Role of the cellular factor CTCF in the regulation of bovine leukemia virus latency and three-dimensional chromatin organization.

Bovine leukemia virus (BLV)-induced tumoral development is a multifactorial phenomenon that remains incompletely understood. Here, we highlight the critical role of the cellular CCCTC-binding factor (CTCF) both in the regulation of BLV transcriptional activities and in the deregulation of the three-dimensional (3D) chromatin architecture surrounding the BLV integration site. We demonstrated the in vivo recruitment of CTCF to three conserved CTCF binding motifs along the provirus.

The Molecular Biology of HIV Latency.

HIV remains incurable due to the existence of a reservoir of cells that harbor intact integrated genomes of the virus in the absence of viral replication. This population of infected cells remains invisible to the immune system and is not targeted by the drugs used in the current antiretroviral therapies (cART). Reversal of latency by the use of inhibitors of chromatin-remodeling enzymes has been studied extensively in an attempt to purge this reservoir of latent HIV but has thus far not shown any success in clinical trials.

Molecular Control of HIV and SIV Latency.

The HIV latent reservoirs are considered as the main hurdle to viral eradication. Numerous mechanisms lead to the establishment of HIV latency and act at the transcriptional and post-transcriptional levels. A better understanding of latency is needed in order to ultimately achieve a cure for HIV.

Primate lentiviruses use at least three alternative strategies to suppress NF-κB-mediated immune activation.

Primate lentiviruses have evolved sophisticated strategies to suppress the immune response of their host species. For example, HIV-2 and most simian immunodeficiency viruses (SIVs) use their accessory protein Nef to prevent T cell activation and antiviral gene expression by downmodulating the T cell receptor CD3. This Nef function was lost in HIV-1 and other vpu-encoding viruses suggesting that the acquisition of Vpu-mediated NF-κB inhibition reduced the selection pressure for inhibition of T cell activation by Nef.

Repression of Human T-lymphotropic virus type 1 Long Terminal Repeat sense transcription by Sp1 recruitment to novel Sp1 binding sites.

Human T-lymphotropic Virus type 1 (HTLV-1) infection is characterized by viral latency in the majority of infected cells and by the absence of viremia. These features are thought to be due to the repression of viral sense transcription in vivo. Here, our in silico analysis of the HTLV-1 Long Terminal Repeat (LTR) promoter nucleotide sequence revealed, in addition to the four Sp1 binding sites previously identified, the presence of two additional potential Sp1 sites within the R region.

HIV Latency: Should We Shock or Lock?

Combinatory antiretroviral therapy (cART) increases the survival and quality of life of HIV-1-infected patients. However, interruption of therapy almost invariably leads to the re-emergence of detectable viral replication because HIV-1 persists in viral latent reservoirs. Improved understanding of the molecular mechanisms involved in HIV-1 latency has paved the way for innovative strategies that attempt to purge latent virus.

Reactivation capacity by latency-reversing agents ex vivo correlates with the size of the HIV-1 reservoir.

Objective: HIV-1 reservoirs are the major hurdle to virus clearance in combination antiretroviral therapy (cART)-treated patients. An approach to eradicating HIV-1 involves reversing latency in cART-treated patients to make latent cells visible to the host immune system. Stimulation of patient cell cultures with latency-reversing agents (LRAs) ex vivo results in heterogeneous responses among HIV-infected patients.

HIC1 controls cellular- and HIV-1- gene transcription via interactions with CTIP2 and HMGA1.

Among many cellular transcriptional regulators, Bcl11b/CTIP2 and HGMA1 have been described to control the establishment and the persistence of HIV-1 latency in microglial cells, the main viral reservoir in the brain. In this present work, we identify and characterize a transcription factor i.e.

Characterization of new RNA polymerase III and RNA polymerase II transcriptional promoters in the Bovine Leukemia Virus genome.

