Soonmoon Yoo, Ph.D. - US grants

DESCRIPTION (provided by applicant): Axonal damage to the central nervous system (CNS) do not spontaneously regenerate because of the inherent low repair capacity and cannot currently be repaired or replaced by any treatment, resulting in devastating and permanent loss of neurophysiological function. By contrast, spontaneous axon regeneration occurs following injury of peripheral nerves, although the regenerative capacity declines as the PNS neurons get maturate. Distinguishing the differences between these and finding the key regulators and related target genes responsible for successful regeneration will bring key insight into the reasons why CNS neurons fail to regenerate. The long-term goal of my research is to understand the molecular mechanisms that induce regenerative response in PNS neurons and to develop new therapeutic strategies for CNS nerve regeneration. Injury to axons in PNS induces rapid and local regenerative responses for which local protein synthesis in axons is essential to initiate damage repair, to form a new growth cone and to generate retrogradely transporting injury signal. Small non-coding RNAs, including microRNAs (miRNA), have recently been recognized as a prominent player in post-transcriptional regulation of local protein synthesis. Although it is now certain that the miRNAs localize into axons and play a role in the coordinated regulation of local protein synthesis in regenerating axons, how these non-coding RNAs translocate into distal process of neurons is completely unknown to date. Interestingly, several recent studies show the presence of Dicer and components of the miRNA-induced silencing complex (miRISC) that are required for processing precursor miRNAs (pre- miRNAs) to mature functional miRNAs both in dendrites and axons. Processing of a pre-miRNA to mature miRNA locally in neuronal processes could confer a unique advantage for coordinately altering the population of proteins generated in growth cones by targeting mRNA cohorts. Here, we propose to characterize the population of pre-miRNAs transported into axons and to determine the RNA structures and the trans-acting factor(s) underlying this transport of pre-miRNA into distal process of sensory neurons. The specific aims of this proposal are: (1) to profile changes of the precursor and mature miRNA expression levels in the axonal compartments following sciatic nerve injury; (2) To identify the cis-acting region(s) of precursor miRNAs and trans-acting factor(s) that are required for localization. On completion of these studies, we will have characterized a novel set of neuronal miRNAs that regulate local protein synthesis in distal axons to promote nerve regeneration. In addition, the proposed work will, for the first time, demonstrate mRNA-independent localization of precursor miRNAs into distal axons and a subsequent local maturation to the mature functional miRNA upon a signal arrival. We also anticipate that these studies should identify new targets that could be manipulated to promote axonal regrowth in various neurological disorders including spinal cord injury and demyelinating neuropathies.

Axons in the peripheral nervous system (PNS) spontaneously regenerate, allowing functional recovery after injury, which is largely absent in adult mammalian central nervous system. Intra- axonal protein synthesis is a key component of spontaneous regenerative responses in the PNS and this can be drastically enhanced in nerves that have been pre-conditioned by a prior injury. Our work has shown selective localization and differential translation of many axonal mRNAs in adult sensory neurons and axons of peripheral nerves. However, it is still not clear how translation of axonal mRNAs is regulated. Our long-term goal of this research is to understand the molecular mechanisms that provide spatial and temporal regulation for intra-axonal protein synthesis. Small non-coding RNAs (sncRNAs) have emerged as key controllers of gene expression in the nervous system. Among these, PIWI-interacting RNAs (piRNAs) are primarily known for their roles in silencing transposable elements in germline cells. Recently, piRNAs are suggested to contribute to neuronal development and function. However, the possibility that the piRNAs are acting locally within neuronal processes to regulate axonal mRNA translation has not been tested. We anticipate that axonal piRNAs could confer a unique advantage for coordinately altering the population of proteins generated in growth cones by targeting mRNA cohorts. Our preliminary work shows that piRNA-like small RNAs (piLRNAs) are present in axons of sciatic nerve and that depletion of MIWI protein, a murine homolog of PIWI, increases axon growth and decreases axon retraction after injury. In this proposal, we hypothesize that the MIWI protein and specific piLRNAs are selectively enriched in axons of neurons and are functionally important for neuronal morphology including axon growth and regeneration following injury. In Aim 1, we will determine whether piLRNAs are selectively enriched in axons of neuronal cells by directly contrasting levels of piLRNAs in axon vs. cell body compartment. In Aim 2, we will focus on piLRNAs-5567, 1199, 5901, 5598, and 5595 to examine whether these axonally enriched piLRNAs control axon growth and/or regeneration. Once completed, we will, for the first time, demonstrate the localization of neuronal piLRNAs in the axon compartment, and identify novel roles of the axonally enriched piLRNAs in sensory neurons in axon growth and/or regeneration.