Both Human α/β and γ Interferons Upregulate the Expression of CD48 Cell Surface Molecules

International audienceWe have established a cDNA library from interferon (IFN)-treated human lymphoblastoid Daudi cells and made use of differential screening to search for yet unidentified IFN-regulated genes. In the course of these studies, we have isolated a human cDNA coding for the glycosyl-phosphatidylinositol-linked (GPI) membrane glycoprotein CD48 (TCT-1, Blast-1). Various studies demonstrated that the murine CD48 is the predominant counterreceptor for the mouse CD2 and is involved in the regulation of T cell activation. Since the murine CD48 is functionally homologous to the human CD2 ligand LFA-3 (CD48), the function of the human CD48 remains unknown. In this report, we show that both Hu-IFN-alpha/beta and Hu-IFN-gamma increase the level of CD48 mRNA and upregulate the expression of CD48 proteins at the surface of various cultured human cell lines. However, the IFN have no effect on the expression of LFA-3. In addition, we show that IFN increase CD48 expression on peripheral blood mononuclear CD3+, CD14+, and CD19+ subpopulations. These data suggest that in addition to modulation of the conventional MHC class I and class II-restricted interactions, the IFN might promote MHC-unrestricted interactions of target cells with the immune cells by inducing the expression of the cell surface CD48 molecule

Exploring mammalian genome within phase-separated nuclear bodies: Experimental methods and implications for gene expression

The importance of genome organization at the supranucleosomal scale in the control of gene expression is increasingly recognized today. In mammals, Topologically Associating Domains (TADs)andtheactive/inactivechromosomalcompartmentsaretwoofthemainnuclearstructuresthat contribute to this organization level. However, recent works reviewed here indicate that, at specific loci, chromatin interactions with nuclear bodies could also be crucial to regulate genome functions, in particular transcription. They moreover suggest that these nuclear bodies are membrane-less organelles dynamically self-assembled and disassembled through mechanisms of phase separation. We have recently developed a novel genome-wide experimental method, High-salt Recovered Sequences sequencing (HRS-seq), which allows the identification of chromatin regions associated with large ribonucleoprotein (RNP) complexes and nuclear bodies. We argue that the physical nature of such RNP complexes and nuclear bodies appears to be central in their ability to promote efficient interactions between distant genomic regions. The development of novel experimental approaches,includingourHRS-seqmethod,isopeningnewavenuestounderstandhowself-assembly of phase-separated nuclear bodies possibly contributes to mammalian genome organization and gene expression

Opposite Effects of Transforming Growth Factor-β Activation and Rho-Associated Kinase Inhibition on Human Trophoblast Migration in a Reconstituted Placental-Endometrial Coculture System

International audiencePlacental implantation involves highly regulated trophoblast invasion of the endometrial stroma. TGF is a known regulator of this process. This study examines the effect of TGF on extravillous cytotrophoblastic cell (EVCT) migration in cocultures of first-trimester human chorionic villus explants and primary human endometrial fibroblasts. Migration of EVCTs was followed by phase-contrast time-lapse micros-copy and was shown to highly depend on the endometrial fibroblast matrix. Interstitial EVCT invasion was also analyzed by confocal microscopy of fluorescently prelabeled trophoblasts and endometrial fibroblasts. As expected, addition of TGF led to inhibition of EVCT invasion of the endometrial cell layer. This inhibition was characterized by formation of compact EVCT stacks at migration fronts and displacement of endometrial fibroblasts. We tested the role of the RhoA/Rho-associated kinase (ROCK) pathway, a TGF-dependent pathway known to regulate cell migration. Interestingly, blocking ROCK with the chemical in-hibitor Y27632 had an effect opposite to TGF activation because it promoted superficial EVCT migration on the en-dometrial cell layer. These data suggest a role for ROCK in the TGF-dependent control of trophoblast migration. Furthermore , they indicate that even though ROCK signaling plays a role in human trophoblast cell invasion, EVCT migration can still occur in the absence of ROCK activity. (Endocrinology 149: 4475– 4485, 2008

Exploring Mammalian Genome within Phase-Separated Nuclear Bodies: Experimental Methods and Implications for Gene Expression

International audienceThe importance of genome organization at the supranucleosomal scale in the control of gene expression is increasingly recognized today. In mammals, Topologically Associating Domains (TADs) and the active/inactive chromosomal compartments are two of the main nuclear structures that contribute to this organization level. However, recent works reviewed here indicate that, at specific loci, chromatin interactions with nuclear bodies could also be crucial to regulate genome functions, in particular transcription. They moreover suggest that these nuclear bodies are membrane-less organelles dynamically self-assembled and disassembled through mechanisms of phase separation. We have recently developed a novel genome-wide experimental method, High-salt Recovered Sequences sequencing (HRS-seq), which allows the identification of chromatin regions associated with large ribonucleoprotein (RNP) complexes and nuclear bodies. We argue that the physical nature of such RNP complexes and nuclear bodies appears to be central in their ability to promote efficient interactions between distant genomic regions. The development of novel experimental approaches, including our HRS-seq method, is opening new avenues to understand how self-assembly of phase-separated nuclear bodies possibly contributes to mammalian genome organization and gene expression

Endogenous Retroviral Sequences Behave as Putative Enhancers Controlling Gene Expression through HP1-Regulated Long-Range Chromatin Interactions

International audienceAbout half of the mammalian genome is constituted of repeated elements, among which endogenous retroviruses (ERVs) are known to influence gene expression and cancer development. The HP1 (Heterochromatin Protein 1) proteins are known to be essential for heterochromatin establishment and function and its loss in hepatocytes leads to the reactivation of specific ERVs and to liver tumorigenesis. Here, by studying two ERVs located upstream of genes upregulated upon loss of HP1, Mbd1 and Trim24, we show that these HP1-dependent ERVs behave as either alternative promoters or as putative enhancers forming a loop with promoters of endogenous genes depending on the genomic context and HP1 expression level. These ERVs are characterised by a specific HP1-independent enrichment in heterochromatin-associated marks H3K9me3 and H4K20me3 as well as in the enhancer-specific mark H3K4me1, a combination that might represent a bookmark of putative ERV-derived enhancers. These ERVs are further enriched in a HP1-dependent manner in H3K27me3, suggesting a critical role of this mark together with HP1 in the silencing of the ERVs, as well as for the repression of the associated genes. Altogether, these results lead to the identification of a new regulatory hub involving the HP1-dependent formation of a physical loop between specific ERVs and endogenous genes

A novel function for Cyclin A2: control of cell invasion via RhoA signaling.

International audienceCyclin A2 plays a key role in cell cycle regulation. It is essential in embryonic cells and in the hematopoietic lineage yet dispensable in fibroblasts. In this paper, we demonstrate that Cyclin A2-depleted cells display a cortical distribution of actin filaments and increased migration. These defects are rescued by restoration of wild-type Cyclin A2, which directly interacts with RhoA, or by a Cyclin A2 mutant unable to associate with Cdk. In vitro, Cyclin A2 potentiates the exchange activity of a RhoA-specific guanine nucleotide exchange factor. Consistent with this, Cyclin A2 depletion enhances migration of fibroblasts and invasiveness of transformed cells via down-regulation of RhoA activity. Moreover, Cyclin A2 expression is lower in metastases relative to primary colon adenocarcinoma in matched human tumors. All together, these data show that Cyclin A2 negatively controls cell motility by promoting RhoA activation, thus demonstrating a novel Cyclin A2 function in cytoskeletal rearrangements and cell migration

High-salt Recovered Sequences are associated with the active chromosomal compartment and with large ribonucleoprotein complexes including nuclear bodies

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