O'Carroll Group

MicroRNAs (miRNAs) are small non-coding RNA molecules that have been identified as potent negative regulators of gene expression. miRNA-mediated gene silencing is executed by the multiprotein RNA-induced silencing complex (RISC). At the core of RISC is an Argonaute (Ago) protein that binds miRNA and executes their function. Using hematopoiesis in mice as a model system to study the physiological function of Ago proteins and the mechanism of miRNA-mediated gene silencing in vivo, we found that Ago2 selectively controls early development of B lymphoid and erythroid cells. We showed that the unique and defining feature of Ago2, the Slicer endonuclease activity, is dispensable for hematopoiesis. Instead, we have identified Ago2 as a key regulator of miRNA homeostasis, with deficiency in Ago2 impairing miRNA biogenesis from precursor- miRNAs. Our current interests regarding RISC now focus on the post-translational regulation of RISC in vivo.Having identified erythroid development as being sensitive tomiRNA dosage, we now strive to mechanistically understand how key miRNA loci regulate terminal erythropoiesis. In addition, we are exploring the contribution of individual miRNAs to the development and homeostasis of spermatogenesis.

Transposable elements are mobile genetic elements that constitute a large fraction of mammalian genomes. In mice, these highly mutagenic elements are epigenetically silenced throughout life to avoid the deleterious effects of transposition. Transposon silencing is established during germ cell development and maintained post-fertilisation in all somatic cells. This epigenetic process of transposons silencing is of fundamental importance for germ cell development and the genomic integrity of the gametes and thus the future soma. Highly conserved ribonuclear particles (RNPs) have evolved that target transposons for DNA methylation and subsequent epigenetic silencing. In the male germline members of the Piwi subclade of the Argonaute family of proteins, Mili and Miwi2 are essential for de novo DNA methylation of transposons and spermatogenesis. BothMili andMiwi2 bind a class of small non-coding RNAs known as Piwi-interacting RNAs (piRNAs) that are believed to act as guides for targeting of the respective RNPs. We currently address several basic questions on the intrinsic mechanism and function of mammalian Piwi proteins function.

Future projects and goals

• Identify miRNAs and their respective targets that control erythropoiesis;
• determine the in vivo significance of post-translation modifications of Ago2;
• explore the function of microRNAs during spermatogenesis;
• understand the mechanism of Miwi2 and Mili in the process of epigenetic transposon silencing.

We currently address our goals using state-of-the-art mouse genetic strategies coupled with high throughput sequencing approaches.