Proteometabolomic characterization of the spider web components Nephila clavipes used in prey capture strategy

Author(s)

Franciele Grego Esteves

• Document type: Master's Dissertation
• Press: Rio Claro. 2017-04-25.
• Institution: Universidade Estadual Paulista (Unesp). Instituto de Biociências. Rio Claro
• Defense date: 2017-03-22
• Advisor: Mario Sergio Palma; José Roberto Aparecido dos Santos Pinto

Abstract

Nephila clavipes belongs to the group of orb-weavings spiders that have developed the ability to synthesize adhesive threads. Such adhesive threads are found in the core circles of the orb-webs, and present oily droplets which in turn contain many vesicles in suspension, entrapping solutions of proteins, peptides and many small-molecular mass compounds. Chemical analysis studies of these droplets identified toxins, saturated fatty acids and even alkaloids. It is speculated that when an insect-prey is trapped by the web, the fatty acids of these droplets aid in the process of destabilizing the insect cuticle allowing the diffusion of the toxins into the prey body. Some studies also reported alkaloids that act as repellents to predators in some webs, and as insect-toxins in others. The presence of these molecules is evidence that the web is not a simple tool for mechanical capture and imprisonment of prey; but rather a complex structure that seems to play a strategic "active" role in capturing its prey. Considering this, the aim of this study was to analyze the ultrastructure, the disposition of the droplets on the silk fibers, and to visualize the presence of lipid vesicles extracted from the web’s droplets of N. clavipes spider, by microscopy. In addition, the richness of the chemical profile of the small-molecular mass compounds in the web-silk was explored through comprehensive two-dimensional gas chromatography coupled to a mass detector; and finally to investigate the richness of proteins present in web-silk and silk-producing glands through in solution proteolytic digestion, liquid chromatography, and mass spectrometry, highlighting the possible toxins involved in the prey paralysis. First, was performed scanning electron microscopy study of the droplets deposited on the web-silk and light microscopy of the suspended lipid vesicles retained within the aqueous contents of the droplets. Posteriorly in the profiling chemical analysis were identified 316 compounds, among these 25 were identified from chemical standards, most of them are hydrocarbons saturated and some fatty acids. This study also demonstrated through repellence bioassays, that some of the fatty acids and hydrocarbons identified in the web's droplets presented potential as repellents to invasive ants. While the palmitic acid presented potential action in the process of dissolution / disintegration superficial layer of cuticle bees, to allow the difusion of toxins into the prey body. In the proteomic analysis a total of 2051 proteins were identified in the web-silk, of which 163 of these proteins are toxins. A total of 927, 1961, 849, and 860 proteins were also identified in the aggregate, major ampullate, flagelliform and minor ampullate glands, respectively; and of these totals, 194, 78, 32 and 30 are toxins belonging to each gland mentioned above, respectively. This study suggests that silkproducing glands, especially the aggregate gland, can synthesize and deposit on the silk-web important toxins, which are common to some animal venoms, thereby making the webs an active structure in the prey capture. From the obtained results it was possible to elaborate a hypothesis that relates the role of the droplets, lipid vesicles and the compounds identified as part of the chemical strategy of the web in the pre-digestion and prey paralysis. Thus, this study provided a better understanding of the chemical-ecological prey capture by N. clavipes spider web, in addition to information that may enable the use of these compounds in the development of insecticide-selective or even possible pharmacological applications. (AU)

• FAPESP's process: 15/14220-8 - Proteometabolomic characterization of the spider web components Nephila clavipes used in prey capture strategy
• Grantee: Franciele Grego Esteves
• Support Opportunities: Scholarships in Brazil - Master