Genome-wide analysis of cancer/testis gene expression.

Cancer/Testis (CT) genes, normally expressed in germ line cells but also activated in a wide range of cancer types, often encode antigens that are immunogenic in cancer patients, and present potential for use as biomarkers and targets for immunotherapy. Using multiple in silico gene expression analysis technologies, including twice the number of expressed sequence tags used in previous studies, we have performed a comprehensive genome-wide survey of expression for a set of 153 previously described CT genes in normal and cancer expression libraries. We find that although they are generally highly expressed in testis, these genes exhibit heterogeneous gene expression profiles, allowing their classification into testis-restricted (39), testis/brain-restricted (14), and a testis-selective (85) group of genes that show additional expression in somatic tissues. The chromosomal distribution of these genes confirmed the previously observed dominance of X chromosome location, with CT-X genes being significantly more testis-restricted than non-X CT. Applying this core classification in a genome-wide survey we identified >30 CT candidate genes; 3 of them, PEPP-2, OTOA, and AKAP4, were confirmed as testis-restricted or testis-selective using RT-PCR, with variable expression frequencies observed in a panel of cancer cell lines. Our classification provides an objective ranking for potential CT genes, which is useful in guiding further identification and characterization of these potentially important diagnostic and therapeutic targets.

Jasmonate and salicylate as global signals for defense gene expression.

Remarkably, only a few low molecular mass signals, including jasmonic acid, ethylene and salicylic acid, upregulate the expression of scores of defense-related genes. Using these regulators, the plant fine-tunes its defense gene expression against aggressors which, in some cases, may be able to disrupt or amplify plant defense signal pathways to their own ends.

Targeted Gene Deletion and In Vivo Analysis of Putative Virulence Gene Function in the Pathogenic Dermatophyte Arthroderma benhamiae.

Dermatophytes cause the majority of superficial mycoses in humans and animals. However, little is known about the pathogenicity of this specialized group of filamentous fungi, for which molecular research has been limited thus far. During experimental infection of guinea pigs by the human pathogenic dermatophyte Arthroderma benhamiae, we recently detected the activation of the fungal gene encoding malate synthase AcuE, a key enzyme of the glyoxylate cycle. By the establishment of the first genetic system for A. benhamiae, specific ΔacuE mutants were constructed in a wild-type strain and, in addition, in a derivative in which we inactivated the nonhomologous end-joining pathway by deletion of the A. benhamiae KU70 gene. The absence of AbenKU70 resulted in an increased frequency of the targeted insertion of linear DNA by homologous recombination, without notably altering the monitored in vitro growth abilities of the fungus or its virulence in a guinea pig infection model. Phenotypic analyses of ΔacuE mutants and complemented strains depicted that malate synthase is required for the growth of A. benhamiae on lipids, major constituents of the skin. However, mutant analysis did not reveal a pathogenic role of the A. benhamiae enzyme in guinea pig dermatophytosis or during epidermal invasion of the fungus in an in vitro model of reconstituted human epidermis. The presented efficient system for targeted genetic manipulation in A. benhamiae, paired with the analyzed infection models, will advance the functional characterization of putative virulence determinants in medically important dermatophytes.

The gene for the ligand binding chain of the human interferon gamma receptor.

In order to characterize the gene encoding the ligand binding (1(st); alpha) chain of the human IFN-gamma receptor, two overlapping cosmid clones were analyzed. The gene spans over 25 kilobases (kb) of the genomic DNA and has seven exons. The extracellular domain is encoded by exons 1 to 5 and by part of exon 6. The transmembrane region is also encoded by exon 6. Exon 7 encodes the intracellular domain and the 3' untranslated portion. The gene was located on chromosome 6q23.1, as determined by in situ hybridization. The 4 kb region upstream (5') of the gene was sequenced and analyzed for promoter activity. No consensus-matching TATA or CAAT boxes in the 5' region were found. Potential binding sites for Sp1, AP-1, AP-2, and CREB nuclear factors were identified. Compatible with the presence of the Sp1/AP-2 sites and the lack of TATA box, S1-nuclease mapping experiments showed multiple transcription initiation sites. Promoter activity of the 5' flanking region was analyzed with two different reporter genes: the Escherichia coli chloramphenicol acetyltransferase and human growth hormone. The smallest 5' region of the gene that still had full promoter activity was 692 base pairs in length. In addition, we found sequences belonging to the oldest family of Alu repeats, 2 - 3 kb upstream of the gene, which could be useful for genetic studies.

