Anna Christina M. Salim

Swine and Poultry Pathogens: the Complete Genome Sequences of Two Strains of Mycoplasma hyopneumoniae and a Strain of Mycoplasma synoviae

This work reports the results of analyses of three complete mycoplasma genomes, a pathogenic (7448) and a nonpathogenic (J) strain of the swine pathogen Mycoplasma hyopneumoniae and a strain of the avian pathogen Mycoplasma synoviae; the genome sizes of the three strains were 920,079 bp, 897,405 bp, and 799,476 bp, respectively. These genomes were compared with other sequenced mycoplasma genomes reported in the literature to examine several aspects of mycoplasma evolution. Strain-specific regions, including integrative and conjugal elements, and genome rearrangements and alterations in adhesin sequences were observed in the M. hyopneumoniae strains, and all of these were potentially related to pathogenicity. Genomic comparisons revealed that reduction in genome size implied loss of redundant metabolic pathways, with maintenance of alternative routes in different species. Horizontal gene transfer was consistently observed between M. synoviae and Mycoplasma gallisepticum. Our analyses indicated a likely transfer event of hemagglutinin-coding DNA sequences from M. gallisepticum to M. synoviae.

Epigenetic silencing of the adhesion molecule ADAM23 is highly frequent in breast tumors

Altered cell adhesion is causally involved in tumor progression, and the identification of novel adhesion molecules altered in tumors is crucial for our understanding of tumor biology and for the development of new prognostic and therapeutic strategies. Here, we provide evidence for the epigenetic downregulation in breast tumors of the A Desintegrin And Metalloprotease domain 23 gene (ADAM 23), a member of a new family of surface molecules with roles in cell–cell adhesion and/or cell–matrix interactions. We examined the mRNA expression and methylation status of the 5′ upstream region of the ADAM23 gene in different breast tumor cell lines as well as in primary breast tumors. We found ADAM23 5′ hypermethylation in eight out of 12 (66.7%) tumor cell lines and in nine out of 13 (69.2%) primary tumors. Promoter hypermethylation was strongly associated with reductions in both mRNA and protein expression, with a threshold of 40–60% of modified CpG dinucleotides being required for the complete silencing of ADAM23 mRNA expression. Treatment of MCF-7 and SKBR-3 cell lines with 5′-Aza-2′-deoxycytidine led to a reactivation of ADAM23 mRNA expression and a marked decrease in the methylation level. It is worth noting that primary breast tumors with a more advanced grade showed a higher degree of methylation, suggesting that the adhesion molecule ADAM23 may be downregulated during the progression of breast cancer.

Comprehensive evaluation of the effectiveness of gene expression signatures to predict complete response to neoadjuvant chemoradiotherapy and guide surgical intervention in rectal cancer

Neoadjuvant chemoradiotherapy (nCRT) may lead to complete tumor regression in rectal cancer patients. Prediction of complete response to nCRT may allow a personalized management of rectal cancer and spare patients from unnecessary radical total mesorectal excision with or without sphincter preservation. To identify a gene expression signature capable of predicting complete pathological response (pCR) to nCRT, we performed a gene expression analysis in 25 pretreatment biopsies from patients who underwent 5FU-based nCRT using RNA-Seq. A supervised learning algorithm was used to identify expression signatures capable of predicting pCR, and the predictive value of these signatures was validated using independent samples. We also evaluated the utility of previously published signatures in predicting complete response in our cohort. We identified 27 differentially expressed genes between patients with pCR and patients with incomplete responses to nCRT. Predictive gene signatures using subsets of these 27 differentially expressed genes peaked at 81.8% accuracy. However, signatures with the highest sensitivity showed poor specificity, and vice-versa, when applied in an independent set of patients. Testing previously published signatures on our cohort also showed poor predictive value. Our results indicate that currently available predictive signatures are highly dependent on the sample set from which they are derived, and their accuracy is not superior to current imaging and clinical parameters used to assess response to nCRT and guide surgical intervention.

