Development and ultrastructure of the rigid dorsal and flexible ventral cuticles of the elytron of the red flour beetle, Tribolium castaneum Mi Young Noh a , Subbaratnam Muthukrishnan b , Karl J. Kramer b , Yasuyuki Arakane a, * a Department of Applied Biology, Chonnam National University, Gwangju 500-757, South Korea b Department of Biochemistry and Molecular Biophysics, Kansas State University, Manhattan, KS 66506, USA article info Article history: Received 13 August 2017 Received in revised form 2 November 2017 Accepted 2 November 2017 Available online 5 November 2017 Keywords: Tribolium castaneum Elytron Cuticle/exoskeleton Chitin Transmission electron microscopy (TEM) Pore canal abstract Insect exoskeletons are composed of the cuticle, a biomaterial primarily formed from the linear and relatively rigid polysaccharide, chitin, and structural proteins. This extracellular material serves both as a skin and skeleton, protecting insects from environmental stresses and mechanical damage. Despite its rather limited compositional palette, cuticles in different anatomical regions or developmental stages exhibit remarkably diverse physicochemical and mechanical properties because of differences in chemical composition, molecular interactions and morphological architecture of the various layers and sublayers throughout the cuticle including the envelope, epicuticle and procuticle (exocuticle and endocuticle). Even though the ultrastructure of the arthropod cuticle has been studied rather extensively, its temporal developmental pattern, in particular, the synchronous development of the functional layers in different cuticles during a molt, is not well understood. The beetle elytron, which is a highly modified and sclerotized forewing, offers excellent advantages for such a study because it can be easily isolated at precise time points during development. In this study, we describe the morphogenesis of the dorsal and ventral cuticles of the elytron of the red flour beetle, Tribolium castaneum, during the period from the 0 d- old pupa to the 9 d-old adult. The deposition of exocuticle and mesocuticle is substantially different in the two cuticles. The dorsal cuticle is four-fold thicker than the ventral. Unlike the ventral cuticle, the dorsal contains a thicker exocuticle consisting of a large number of horizontal laminae and vertical pore canals with pore canal fibers and rib-like veins and bristles as well as a mesocuticle, lying right above the enodcuticle. The degree of sclerotization appears to be much greater in the dorsal cuticle. All of these differences result in a relatively thick and tanned rigid dorsal cuticle and a much thinner and less pig- mented membrane-like ventral cuticle. © 2017 Elsevier Ltd. All rights reserved. 1. Introduction Molting of the exoskeleton or cuticle is an essential develop- mental process for many invertebrate species in which the extra- cellular matrix or cuticle is replaced and remodeled during growth. The insect cuticle is produced by the underlying epidermal cells of ectodermal origin. It provides protection against mechanical injury, pathogens and parasites, while also providing sensory perception, muscle attachment points, coloration, camouflage and other physiological functions. Since the cuticle is present at all develop- mental stages and essentially covers the entire body, it must exhibit great diversity in its physicochemical and mechanical properties to allow for growth and to accommodate the functions of the tissues and organs that are protected. Thus cuticles from different parts of the insect's anatomy or the same cuticle present at different developmental stages may have widely different mechanical properties that arise most likely from differences in chemical composition and molecular interactions, as well as the thickness and arrangement of morphologically distinct layers within the cuticle. Insect cuticle is composed primarily of chitin, which is a rather rigid polysaccharide that accounts for much of the mechanical strength of the cuticle, and a whole assortment of proteins, some structural many of which bind to chitin and others are enzymes that catalyze chitin deacetylation, tanning or cross-linking of structural proteins to one another. The ultrastructure of the cuticle has been studied extensively by optical microscopy, scanning * Corresponding author. Department of Applied Biology, Chonnam National University, 300 Yongbong-dong, Buk-gu, Gwangju 500-757, South Korea. E-mail address: arakane@chonnam.ac.kr (Y. Arakane). Contents lists available at ScienceDirect Insect Biochemistry and Molecular Biology journal homepage: www.elsevier.com/locate/ibmb https://doi.org/10.1016/j.ibmb.2017.11.003 0965-1748/© 2017 Elsevier Ltd. All rights reserved. Insect Biochemistry and Molecular Biology 91 (2017) 21e33