Embryonic vascular endothelial cells are malleable to reprogramming via Prox1 to a lymphatic gene signature.

BACKGROUND: In vivo studies demonstrate that the Prox1 transcription factor plays a critical role in the development of the early lymphatic system. Upon Prox1 expression, early lymphatic endothelial cells differentiate from the cardinal vein and begin to express lymphatic markers such as VEGFR-3, LYVE-1 and Podoplanin. Subsequent in vitro studies have found that differentiated vascular endothelial cells can be reprogrammed by Prox1 to express a lymphatic gene profile, suggesting that Prox1 can initiate the expression of a unique gene signature during lymphangiogenesis. While the in vitro data suggest that gene reprogramming occurs upon Prox1 expression, it is not clear if this is a direct result of Prox1 in vascular endothelial cells in vivo. RESULTS: Overexpression of Prox1 in vascular endothelial cells during embryonic development results in the reprogramming of genes to that of a more lymphatic signature. Consequent to this overexpression, embryos suffer from gross edema that results in embryonic lethality at E13.5. Furthermore, hemorrhaging and anemia is apparent along with clear defects in lymph sac development. Alterations in junctional proteins resulting in an increase in vascular permeability upon Prox1 overexpression may contribute to the complications found during embryonic development. CONCLUSION: We present a novel mouse model that addresses the importance of Prox1 in early embryonic lymphangiogenesis. It is clear that there needs to be a measured pattern of expression of Prox1 during embryonic development. Furthermore, Prox1 reprograms vascular endothelial cells in vivo by creating a molecular signature to that of a lymphatic endothelial cell.

Chromosomal rearrangements do not seem to affect the gene flow in hybrid zones between karyotypic races of the common shrew (Sorex araneus).

Chromosomal rearrangements are proposed to promote genetic differentiation between chromosomally differentiated taxa and therefore promote speciation. Due to their remarkable karyotypic polymorphism, the shrews of the Sorex araneus group were used to investigate the impact of chromosomal rearrangements on gene flow. Five intraspecific chromosomal hybrid zones characterized by different levels of karyotypic complexity were studied using 16 microsatellites markers. We observed low levels of genetic differentiation even in the hybrid zones with the highest karyotypic complexity. No evidence of restricted gene flow between differently rearranged chromosomes was observed. Contrary to what was observed at the interspecific level, the effect of chromosomal rearrangements on gene flow was undetectable within the S. araneus species.

Network-guided analysis of genes with altered somatic copy number and gene expression reveals pathways commonly perturbed in metastatic melanoma.

Cancer genomes frequently contain somatic copy number alterations (SCNA) that can significantly perturb the expression level of affected genes and thus disrupt pathways controlling normal growth. In melanoma, many studies have focussed on the copy number and gene expression levels of the BRAF, PTEN and MITF genes, but little has been done to identify new genes using these parameters at the genome-wide scale. Using karyotyping, SNP and CGH arrays, and RNA-seq, we have identified SCNA affecting gene expression ('SCNA-genes') in seven human metastatic melanoma cell lines. We showed that the combination of these techniques is useful to identify candidate genes potentially involved in tumorigenesis. Since few of these alterations were recurrent across our samples, we used a protein network-guided approach to determine whether any pathways were enriched in SCNA-genes in one or more samples. From this unbiased genome-wide analysis, we identified 28 significantly enriched pathway modules. Comparison with two large, independent melanoma SCNA datasets showed less than 10% overlap at the individual gene level, but network-guided analysis revealed 66% shared pathways, including all but three of the pathways identified in our data. Frequently altered pathways included WNT, cadherin signalling, angiogenesis and melanogenesis. Additionally, our results emphasize the potential of the EPHA3 and FRS2 gene products, involved in angiogenesis and migration, as possible therapeutic targets in melanoma. Our study demonstrates the utility of network-guided approaches, for both large and small datasets, to identify pathways recurrently perturbed in cancer.