The Glucuronyltransferase GlcAT-P Is Required for Stretch Growth of Peripheral Nerves in Drosophila Rahul Pandey, Jorge Blanco, Gerald Udolph* Neural Development and Repair, Institute of Medical Biology, Singapore, Singapore Abstract During development, the growth of the animal body is accompanied by a concomitant elongation of the peripheral nerves, which requires the elongation of integrated nerve fibers and the axons projecting therein. Although this process is of fundamental importance to almost all organisms of the animal kingdom, very little is known about the mechanisms regulating this process. Here, we describe the identification and characterization of novel mutant alleles of GlcAT-P, the Drosophila ortholog of the mammalian glucuronyltransferase b3gat1. GlcAT-P mutants reveal shorter larval peripheral nerves and an elongated ventral nerve cord (VNC). We show that GlcAT-P is expressed in a subset of neurons in the central brain hemispheres, in some motoneurons of the ventral nerve cord as well as in central and peripheral nerve glia. We demonstrate that in GlcAT-P mutants the VNC is under tension of shorter peripheral nerves suggesting that the VNC elongates as a consequence of tension imparted by retarded peripheral nerve growth during larval development. We also provide evidence that for growth of peripheral nerve fibers GlcAT-P is critically required in hemocytes; however, glial cells are also important in this process. The glial specific repo gene acts as a modifier of GlcAT-P and loss or reduction of repo function in a GlcAT-P mutant background enhances VNC elongation. We propose a model in which hemocytes are required for aspects of glial cell biology which in turn affects the elongation of peripheral nerves during larval development. Our data also identifies GlcAT-P as a first candidate gene involved in growth of integrated peripheral nerves and therefore establishes Drosophila as an amenable in-vivo model system to study this process at the cellular and molecular level in more detail. Citation: Pandey R, Blanco J, Udolph G (2011) The Glucuronyltransferase GlcAT-P Is Required for Stretch Growth of Peripheral Nerves in Drosophila. PLoS ONE 6(11): e28106. doi:10.1371/journal.pone.0028106 Editor: Michael A. Fox, Virginia Commonwealth University Medical Center, United States of America Received July 22, 2011; Accepted November 1, 2011; Published November 23, 2011 Copyright: 2011 Pandey et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was funded by the Institute of Medical Biology. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the mansucript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: gerald.udolph@gmail.com Introduction During animal development and growth, the nervous system needs to expand in conjunction with the general expansion of the body. Crucial to this process is the extension of integrated nerve fibers, which contain axons connecting motoneurons in the central nervous system (CNS) to their peripheral targets, muscles and sensory neurons in the peripheral nervous system (PNS) connecting to their synaptic partners in the CNS. Neurite outgrowth, via growth-cone mediated mechanisms and growth- cone guidance, towards a target has been extensively studied [1,2]. Current models suggest that during development axonal elongation occurs by extension of the growth cone [3]. However, once the growth cone reaches its final target and is fully connected and thus tightly integrated, growth-cone related axonal elongation mechanisms are unlikely functional any longer. Mechanical forces have been postulated to stimulate elongation of integrated axons as the animal grows [4]. This distinct process has been recently referred to as ‘‘stretch growth of integrated axon tracts’’ [5]. Examples of such extreme axonal stretch growth, where animal growth supplies constant me- chanical tension on nerves and white matter tracts, are observed throughout the animal kingdom [6,7]. As the anterior-posterior body axis of an animal extends during growth, the distance between most neuronal somata in the CNS and PNS and their respective target cells increases dramatically, exerting tensile forces on the axons within the nerves. These forces would normally stimulate the addition of cytoskeletal elements, axolemma and other building materials along the axon to compensate for the mechanical strain. In vitro studies have demonstrated that integrated axon tracts can undergo stretch growth [5,8,9]. Several studies have been reported using explanted neurons from various organisms to understand stretch induced axonal elongation in cell culture in vitro [10,11,12,13,14,15] (reviewed in [16]). However, elongation of integrated axons is not understood at the molecular level particularly at the stage of a developing organism in vivo, partly due to the lack of mutants of genes involved in this process. Drosophila offers an amenable model system to study the process of stretch growth of integrated axons, due to the high growth rate of its larval body during development as well as its compliance to genetic analyses [17]. Importantly, extensive growth is observed during the four days of larval development. During this phase, motoneuron axons that established connectivity to their muscle targets during embryogenesis have to elongate in parallel to the expansion of the larval body. However, very little is known about the cellular and molecular mechanisms controlling the extension of integrated peripheral nerves/axons during Drosophila larval development. Post-translational modifications of proteins are known to be involved in nervous system development [18]. In particular, glycosylation of membrane-targeted and secreted proteins is an PLoS ONE | www.plosone.org 1 November 2011 | Volume 6 | Issue 11 | e28106