Activation of LTRs from Different Human Endogenous Retrovirus (HERV) Families by the HTLV-1 Tax Protein and T-Cell Activators

Human endogenous retroviruses (HERVs) represent approximately 8% of our genome. HERVs influence cellular gene expression and contribute to normal physiological processes such as cellular differentiation and morphogenesis. HERVs have also been associated with certain pathological conditions, including cancer and neurodegenerative diseases. As HTLV-1 causes adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and has been shown to modulate host gene expression mainly through the expression of the powerful Tax transactivator, herein we were interested in looking at the potential modulation capacity of HTLV-1 Tax on HERV expression. In order to evaluate the promoter activity of different HERV LTRs, pHERV-LTR-luc constructs were co-transfected in Jurkat T-cells with a Tax expression vector. Tax expression potently increased the LTR activity of HERV-W8 and HERV-H (MC16). In parallel, Jurkat cells were also stimulated with different T-cell-activating agents and HERV LTRs were observed to respond to different combination of Forskolin, bpV[pic] a protein tyrosine phosphatase inhibitor, and PMA. Transfection of expression vectors for different Tax mutants in Jurkat cells showed that several transcription factors including CREB appeared to be important for HERV-W8 LTR activation. Deletion mutants were derived from the HERV-W8 LTR and the region from −137 to −123 was found to be important for LTR response following Tax expression in Jurkat cells, while a different region was shown to be required in cells treated with activators. Our results thus demonstrated that HTLV-1 Tax activates several HERV LTRs. This raises the possibility that upregulated HERV expression could be involved in diseases associated with HTLV-1 infection

A Mathematical Model for Suppression Subtractive Hybridization

Suppression subtractive hybridization (SSH) is frequently used to unearth differentially expressed genes on a whole-genome scale. Its versatility is based on combining cDNA library subtraction and normalization, which allows the isolation of sequences of varying degrees of abundance and differential expression. SSH is a complex process with many adjustable parameters that affect the outcome of gene isolation.We present a mathematical model of SSH based on DNA hybridization kinetics for assessing the effect of various parameters to facilitate its optimization. We derive an equation for the probability that a particular differentially expressed species is successfully isolated and use this to quantify the effect of the following parameters related to the cDNA sample: (a) mRNA abundance; (b) partial sequence complementarity to other species; and (3) degree of differential expression. We also evaluate the effect of parameters related to the process, including: (a) reaction times; and (b) extent of driver excess used in the two hybridization reactions. The optimum set of process parameters for successful isolation of differentially expressed species depends on transcript abundance. We show that the reaction conditions have a significant effect on the occurrence of false-positives and formulate strategies to isolate specific subsets of differentially expressed genes. We also quantify the effect of non-specific hybridization on the false-positive results and present strategies for spiking cDNA sequences to address this problem

Widely variable endogenous retroviral methylation levels in human placenta

It is generally assumed that transposable elements, including endogenous retroviruses (ERVs), are silenced by DNA methylation/chromatin structure in mammalian cells. However, there have been very few experimental studies to examine the methylation status of human ERVs. In this study, we determined and compared the methylation status of the 5′ long terminal repeats (LTRs) of different copies of the human endogenous retrovirus (HERV) family HERV-E, which are inserted in various genomic contexts. We found that three HERV-E LTRs which function as alternative gene promoters in placenta are unmethylated in that tissue but heavily methylated in blood cells, where these LTRs are not active promoters. This difference is not solely due to global hypomethylation in placenta, since two general measures of methylation levels of HERV-E and HERV-K LTRs suggest only 10–15% lower overall HERV methylation in placenta compared to blood. Comparisons between methylation levels of the LTR-derived gene promoters and six random HERV-E LTRs in placenta showed that the former display significantly lower methylation levels than random LTRs. Moreover, the differences in methylation between LTRs cannot always be explained by their genomic environment, since methylation of flanking sequences can be very different from methylation of the LTR itself

New insights into HIV-1-primary skin disorders

Since the first reports of AIDS, skin involvement has become a burdensome stigma for seropositive patients and a challenging task for dermatologist and infectious disease specialists due to the severe and recalcitrant nature of the conditions. Dermatologic manifestations in AIDS patients act as markers of disease progression, a fact that enhances the importance of understanding their pathogenesis

Identification of a human endogenous LTR-like sequence using HIV-1 LTR specific primers.

Using 'consensus' primers derived from the LTR region of 15 HIV-1 isolates, a fragment of 583 bp was amplified from human DNA. Even though specificity was confirmed by Southern blot analysis with a conserved LTR oligonucleotide probe, no significant homologies were detected to either retroviral regions or human or non-human published sequences. Nevertheless, when used as a probe, the 583-bp fragment identified a unique DNA sequence in the human genome on chromosome 1, and cross-reactive sequences in monkey, but not mouse, DNA. This novel, unique and conserved sequence of 583 bp was used to isolate a human HS-1 clone in which the structural property of a viral LTR could be identified