Comparison of motif-based and whole-unique-sequence-based analyses of phage display library datasets generated by biopanning of anti-Borrelia burgdorferi immune sera.

Detection of protection-associated epitopes via reverse vaccinology is the first step for development of subunit vaccines against microbial pathogens. Mapping subunit vaccine targets requires high throughput methods, which would allow delineation of epitopes recognized by protective antibodies on a large scale. Phage displayed random peptide library coupled to Next Generation Sequencing (PDRPL/NGS) is the universal platform that enables high-yield identification of peptides that mimic epitopes (mimotopes).

Delineating Surface Epitopes of Lyme Disease Pathogen Targeted by Highly Protective Antibodies of New Zealand White Rabbits.

Lyme disease (LD), the most prevalent vector-borne illness in the United States and Europe, is caused by No vaccine is available for humans. Dogmatically, can establish a persistent infection in the mammalian host (e.g.

New Zealand White Rabbits Effectively Clear Borrelia burgdorferi B31 despite the Bacterium's Functional Antigenic Variation System.

is a tick-borne bacterium responsible for approximately 300,000 annual cases of Lyme disease (LD) in the United States, with increasing incidences in other parts of the world. The debilitating nature of LD is mainly attributed to the ability of to persist in patients for many years despite strong anti- antibody responses. Antimicrobial treatment of persistent infection is challenging.

Identification of Surface Epitopes Associated with Protection against Highly Immune-Evasive VlsE-Expressing Lyme Disease Spirochetes.

The tick-borne pathogen is responsible for approximately 300,000 Lyme disease (LD) cases per year in the United States. Recent increases in the number of LD cases, in addition to the spread of the tick vector and a lack of a vaccine, highlight an urgent need for designing and developing an efficacious LD vaccine. Identification of protective epitopes that could be used to develop a second-generation (subunit) vaccine is therefore imperative.

Identification of cancer-specific motifs in mimotope profiles of serum antibody repertoire.

Background: For fighting cancer, earlier detection is crucial. Circulating auto-antibodies produced by the patient's own immune system after exposure to cancer proteins are promising bio-markers for the early detection of cancer. Since an antibody recognizes not the whole antigen but 4-7 critical amino acids within the antigenic determinant (epitope), the whole proteome can be represented by a random peptide phage display library.

Antibody Response to Lyme Disease Spirochetes in the Context of VlsE-Mediated Immune Evasion.

Lyme disease (LD), the most prevalent tick-borne illness in North America, is caused by Borrelia burgdorferi The long-term survival of B. burgdorferi spirochetes in the mammalian host is achieved though VlsE-mediated antigenic variation. It is mathematically predicted that a highly variable surface antigen prolongs bacterial infection sufficiently to exhaust the immune response directed toward invariant surface antigens.

Nrf2 amplifies oxidative stress via induction of Klf9.

Reactive oxygen species (ROS) activate NF-E2-related transcription factor 2 (Nrf2), a key transcriptional regulator driving antioxidant gene expression and protection from oxidant injury. Here, we report that in response to elevation of intracellular ROS above a critical threshold, Nrf2 stimulates expression of transcription Kruppel-like factor 9 (Klf9), resulting in further Klf9-dependent increases in ROS and subsequent cell death. We demonstrated that Klf9 independently causes increased ROS levels in various types of cultured cells and in mouse tissues and is required for pathogenesis of bleomycin-induced pulmonary fibrosis in mice.

Serum Antibody Repertoire Profiling Using In Silico Antigen Screen.

Serum antibodies are valuable source of information on the health state of an organism. The profiles of serum antibody reactivity can be generated by using a high throughput sequencing of peptide-coding DNA from combinatorial random peptide phage display libraries selected for binding to serum antibodies. Here we demonstrate that the targets of immune response, which are recognized by serum antibodies directed against sequential epitopes, can be identified using the serum antibody repertoire profiles generated by high throughput sequencing.

A high throughput method for identifying personalized tumor-associated antigens.

