints, as specified by synaptic connectivity. Using a newly compiled version of the Caenorhabditis elegans wiring diagram, we solve for the optimal layout of 279 nonpharyngeal neurons. In the optimal layout, most neurons are located close to their actual positions, suggesting that wiring minimization is an important factor. Yet some neurons exhibit strong deviations from "optimal" position. We propose that biological factors relating to axonal guidance and command neuron functions contribute to these deviations. We capture these factors by proposing a modified wiring cost function.

BACKGROUND: The best studied insect-symbiont system is that of aphids and their primary bacterial endosymbiont Buchnera aphidicola. Buchnera inhabits specialized host cells called bacteriocytes, provides nutrients to the aphid and has co-speciated with its aphid hosts for the past 150 million years. We have used a single microarray to examine gene expression in the pea aphid, Acyrthosiphon pisum, and its resident Buchnera. Very little is known of gene expression in aphids, few studies have examined gene expression in Buchnera, and no study has examined simultaneously the expression profiles of a host and its symbiont. Expression profiling of aphids, in studies such as this, will be critical for assigning newly discovered A. pisum genes to functional roles. In particular, because aphids possess many genes that are absent from Drosophila and other holometabolous insect taxa, aphid genome annotation efforts cannot rely entirely on homology to the best-studied insect systems. Development of this dual-genome array represents a first attempt to characterize gene expression in this emerging model system.

RESULTS: We chose to examine heat shock response because it has been well characterized both in Buchnera and in other insect species. Our results from the Buchnera of A. pisum show responses for the same gene set as an earlier study of heat shock response in Buchnera for the host aphid Schizaphis graminum. Additionally, analyses of aphid transcripts showed the expected response for homologs of known heat shock genes as well as responses for several genes with unknown functional roles.

CONCLUSION: We examined gene expression under heat shock of an insect and its bacterial symbiont in a single assay using a dual-genome microarray. Further, our results indicate that microarrays are a useful tool for inferring functional roles of genes in A. pisum and other insects and suggest that the pea aphid genome may contain many gene paralogs that are differentially regulated.

Aphids are the leading pests in agricultural crops. A large-scale sequencing of 40,904 ESTs from the pea aphid Acyrthosiphon pisum was carried out to define a catalog of 12,082 unique transcripts. A strong AT bias was found, indicating a compositional shift between Drosophila melanogaster and A. pisum. An in silico profiling analysis characterized 135 transcripts specific to pea-aphid tissues (relating to bacteriocytes and parthenogenetic embryos). This project is the first to address the genetics of the Hemiptera and of a hemimetabolous insect.

The Lrp/AsnC family of transcriptional regulatory proteins is found in both archaea and bacteria. Members of the family influence cellular metabolism in both a global (Lrp) and specific (AsnC) manner, often in response to exogenous amino acid effectors. In the present study we have determined both the first bacterial and the highest resolution structures for members of the family. Escherichia coli AsnC is a specific gene regulator whose activity is triggered by asparagine binding. Bacillus subtilis LrpC is a global regulator involved in chromosome condensation. Our AsnC-asparagine structure is the first for a regulator-effector complex and is revealed as an octameric disc. Key ligand recognition residues are identified together with a route for ligand access. The LrpC structure reveals a stable octamer supportive of a topological role in dynamic DNA packaging. The structures yield significant clues to the functionality of Lrp/AsnC-type regulators with respect to ligand binding and oligomerization states as well as to their role in specific and global DNA regulation.

Persistent neural activity that outlasts an initial stimulus is thought to provide a mechanism for the transient storage of memory. In this issue of Neuron, Fransén et al. identify important principles for a cell-autonomous mechanism of graded persistent firing using an elegant combination of experimental and computational approaches.

An important aspect of understanding a biological pathway is to delineate the transcriptional regulatory mechanisms of the genes involved. Two important tasks are often encountered when studying transcription regulation, i.e., (1) the identification of common transcriptional regulators of a set of coexpressed genes; (2) the identification of genes that are regulated by one or several transcription factors. In this study, a systematic and statistical approach was taken to accomplish these tasks by establishing an integrated model considering all of the promoters and characterized transcription factors (TFs) in the genome. A promoter analysis pipeline (PAP) was developed to implement this approach. PAP was tested using coregulated gene clusters collected from the literature. In most test cases, PAP identified the transcription regulators of the input genes accurately. When compared with chromatin immunoprecipitation experiment data, PAP’s predictions are consistent with the experimental observations. When PAP was used to analyze one published expression-profiling data set and two novel coregulated gene sets, PAP was able to generate biologically meaningful hypotheses. Therefore, by taking a systematic approach of considering all promoters and characterized TFs in our model, we were able to make more reliable predictions about the regulation of gene expression in mammalian organisms.

Virus assembly has not been routinely targeted in the development of antiviral drugs, in part because of the lack of tractable methods for screening in vitro. We have developed an in vitro assay of hepatitis B virus (HBV) capsid assembly, based on fluorescence quenching of dye-labeled capsid protein, for testing potential inhibitors. This assay is adaptable to high-throughput screening and can identify small-molecule inhibitors of virus assembly that prevent, inappropriately accelerate and/or misdirect capsid formation to yield aberrant particles. An in vitro primary screen has the advantage of identifying promising lead compounds affecting assembly without the requirement that they be taken up by cells in culture and be nontoxic. Our approach may facilitate the identification of antivirals targeting viruses other than HBV, such as avian influenza and HIV.

The additive and synergistic therapeutic effects derived from combinations of chemotherapeutic drugs and radiation have established an indispensable paradigm: cancer must be attacked on multiple fronts. However, the increased antitumor efficacy of such combinational regimens is also associated with severe systemic toxicity, as these drugs cannot selectively target tumor cells. Monoclonal antibodies (mAbs), which have exquisite specificity for their antigens, are becoming an increasingly important class of antitumor agents, as they enhance the efficacy of various therapeutic regimens without significantly increasing systemic toxicity. Furthermore, preclinical and early clinical evidence indicate that combinations of antibody-based drugs provide even greater efficacy with minimal side effects. Unfortunately, the research, manufacturing and regulatory costs of mAb development pose a significant barrier to the use of antibody-based combination therapies. An emerging alternative is the use of dual-targeting bispecific antibodies (BsAbs). BsAbs are derived from the recombination of variable domains of two antibodies with different specificities; BsAbs are thus capable of binding both antigens of their parental antibodies. With the recent progress that has been made in antibody engineering technology, BsAbs that simultaneously target two tumor-associated molecules (eg, growth factor receptors) are being heralded for their potential to deliver two therapeutic moieties in a single molecule.

The multisubunit transcription factor TFIID is essential for directing eukaryotic promoter recognition and mediating interactions with activators/cofactors during assembly of the preinitiation complex. Despite its central role in transcription initiation and regulation, structural knowledge of the TFIID complex has so far been largely limited to electron microscopy studies of negatively stained samples. Here, we present a cryo-electron microscopy 3D reconstruction of the large endogenous human TFIID complex. The improved cryopreservation has allowed for a more detailed definition of the structural elements in the complex and for the detection, by an extensive statistical analysis of the data, of a conformational opening and closing of the cavity central to the TFIID architecture. We propose that these density rearrangements in the structure are a likely reflection of the plasticity of the interactions between TFIID and its many partner proteins.