Abyss1: a novel L2-like non-LTR retroelement of the snakelocks anemone (Anemonia sulcata).

Non-LTR retrotransposons are a diverse and taxonomically widely dispersed group of retroelements that can be divided into at least 14 distinguishable clades. Basal metazoans have not been examined in great detail for their retrotransposon content. In order to screen for the presence of reverse transcriptase (RT) related sequences in Cnidaria and Ctenophora, basal phyla of metazoans, PCR with highly degenerate oligonucleotides was performed and an RT-like sequence was identified from the sea anemone species Anemonia sulcata. Further screening identified a related element in another anemone species Actinia equina. Significant homology to non-LTR retrotransposon RTs was observed, particularly to L2-like elements of fish such as Maui. The sequence was not detected among other cnidarians and we have designated the A. sulcata and A. equina elements Abyss1 and Abyss2 respectively. Phylogenetic analysis of Abyss1 compared with members of 14 known non-LTR retroelement clades suggests that the sequence represents a novel L2 element. &nbsp

Regulation of Human Endogenous Retrovirus (HERV) expression in human brain cells.

The expression profiles of a broad range of HERVs were analyzed in brain samples from patients with schizophrenia, bipolar disorders and healthy controls using a retrovirus specific microarray. Remarkably, upregulation of HERV-K(HML-2) elements was found to be significantly prevalent in both, bipolar disorder- and schizophrenia-associated samples, suggesting a potential association with disease. Therefore, the methylation status of HERV-K(HML-2) loci active in patient samples was investigated in various human brain cells. Preliminary results indicate a correlation between DNA demethylation and transcriptional activity of HERV-K (HML-2) loci.  In addition, we have investigated a potential influence of psychiatric medication on HERV transcription activity, since the majority of patients were treated with neuroleptica and/or antidepressants. Various cell lines derived from human brain (astrocytes and neuronal cells) were treated with different antipsychotic drugs, such as valproic acid, haloperidol, risperidone and clozapine. The drugs were used in non-cytotoxic concentrations. Alterations of HERV expression patterns were monitored with the retrovirus-specific microarray. All medications tested so far showed an influence on expression of at least some HERV families. A dose-dependent upregulation of transcriptional activity after treatment with valproic acid, haloperidol and risperidone was observed for several class I (HERV-Fb, HERV-W, ERV9) and class II HERVs (HERV-KC4). Valproic acid additionally showed an increased activity of HML-3 (class II HERV). Medication-dependent differences of HERV activity were validated by QRT-PCR. Furthermore, preliminary data suggest that members of the HML-2 family are not influenced. Thus, the differential HERV-K10 activity detected in brain samples of patients may rather relate to the disease than to psychiatric medication

RetroArray - a comprehensive diagnostic DNA chip for rapid detection and identificaton of retroviruses, retroviral contaminants, and mistaken identity of cell lines.

Retroviruses not only represent some of the most dangerous pathogens, but also constitute, as remnants of former infections that happened millions of years ago, a large fraction of the human genome. We have established a fast and reliable DNA chip-based assay (RetroArray) for detection and identification of a wide variety of human and other vertebrate exogenous and endogenous retroviruses in biological/clinical samples. The assay combines a pan-retrovirus multiplex polymerase chain reaction (PCR) using fluorochrome-modified primers and DNA chip hybridization. Using RetroArray, distinct transcription profiles of human endogenous retroviruses (HERVS) have been established for a variety of human tissues. Using paired samples (normal vs. Disease) this method can be applied to examine HERV activity in human tumors and can help to identify retrovirus-derived tumor antigens. In addition, RetroArray has been designed to detect human exogenous retroviruses such as human immunodeficity and sensitivity of the assay was demonstrated by detecting traces of pig endogenous retrovirus (PERV) DNA down to ~25 copies in human cDNA samples. Furthermore, retroviral transcripts may be identified in particle preparations from cell culture supernatants. This makes the assay a valuable technique for monitoring packaging cell lines and vector preparations commonly used in human gene therapy applications to exclude cotransfer of replication competent retroviruses (RCRs) or endogenous retrocviruses (ERVs) into target cell. Therefore, RetroArray could improve significantly the safety of human gene therapy, tissue engineering, xenotransplantation and production of therapeutic polypeptides in cell culture. Mistaken identity of human cell lines and frequently observed laboratory contaminations with cells of other species, as well as infection with polytropic animal retroviruses such as squirrel monkey retrocirus (SMRV) or murine leukemia virus (MLV) can influence experimental results and may lead to invalid conclusions. The RetroArray technique is an excellent tool for testing purity and homogeneity of cell lines. Characteristic HERV transcription profiles can be used to assess the cell type and to monitor cell lines for contaminating cells