Bovine leukemia virus latency is a viral strategy used to escape from the host immune system and contribute to tumor development. However, a highly expressed BLV micro-RNA cluster has been reported, suggesting that the BLV silencing is not complete. Here, we demonstrate the in vivo recruitment of RNA polymerase III to the BLV miRNA cluster both in BLV-latently infected cell lines and in ovine BLV-infected primary cells, through a canonical type 2 RNAPIII promoter.

Preclinical shock strategies to reactivate latent HIV-1: an update.

Purpose Of Review: The 'shock and kill' strategy consists of activating HIV-1 expression to allow latently infected cells to die from viral cytopathic effects or host cytolytic immune effectors. This strategy relies on small molecules, called latency reversing agents, which activate HIV transcription.

Recent Findings: Several mechanisms operating at the transcriptional level are involved in the establishment and maintenance of HIV-1 latency, including the absence of crucial inducible host transcription factors, epigenetic silencing, and the sequestration of the positive transcription elongation factor B.

Sequential treatment with 5-aza-2'-deoxycytidine and deacetylase inhibitors reactivates HIV-1.

Reactivation of HIV gene expression in latently infected cells together with an efficient cART has been proposed as an adjuvant therapy aimed at eliminating/decreasing the reservoir size. Results from HIV clinical trials using deacetylase inhibitors (HDACIs) question the efficiency of these latency-reversing agents (LRAs) used alone and underline the need to evaluate other LRAs in combination with HDACIs. Here, we evaluated the therapeutic potential of a demethylating agent (5-AzadC) in combination with clinically tolerable HDACIs in reactivating HIV-1 from latency first in vitro and next ex vivo.

Prdm12 specifies V1 interneurons through cross-repressive interactions with Dbx1 and Nkx6 genes in Xenopus.

V1 interneurons are inhibitory neurons that play an essential role in vertebrate locomotion. The molecular mechanisms underlying their genesis remain, however, largely undefined. Here, we show that the transcription factor Prdm12 is selectively expressed in p1 progenitors of the hindbrain and spinal cord in the frog embryo, and that a similar restricted expression profile is observed in the nerve cord of other vertebrates as well as of the cephalochordate amphioxus.

An In-Depth Comparison of Latency-Reversing Agent Combinations in Various In Vitro and Ex Vivo HIV-1 Latency Models Identified Bryostatin-1+JQ1 and Ingenol-B+JQ1 to Potently Reactivate Viral Gene Expression.

The persistence of latently infected cells in patients under combinatory antiretroviral therapy (cART) is a major hurdle to HIV-1 eradication. Strategies to purge these reservoirs are needed and activation of viral gene expression in latently infected cells is one promising strategy. Bromodomain and Extraterminal (BET) bromodomain inhibitors (BETi) are compounds able to reactivate latent proviruses in a positive transcription elongation factor b (P-TEFb)-dependent manner.

Differential regulation of NF-κB-mediated proviral and antiviral host gene expression by primate lentiviral Nef and Vpu proteins.

NF-κB is essential for effective transcription of primate lentiviral genomes and also activates antiviral host genes. Here, we show that the early protein Nef of most primate lentiviruses enhances NF-κB activation. In contrast, the late protein Vpu of HIV-1 and its simian precursors inhibits activation of NF-κB, even in the presence of Nef.

The Prdm13 histone methyltransferase encoding gene is a Ptf1a-Rbpj downstream target that suppresses glutamatergic and promotes GABAergic neuronal fate in the dorsal neural tube.

The basic helix-loop-helix (bHLH) transcriptional activator Ptf1a determines inhibitory GABAergic over excitatory glutamatergic neuronal cell fate in progenitors of the vertebrate dorsal spinal cord, cerebellum and retina. In an in situ hybridization expression survey of PR domain containing genes encoding putative chromatin-remodeling zinc finger transcription factors in Xenopus embryos, we identified Prdm13 as a histone methyltransferase belonging to the Ptf1a synexpression group. Gain and loss of Ptf1a function analyses in both frog and mice indicates that Prdm13 is positively regulated by Ptf1a and likely constitutes a direct transcriptional target.

