The influence of Argonaute proteins on alternative RNA splicing.

Alternative splicing of precursor RNAs is an important process in multicellular species because it impacts several aspects of gene expression: from the increase of protein repertoire to the level of expression. A large body of evidences demonstrates that factors regulating chromatin and transcription impact the outcomes of alternative splicing. Argonaute (AGO) proteins were known to play key roles in the regulation of gene expression at the post-transcriptional level.

10-million-years AGO: argonaute on chromatin in yeast and human, a conserved mode of action?

Whereas in yeast the function and mode of action of nuclear RNAi are well documented, mammalian nuclear RNAi is a matter of debates. Several papers support a role for nuclear Argonaute in alternative splicing. However, the molecular mechanism remains elusive.

Argonaute proteins couple chromatin silencing to alternative splicing.

Argonaute proteins play a major part in transcriptional gene silencing in many organisms, but their role in the nucleus of somatic mammalian cells remains elusive. Here, we have immunopurified human Argonaute-1 and Argonaute-2 (AGO1 and AGO2) chromatin-embedded proteins and found them associated with chromatin modifiers and, notably, with splicing factors. Using the CD44 gene as a model, we show that AGO1 and AGO2 facilitate spliceosome recruitment and modulate RNA polymerase II elongation rate, thereby affecting alternative splicing.

A subset of the histone H3 lysine 9 methyltransferases Suv39h1, G9a, GLP, and SETDB1 participate in a multimeric complex.

Lysine 9 of histone 3 (H3K9) can be mono-, di-, or trimethylated, inducing distinct effects on gene expression and chromatin compaction. H3K9 methylation can be mediated by several histone methyltransferases (HKMTs) that possess mono-, di-, or trimethylation activities. Here we provide evidence that a subset of each of the main H3K9 HKMTs, G9a/KMT1C, GLP/KMT1D, SETDB1/KMT1E, and Suv39h1/KMT1A, coexist in the same megacomplex.

Tandem affinity purification of miRNA target mRNAs (TAP-Tar).

MicroRNAs (miRNAs) bind to Argonaute proteins, and together they form the RISC complex and regulate target mRNA translation and/or stability. Identification of mRNA targets is key to deciphering the physiological functions and mode of action of miRNAs. In mammals, miRNAs are generally poorly homologous to their target sequence, and target identification cannot be based solely on bioinformatics.

Physical and functional interaction between heterochromatin protein 1alpha and the RNA-binding protein heterogeneous nuclear ribonucleoprotein U.

By combining biochemical purification and mass spectrometry, we identified proteins associated with human heterochromatin protein 1alpha (HP1alpha) both in the nucleoplasm and in chromatin. Some of these are RNA-binding proteins, and among them is the protein heterogeneous nuclear ribonucleoprotein U (hnRNP U)/SAF-A, which is linked to chromatin organization and transcriptional regulation. Here, we demonstrate that hnRNP U is a bona fide HP1alpha-interacting molecule.

[Micro-RNAs and muscle differentiation].

Deciphering the mechanisms underlying skeletal muscle differentiation in mammals is an important challenge. Cell differentiation involves complex pathways regulated at both transcriptional and post-transcriptional levels. Recent observations have revealed the importance of small (20-25 base pairs) non-coding RNAs (microRNAs or miRNAs) that are expressed in both lower organisms and in mammals.

Dimer composition and promoter context contribute to functional cooperation between AP-1 and NFAT.

The transcription factors activator protein 1 (AP-1) and nuclear factor of activated T-cells (NFAT) cooperate to induce the expression of cytokines during the immune response. While much is known about the signaling pathways and physical interactions between NFAT and AP-1 dimers following lymphocyte activation, few studies have addressed the role of AP-1 composition in modulating NFAT:AP-1-dependent transcription. We examined the function of specific AP-1 complexes using "tethered" AP-1 dimers with defined composition.

The microRNA miR-181 targets the homeobox protein Hox-A11 during mammalian myoblast differentiation.

Deciphering the mechanisms underlying skeletal muscle-cell differentiation in mammals is an important challenge. Cell differentiation involves complex pathways regulated at both transcriptional and post-transcriptional levels. Recent observations have revealed the importance of small (20-25 base pair) non-coding RNAs (microRNAs or miRNAs) that are expressed in both lower organisms and in mammals.

siRNA as a route to new cancer therapies.

The RNA interference (RNAi) gene-silencing mechanism is induced by double-stranded RNA (dsRNA) and is highly sequence-specific. It is an extremely powerful tool for silencing gene expression in vitro, and might be used as therapy in human pathologies such as cancer, viral infections and genetic disorders. RNAi was initially discovered in plants, but it has become clear that it is also conserved in animal species.

AP-1 dimers regulate transcription of the p14/p19ARF tumor suppressor gene.

Evidence is accumulating about the role of individual AP-1 components in cell proliferation and transformation. Notably, Ras-mediated transformation is characterized by the upregulation of particular AP-1 members, such as c-Jun and Fra-1. The p14/p19ARF tumor suppressor gene is a key link between oncogenic Ras signaling and the p53 pathway.

Role of p53 in the sensitization of tumor cells to apoptotic cell death.

Immunotherapy of cancer has always represented a very attractive fourth-modality therapeutic approach. Over the past few years, advances in the identification of tumor antigens have opened new perspectives and provided new opportunities for a more accurate immunotherapy of cancer. However, when applied to patients with established tumors, it rarely leads to an objective response.

Wild-type p53 induced sensitization of mutant p53 TNF-resistant cells: role of caspase-8 and mitochondria.

In the present study, we have investigated the mechanisms by which the restoration of wild-type (wt) p53 functions in p53 mutant cells increases their susceptibility to the cytotoxic action of tumor necrosis factor (TNF). Our data indicate that the resistance of p53-mutated cl.1001 cells to TNF-induced cell death was not due to a defect in the expression of TRADD and FADD, yet correlated with a reduced caspase-8 activation as well as a deficient mitochondrial membrane permeabilization.