Noncognate Signals Drive Enhanced Effector CD8 T Cell Responses through an IFNAR1-Dependent Pathway after Infection with the Prototypic Vaccine, 0ΔNLS, against Herpes Simplex Virus 1.

Previous studies by our group identified a highly efficacious vaccine 0ΔNLS (deficient in the nuclear localization signal of infected cell protein 0) against herpes simplex virus 1 (HSV-1) in an experimental ocular mouse model. However, details regarding fundamental differences in the initial innate and adaptive host immune response were not explored. Here, we present a side-by-side analysis of the primary infection characterizing differences of the host immune response in mice infected with 0ΔNLS versus the parental, GFP105.

Digital Data Storage Using DNA Nanostructures and Solid-State Nanopores.

Solid-state nanopores are powerful tools for reading the three-dimensional shape of molecules, allowing for the translation of molecular structure information into electric signals. Here, we show a high-resolution integrated nanopore system for identifying DNA nanostructures that has the capability of distinguishing attached short DNA hairpins with only a stem length difference of 8 bp along a DNA double strand named the DNA carrier. Using our platform, we can read up to 112 DNA hairpins with a separating distance of 114 bp attached on a DNA carrier that carries digital information.

Small molecule protein interaction profiling with functional protein microarrays.

Small molecules possess the ability to interact with proteins and perturb their specific functions, a property that has been exploited for numerous research applications and to produce therapeutic agents in disease treatment. However, commonly utilized mass spectrometry-based approaches for identifying the target proteins for a small molecule have a number of limitations, particularly in terms of throughput and time and resource consumption. In addition, current technologies lack a mechanism to broadly assess the selectivity profile of the small molecule, which may be important for understanding off-target effects of the compound.

Identification and quantification of proteins differentially secreted by a pair of normal and malignant breast-cancer cell lines.

Secreted proteins play a pivotal role in cellular functions. To better understand malignant behavior, we adapted stable isotopic labeling with amino acids in cell culture technology to identify and quantify proteins differentially released into the extracellular media by a pair of normal and malignant breast-cancer cell lines. Approximately 380 non-redundant proteins were quantified in serum-free media.

Identification of small molecule targets on functional protein microarrays.

Small molecules, such as metabolites and hormones, interact with proteins to regulate numerous biological pathways, which are often aberrant in disease. Small molecule drugs have been successfully exploited to specifically perturb such processes and thereby, decrease and even eliminate disease progression. Although there are compelling reasons to fully characterize interactions of small molecules with all proteins from an organism for which an intended drug regimen is planned, currently available technologies are not yet up to this task.

Quantitative comparison of IMAC and TiO2 surfaces used in the study of regulated, dynamic protein phosphorylation.

Protein phosphorylation regulates many aspects of cellular function, including cell proliferation, migration, and signal transduction. An efficient strategy to isolate phosphopeptides from a pool of unphosphorylated peptides is essential to global characterization using mass spectrometry. We describe an approach employing isotope tagging reagents for relative and absolute quantification (iTRAQ) labeling to compare quantitatively commercial and prototypal immobilized metal affinity chelate (IMAC) and metal oxide resins.

A critical role for cortactin phosphorylation by Abl-family kinases in PDGF-induced dorsal-wave formation.

Proper regulation of cell morphogenesis and migration by adhesion and growth-factor receptors requires Abl-family tyrosine kinases [1-3]. Several substrates of Abl-family kinase have been identified, but they are unlikely to mediate all of the downstream actions of these kinases on cytoskeletal structure. We used a human protein microarray to identify the actin-regulatory protein cortactin as a novel substrate of the Abl and Abl-related gene (Arg) nonreceptor tyrosine kinases.

A pooling-deconvolution strategy for biological network elucidation.

The generation of large-scale data sets is a fundamental requirement of systems biology. But despite recent advances, generation of such high-coverage data remains a major challenge. We developed a pooling-deconvolution strategy that can dramatically decrease the effort required.

Protein microarrays: a new tool for profiling antibody cross-reactivity.

Antibody cross-reactivity can compromise interpretation of experiments and derail therapeutic antibody development. Standard techniques such as immunohistochemistry or Western analysis provide important but often inadequate approaches to assess antibody specificity. Protein microarrays are providing a new approach to rapidly characterize antibody cross-reactivity against 1,000s of proteins simultaneously.

Global analysis of protein phosphorylation in yeast.

