Transcriptional Profiling of Midgut Immunity Response and Degeneration in the Wandering Silkworm, Bombyx mori Qiuyun Xu1., Anrui Lu1., Guohua Xiao 1 , Bing Yang1, Jie Zhang 1 , Xuquan Li1, Jingmin Guan 1 , 1 2 3 1 1 Qimiao Shao , Brenda T. Beerntsen , Peng Zhang , Chengshu Wang , Erjun Ling * 1 Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People’s Republic of China, 2 Department of Veterinary Pathobiology, University of Missouri, Columbia, Missouri, United States of America, 3 National Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, People’s Republic of China Abstract Background: Lepidoptera insects have a novel development process comprising several metamorphic stages during their life cycle compared with vertebrate animals. Unlike most Lepidoptera insects that live on nectar during the adult stage, the Bombyx mori silkworm adults do not eat anything and die after egg-laying. In addition, the midguts of Lepidoptera insects produce antimicrobial proteins during the wandering stage when the larval tissues undergo numerous changes. The exact mechanisms responsible for these phenomena remain unclear. Principal Findings: We used the silkworm as a model and performed genome-wide transcriptional profiling of the midgut between the feeding stage and the wandering stage. Many genes concerned with metabolism, digestion, and ion and small molecule transportation were down-regulated during the wandering stage, indicating that the wandering stage midgut loses its normal functions. Microarray profiling, qRT-PCR and western blot proved the production of antimicrobial proteins (peptides) in the midgut during the wandering stage. Different genes of the immune deficiency (Imd) pathway were up- regulated during the wandering stage. However, some key genes belonging to the Toll pathway showed no change in their transcription levels. Unlike butterfly (Pachliopta aristolochiae), the midgut of silkworm moth has a layer of cells, indicating that the development of midgut since the wandering stage is not usual. Cell division in the midgut was observed only for a short time during the wandering stage. However, there was extensive cell apoptosis before pupation. The imbalance of cell division and apoptosis probably drives the continuous degeneration of the midgut in the silkworm since the wandering stage. Conclusions: This study provided an insight into the mechanism of the degeneration of the silkworm midgut and the production of innate immunity-related proteins during the wandering stage. The imbalance of cell division and apoptosis induces irreversible degeneration of the midgut. The Imd pathway probably regulates the production of antimicrobial peptides in the midgut during the wandering stage. Citation: Xu Q, Lu A, Xiao G, Yang B, Zhang J, et al. (2012) Transcriptional Profiling of Midgut Immunity Response and Degeneration in the Wandering Silkworm, Bombyx mori. PLoS ONE 7(8): e43769. doi:10.1371/journal.pone.0043769 Editor: Christian Scho¨ nbach, Kyushu Institute of Technology, Japan Received March 31, 2012; Accepted July 25, 2012; Published August 24, 2012 Copyright: 2012 Xu 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 supported by the National Basic Research Program of China (2012CB114605), National Natural Science Foundation of China (30970408, 31172151), Ministry of Agriculture of China (2011ZX08009-003, 2012ZX08011001-004), Chinese Academy of Sciences (KSCX2-EW-J-12). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: erjunling@sippe.ac.cn . These authors contributed equally to this work. Introduction Insects such as Drosophila melanogaster live on rotten fruit and food containing many microbes, yet they still survive. The insect midgut provides innate immunity during the feeding stage against many pathogens ingested with their food. Under the delicate control of the midgut innate immune system, the pathogenic microbes can be specifically eliminated with minimal disruption to commensal and mutualistic bacteria [1,2]. Thus, these insects have evolved an effective defense system, which has become a research focus. The insect gut is a continuous tube that starts from the mouth and ends at the anus. It is composed of three parts: the foregut, midgut, and hindgut. In insects, ingested food is stored and partially digested in the foregut, whereas the midgut is the primary site of digestion and absorption of nutrients. In the hindgut, some water and salts are absorbed to balance the hemolymph osmotic pressure during the process of feces formation [3]. All insects undergo metamorphosis, by which they develop from larvae into adults under the control of juvenile hormone (JH) and 20-hydroxyecdysone (20-E) [4,5]. For insects undergoing incom- plete metamorphosis, they have three stages: egg, nymph, and PLOS ONE | www.plosone.org 1 August 2012 | Volume 7 | Issue 8 | e43769