Circulating autoantibodies against tumor-associated antigens (TAAs) and their pattern of glycosylation can be used as diagnostic indicators of cancer. Using random peptide library screening, we identified patient-specific sets of peptides recognized by colon cancer patients' serum IgG and IgM antibodies. We demonstrate a strategy for analyzing BLAST search results for identifying tumor-associated antigens represented by peptides that mimic sequential epitopes.

Par-3 partitioning defective 3 homolog (C. elegans) and androgen-induced prostate proliferative shutoff associated protein genes are mutationally inactivated in prostate cancer cells.

Background: Gene identification by nonsense-mediated mRNA decay inhibition (GINI) has proven its usefulness in identifying mutant genes in cancer cell lines. An increase in transcription in response to NMD inhibition of a subset of genes is a major cause of false positives when genes are selected for sequencing analysis. To distinguish between mRNA accumulations caused by stress response-induced transcription and nonsense-containing mRNA stabilizations is a challenge in identifying mutant genes using GINI.

Truncating mutations in the ACVR2 gene attenuates activin signaling in prostate cancer cells.

Activins are classified as members of the TGFbeta superfamily of signaling molecules and both activin and TGFbeta ligands signal through structurally and functionally related serine/threonine kinase receptors. Defects in these signaling pathways have been associated with the initiation and progression of the cancer phenotype. Inactivating mutations in the TGFbeta type II receptor gene, TGFbetaR2, have been identified in a variety of tumors and cell lines, particularly those with microsatellite instability (MSI).

Identification of inactivating mutations in the JAK1, SYNJ2, and CLPTM1 genes in prostate cancer cells using inhibition of nonsense-mediated decay and microarray analysis.

We have developed a simple analytical method that increases the efficiency of identifying mutant genes in cell lines after the inhibition of nonsense-mediated decay (NMD). The approach assumes that the spectra of mutant genes differ between cell lines of the same tumor origin. Thus, by analyzing more than one cell line in parallel and taking into account not only changes in mRNA levels after the inhibition of NMD, but also comparing mRNA levels between cell lines before the inhibition of NMD, the vast majority of false positives were eliminated from the analysis.

Manipulation of nonsense mediated decay identifies gene mutations in colon cancer Cells with microsatellite instability.

Cancer cells showing microsatellite instability (MSI) demonstrate a high frequency of acquired frameshift mutations that result in the generation of nonsense mutations. RNA transcripts carrying these nonsense mutations are usually targeted for degradation through the nonsense mediated decay (NMD) pathway. Blocking this pathway with drugs such as emitine, results in the 'stabilization' of these mutant transcripts, which can now be detected on cDNA arrays.

A role for p300/CREB binding protein genes in promoting cancer progression in colon cancer cell lines with microsatellite instability.

Our manipulation of the nonsense-mediated decay pathway in microsatellite unstable colon cancer cell lines identified the p300 gene as a potential tumor suppressor in this subtype of cancer. Here, we have demonstrated that not only the p300 gene but also the highly homologous cAMP-response element-binding protein (CREB) binding protein (CBP) gene together are mutated in >85% of microsatellite instability (MSI)+ colon cancer cell lines. A limited survey of primary tumors with MSI+ shows that p300 is also frequently mutated in these cancers, demonstrating that these mutations are not consequences of in vitro growth.

Pim-1 protein kinase is nuclear in Burkitt's lymphoma: nuclear localization is necessary for its biologic effects.

Background: The Pim-1 33-kDa protein is a serine/threonine protein kinase that is capable of enhancing the rate of occurrence of c-Myc-induced lymphomas, and functions to block factor-withdrawal and genotoxic stress-induced apoptosis.

Materials And Methods: We used human lymphoma samples and tissue culture cells to examine the cellular location of this protein and its mechanism of action.

Results: We found that Pim-1 can be located in the cytoplasm, the cytoplasm and nucleus, or the nucleus of cells of normal lymph nodes, but is only located in the nucleus in Burkitt's lymphoma cells.