Distinct patterns of somatic alterations in a lymphoblastoid and a tumor genome derived from the same individual

Although patterns of somatic alterations have been reported for tumor genomes, little is known on how they compare with alterations present in non-tumor genomes. A comparison of the two would be crucial to better characterize the genetic alterations driving tumorigenesis. We sequenced the genomes of a lymphoblastoid (HCC1954BL) and a breast tumor (HCC1954) cell line derived from the same patient and compared the somatic alterations present in both. The lymphoblastoid genome presents a comparable number and similar spectrum of nucleotide substitutions to that found in the tumor genome. However, a significant difference in the ratio of non-synonymous to synonymous substitutions was observed between both genomes (P = 0.031). Protein–protein interaction analysis revealed that mutations in the tumor genome preferentially affect hub-genes (P = 0.0017) and are co-selected to present synergistic functions (P < 0.0001). KEGG analysis showed that in the tumor genome most mutated genes were organized into signaling pathways related to tumorigenesis. No such organization or synergy was observed in the lymphoblastoid genome. Our results indicate that endogenous mutagens and replication errors can generate the overall number of mutations required to drive tumorigenesis and that it is the combination rather than the frequency of mutations that is crucial to complete tumorigenic transformation.

Identification of 9 novel transcripts and two RGSL genes within the hereditary prostate cancer region (HPC1) at 1q25

We applied a systematic bioinformatics approach, followed by careful manual inspection and experimental validation to identify additional expressed sequences located at the Hereditary Prostate Cancer Region (HPC1) between D1S2818 and D1S1642 on chromosome 1q25. All transcripts already described for the 1q25 region were identified and we were able to define 11 additional expressed sequences within this region (three full-length cDNA clone sequences and eight ESTs), increasing the total number of gene count in this region by 38%. Five out of the 11 expressed sequences identified were shown to be expressed in prostate tissue and thus represent novel disease gene candidates for the HPC1 region. Here, we report a detailed characterization of these five novel disease gene candidates, their expression pattern in various tissues, their genomic organization and functional annotation. Two candidates (RGSL1 and RGSL2) correspond to novel members of the RGS family, which is involved in the regulation of G-protein signaling. RGSL1 and RGLS2 expression was detected by real-time polymerase chain reaction in normal prostate tissue, but could not be detected in prostate tumor cell lines, suggesting they might have a role in prostate cancer.

Identification and characterization of a novel mouse gene encoding a Ras-associated guanine nucleotide exchange factor: expression in macrophages and myocarditis elicited by Trypanosoma cruzi parasites.

The ability of Trypanosoma cruzi to activate macrophages is, at least in part, attributed to the glycosylphosphatidylinositol‐anchored mucin‐like glycoproteins (GPI‐mucins) expressed in the surface of the trypomastigote stage of the parasite. The differential display reverse transcriptase‐polymerase chain reaction and the reverse Northern blot were used to study modulation of gene expression in murine macrophages exposed to GPI‐mucins and in cardiac tissues from mice infected with T. cruzi. Among several cDNAs that were more abundant in lanes corresponding to macrophages stimulated with GPI‐mucins as compared with resting cells, we confirmed the differential expression of A1, interleukin‐18, and GPIγ4. Some of these genes were also shown to have enhanced expression in the cardiac tissue (DAP‐12, A1, and GPIγ4) from infected animals. The expression of GPIγ4 was also enhanced in human monocytes stimulated with GPI‐mucins or bacterial lipopolysaccharides. The complete sequence of the GPIγ4 transcript and its gene including the 5′ upstream region was defined. GPIγ4 was encoded by a novel, single copy gene present in mouse as well as human genomes and showed conserved homology to different members of the guanine nucleotide exchange factor family.

Analysis of allelic differential expression in the human genome using allele-specific serial analysis of gene expression tags

Recent reports have demonstrated that a significant proportion of human genes display allelic differential expression (ADE). ADE is associated with phenotypic variability and may contribute to complex genetic diseases. Here, we present a computational analysis of ADE using allele-specific serial analysis of gene expression (SAGE) tags representing 1295 human genes. We identified 472 genes for which unequal representation (>3-fold) of allele-specific SAGE tags was observed in at least one SAGE library, suggesting the occurrence of ADE. For 235 out of these 472 genes, the difference in the expression level between both allele-specific SAGE tags was statistically significant (p < 0.05). Eleven candidate genes were then subjected to experimental validation and ADE was confirmed for 8 out of these 11 genes. Our results suggest that at least 25% of the human genes display ADE and that allele-specific SAGE tags can be efficiently used for the identification of such genes.