Protein phosphorylation is estimated to affect 30% of the proteome and is a major regulatory mechanism that controls many basic cellular processes. Until recently, our biochemical understanding of protein phosphorylation on a global scale has been extremely limited; only one half of the yeast kinases have known in vivo substrates and the phosphorylating kinase is known for less than 160 phosphoproteins. Here we describe, with the use of proteome chip technology, the in vitro substrates recognized by most yeast protein kinases: we identified over 4,000 phosphorylation events involving 1,325 different proteins.

Protein microarray-based screening of antibody specificity.

The increased use of antibodies as therapeutics, as well as the growing demand for large numbers of antibodies for high-throughput protein analyses, has been accompanied by a need for more specific antibodies. An array containing every protein for the relevant organism represents the ideal format for an assay to test antibody specificity since it allows the simultaneous screening of thousands of proteins in relatively normalized quantities. Indeed, the use of a yeast proleome array to profile the specificity of several antibodies directed against yeast proteins has recently been described.

Functional protein microarrays: just how functional are they?

Arrays of immobilized proteins have been developed for the discovery and characterization of protein functions ranging from molecular recognition to enzymatic activity. The success of these applications is highly dependent upon the maintenance of protein structure and function while in an immobilized state - a largely untested hypothesis. However, the immobilization of functional proteins is not without precedent.

The sequence and analysis of duplication-rich human chromosome 16.

Human chromosome 16 features one of the highest levels of segmentally duplicated sequence among the human autosomes. We report here the 78,884,754 base pairs of finished chromosome 16 sequence, representing over 99.9% of its euchromatin.

Development of functional protein microarrays for drug discovery: progress and challenges.

Functional protein microarrays promise new approaches to address longstanding challenges in drug discovery and development, with applications ranging from target identification to clinical trial design. However, their widespread adoption will be contingent upon a robust ability to develop and manufacture arrays in support of these applications. This review will address the major areas of relevance to the development of functional protein microarrays; protein content, surface chemistry, manufacture and assay development.

The DNA sequence and comparative analysis of human chromosome 5.

Chromosome 5 is one of the largest human chromosomes and contains numerous intrachromosomal duplications, yet it has one of the lowest gene densities. This is partially explained by numerous gene-poor regions that display a remarkable degree of noncoding conservation with non-mammalian vertebrates, suggesting that they are functionally constrained. In total, we compiled 177.

Practical applications of rolling circle amplification of DNA templates.

Since its recent implementation at one of the world's largest high-throughput sequencing centers, the utility of MP-RCA for DNA sequencing has been thoroughly validated. However, applications of this technology extend far beyond DNA sequencing. While many of these applications have been explored in this chapter, the future will undoubtedly add to this growing list.

The DNA sequence and biology of human chromosome 19.

Chromosome 19 has the highest gene density of all human chromosomes, more than double the genome-wide average. The large clustered gene families, corresponding high G + C content, CpG islands and density of repetitive DNA indicate a chromosome rich in biological and evolutionary significance. Here we describe 55.

Functional protein microarrays: ripe for discovery.

The manufacture and use of protein microarrays with correctly folded and functional content presents significant challenges. Despite this, the feasibility and utility of such undertakings are now clear, and exciting progress has recently been demonstrated in the areas of content generation, printing strategies and protein immobilization. More importantly, we are now beginning to enjoy the fruits of these efforts as functional protein microarrays are being increasingly employed for biological discovery purposes.

Analyzing antibody specificity with whole proteome microarrays.

Although approximately 10,000 antibodies are available from commercial sources, antibody reagents are still unavailable for most proteins. Furthermore, new applications such as antibody arrays and monoclonal antibody therapeutics have increased the demand for more specific antibodies to reduce cross-reactivity and side effects. An array containing every protein for the relevant organism represents the ideal format for an assay to test antibody specificity, because it allows the simultaneous screening of thousands of proteins for possible cross-reactivity.

Microarrays to characterize protein interactions on a whole-proteome scale.

Protein microarrays contain a defined set of proteins spotted and analyzed at high density, and can be generally classified into two categories; protein profiling arrays and functional protein arrays. Functional protein arrays can be made up of any type of protein, and therefore have a diverse set of useful applications. Advantages of these arrays include low reagent consumption, rapid interpretation of results, and the ability to easily control experimental conditions.

Draft sequencing and comparative genomics of Xylella fastidiosa strains reveal novel biological insights.

Draft sequencing is a rapid and efficient method for determining the near-complete sequence of microbial genomes. Here we report a comparative analysis of one complete and two draft genome sequences of the phytopathogenic bacterium, Xylella fastidiosa, which causes serious disease in plants, including citrus, almond, and oleander. We present highlights of an in silico analysis based on a comparison of reconstructions of core biological subsystems.