In the past few years a series of paradigm-shifting discoveries have radically altered our understanding of mechanisms underlying integrated gene expression programmes, including the widespread transcription of non-coding RNAs. In particular, the discovery of the regulatory small RNAs represents a major breakthrough in recent biomedical history.
In our current research, we are investigating the molecular mechanisms responsible for orchestrating and integrating genome-wide transcriptional and post-transcriptional responses upon changes in extracellular cues critical for diverse biological processes, including developmental, physiological, and pathological events. We focus on previously unsuspected non-coding RNA-mediated strategies in regulating gene expression. These strategies, which underlie genome-wide transcriptional and post-transcriptional responses, are controlled by complex networks of protein/RNA interactions, ~50 percent of which are expected to be cell-type specific. Despite their discovery more than 35 years ago, the fundamental principles by which non-coding RNAs are regulated and functional have been poorly understood. Our recent findings have contributed to unravel novel and unexpected biological and biochemical aspects of such a central role of noncoding RNAs in regulated gene expression programmes. Our works also extended the knowledge of non-coding RNAs toward novel therapeutic and diagnostic implications for many common diseases, including atherosclerosis, diabetes, and cancer.
Presently, many questions about RNA biology in gene expression control are still to be fully addressed, including i) how do fate of RNAs ultimately influence their function? ii) what are the biochemical mechanisms underlying the function of noncoding (nc)RNAs? A major challenge for both questions will be to define the components of the ribonucleoprotein complexes that ultimately determine the fate and function of ncRNAs in timing and cell specific fashion. Overall, we foresee that in vivo experiments and OMICS approaches coupled with proteomic and transcriptomic methodologies will be the most exhaustive path to uncover the role of ncRNAs in physio-pathological events of gene expression. experimental methodologies and OMICS
We set up three main axes:
1- Investigation of novel mechanisms of miRNA-dependent gene expression control.
We recently demonstrated a direct role for let-7b/c to regulate KSRP expression, which in turn controls let-7 processing in competition with Lin28, and the turnover rate of the pituitary terminal differentiation marker aGSU leading to an upregulation of its expression. These findings defined a hierarchical mechanism by which during organogenesis the upregulation of specific miRNAs controls the expression of KSRP and the subsequence fine-tuning of the processing for selected miRNAs and decay rate of mRNAs encoding for differentiation markers.
Figure A : model for the miRNA–dependent control of aGSU transcript (Repetto et al., 2012 Plos Genetics).
Currently, we are using proteomic and high-throughput sequencing approaches coupled with bioinformatics to study the protein/miRNA code to orchestrate the post- transcriptional programme(s) of regulated gene expression.
2- Investigation of the function of RNY-derived small RNAs in macrophage foam cells
Using deep sequencing, we demonstrated that the ~110 nucleotides (nt) long Ro-associated Y-RNAs (or RNYs), are processed into small ~30 nt sequences in apoptotic and lipid-laden macrophages. The expression of RNY–derived small RNAs (referred as s-RNYs) is significantly upregulated in serum of 263 patients with coronary artery disease compared to 514 controls. These findings were patented (n° WO2015055857 A1) and published in BMC Medicine in 2015. As follow up of this primary discovery, we have investigated the role of s-RNYs in macrophages. Loss and gain of function experiments show that s-RNYs activate both caspase-dependent cell death and NF-kappaB-dependent inflammation in macrophages. We found that s-RNYs activate Toll-like receptor 7 (TLR7).These data indicate that s-RNYs are intrinsic components of the machinery regulating lipid-laden macrophage phenotype and function as mediator of inflammation/apoptosis. We are currently investigating the protein complex associated to s-RNYs to study the direct and specific mechanism(s) of s-RNYs inflammation and cell death in macrophages.
Figure B : A model for s-RNY mode of action in lipid-laden macrophages exerting signaling pathway effects (Hizir et al., in press Cell Death & Dis).
3- Role of small RNAs in epigenetic inheritance of metabolic pathologies (conducted by Dr. Valerie Grandjean).