CTIP2 is a negative regulator of P-TEFb.

The positive transcription elongation factor b (P-TEFb) is involved in physiological and pathological events including inflammation, cancer, AIDS, and cardiac hypertrophy. The balance between its active and inactive form is tightly controlled to ensure cellular integrity. We report that the transcriptional repressor CTIP2 is a major modulator of P-TEFb activity.

The AP-1 binding sites located in the pol gene intragenic regulatory region of HIV-1 are important for viral replication.

Our laboratory has previously identified an important intragenic region in the human immunodeficiency virus type 1 (HIV-1) genome, whose complete functional unit is composed of the 5103 fragment, the DNaseI-hypersensitive site HS7 and the 5105 fragment. These fragments (5103 and 5105) both exhibit a phorbol 12-myristate 13-acetate (PMA)-inducible enhancer activity on the herpes simplex virus thymidine kinase promoter. Here, we characterized the three previously identified AP-1 binding sites of fragment 5103 by showing the PMA-inducible in vitro binding and in vivo recruitment of c-Fos, JunB and JunD to this fragment located at the end of the pol gene.

DNA cytosine methylation in the bovine leukemia virus promoter is associated with latency in a lymphoma-derived B-cell line: potential involvement of direct inhibition of cAMP-responsive element (CRE)-binding protein/CRE modulator/activation transcription factor binding.

Bovine leukemia virus (BLV) proviral latency represents a viral strategy to escape the host immune system and allow tumor development. Besides the previously demonstrated role of histone deacetylation in the epigenetic repression of BLV expression, we showed here that BLV promoter activity was induced by several DNA methylation inhibitors (such as 5-aza-2'-deoxycytidine) and that overexpressed DNMT1 and DNMT3A, but not DNMT3B, down-regulated BLV promoter activity. Importantly, cytosine hypermethylation in the 5'-long terminal repeat (LTR) U3 and R regions was associated with true latency in the lymphoma-derived B-cell line L267 but not with defective latency in YR2 cells.

Three novel antibiotic marker cassettes for gene disruption and marker switching in Schizosaccharomyces pombe.

The ease of construction of multiple mutant strains in Schizosaccharomyces pombe is limited by the number of available genetic markers. We describe here three new cassettes for PCR-mediated gene disruption that can be used in combination with commonly used fission yeast markers to make multiple gene deletions. The natMX6, hphMX6 and bleMX6 markers give rise to resistance towards the antibiotics nourseothricin (NAT), hygromycin B and phleomycin, respectively.

Additional vectors for PCR-based gene tagging in Saccharomyces cerevisiae and Schizosaccharomyces pombe using nourseothricin resistance.

The one-step PCR-mediated technique used for modification of chromosomal loci is a powerful tool for functional analysis in yeast. Both Saccharomyces cerevisiae and Schizosaccharomyces pombe are amenable to this technique. However, the scarce availability of selectable markers for Sz.

Glucose deprivation mediates interaction between CTDK-I and Snf1 in Saccharomyces cerevisiae.

Ctk1 is a kinase involved in transcriptional control. We show in the two-hybrid system that Ctk1 interacts with Snf1, a kinase regulating glucose-dependent genes. Co-purification experiments confirmed the two-hybrid interaction but only when cells were grown at low glucose concentrations.

Members of the SAGA and Mediator complexes are partners of the transcription elongation factor TFIIS.

TFIIS, an elongation factor encoded by DST1 in Saccharomyces cerevisiae, stimulates transcript cleavage in arrested RNA polymerase II. Two components of the RNA polymerase II machinery, Med13 (Srb9) and Spt8, were isolated as two-hybrid partners of the conserved TFIIS N-terminal domain. They belong to the Cdk8 module of the Mediator and to a subform of the SAGA co-activator, respectively.