Recently, we and others extended the concept of « epigenetic inheritance » to RNA-mediated paternal heredity. We found that mice derived from embryos microinjected with HFD-sperm or -testis RNA, developed metabolic pathologies, such as obesity and diabetes. Interestingly, miR-19b alone was sufficient to mediate the paternal diet-induced heredity, as we demonstrated by microinjecting it into naive one-cell embryo (Figure).
To gain insight into the mechanism of epigenetic inheritance, we are now exploring the following specific aims: i) To identify the epigenetic signal accumulated in sperm upon continuous paternal HFD feeding for multiple generations. ii) To perform a parallel between mice and human, a deep-sequencing analysis of the RNA transcriptome of normal and obese men will be performed.
Figure C : RNA-mediated epigenetic inheritance of High Fat Diet (HFD)-induced obesity. (Grandjean, et al., 2015, Sci Rep)
[Diagnostic value of YRNA-derived small RNAs for coronary artery disease]
Hizir Z, Trabucchi M, Repetto E
Med Sci (Paris) 2016 Mar;32(3):248-51 PMID : 27011242
RNY-derived small RNAs as a signature of coronary artery disease
Repetto E, Lichtenstein L, Hizir Z, Tekaya N, Benahmed M, Ruidavets JB, Zaragosi LE, Perret B, Bouchareychas L, Genoux A, Lotte R, Ruimy R, Ferrières J, Barbry P, Martinez LO, Trabucchi M
BMC Med 2015;13:259 PMID : 26449324
Regulation of stimulus-inducible gene expression in myeloid cells
Saccani S, Trabucchi M
Semin. Immunol. 2015 Feb;27(1):33-43 PMID : 25820127
Let-7b/c enhance the stability of a tissue-specific mRNA during mammalian organogenesis as part of a feedback loop involving KSRP
Repetto E, Briata P, Kuziner N, Harfe BD, McManus MT, Gherzi R, Rosenfeld MG, Trabucchi M
PLoS Genet. 2012;8(7):e1002823 PMID : 22844247
DICER- and AGO3-dependent generation of retinoic acid-induced DR2 Alu RNAs regulates human stem cell proliferation
Hu Q, Tanasa B, Trabucchi M, Li W, Zhang J, Ohgi KA, Rose DW, Glass CK, Rosenfeld MG
Nat. Struct. Mol. Biol. 2012 Nov;19(11):1168-75 PMID : 23064648
PI3K/AKT signaling determines a dynamic switch between distinct KSRP functions favoring skeletal myogenesis
Briata P, Lin WJ, Giovarelli M, Pasero M, Chou CF, Trabucchi M, Rosenfeld MG, Chen CY, Gherzi R
Cell Death Differ. 2012 Mar;19(3):478-87 PMID : 21886180
KSRP, many functions for a single protein
Briata P, Chen CY, Giovarelli M, Pasero M, Trabucchi M, Ramos A, Gherzi R
Front Biosci (Landmark Ed) 2011;16:1787-96 PMID : 21196264
KSRP Promotes the Maturation of a Group of miRNA Precuresors
Trabucchi M, Briata P, Filipowicz W, Ramos A, Gherzi R, Rosenfeld MG
Adv. Exp. Med. Biol. 2011;700:36-42 PMID : 21755471
How to control miRNA maturation?
Trabucchi M, Briata P, Filipowicz W, Rosenfeld MG, Ramos A, Gherzi R
RNA Biol 2009 Nov-Dec;6(5):536-40 PMID : 19875945
LPS induces KH-type splicing regulatory protein-dependent processing of microRNA-155 precursors in macrophages
Ruggiero T, Trabucchi M, De Santa F, Zupo S, Harfe BD, McManus MT, Rosenfeld MG, Briata P, Gherzi R
FASEB J. 2009 Sep;23(9):2898-908 PMID : 19423639
The RNA-binding protein KSRP promotes the biogenesis of a subset of microRNAs
Trabucchi M, Briata P, Garcia-Mayoral M, Haase AD, Filipowicz W, Ramos A, Gherzi R, Rosenfeld MG
Nature 2009 Jun;459(7249):1010-4 PMID : 19458619