Hirsutellin A: A Paradigmatic Example of the Insecticidal Function of Fungal Ribotoxins

Abstract

The fungal pathogen Hirsutella thompsonii produces an insecticidal protein named hirsutellin A (HtA), which has been described to be toxic to several species of mites, insect larvae, and cells. On the other hand, on the basis of an extensive biochemical and structural characterization, HtA has been considered to be a member of the ribotoxins family. Ribotoxins are fungal extracellular ribonucleases, which inactivate ribosomes by specifically cleaving a single phosphodiester bond located at the large rRNA. Although ribotoxins were brought to light in the 1960s as antitumor agents, their biological function has remained elusive. Thus, the consideration of hirsutellin A, an insecticidal protein, as a singular ribotoxin recalled the idea of the biological activity of these toxins as insecticidal agents. Further studies have demonstrated that the most representative member of the ribotoxin family, α-sarcin, also shows strong toxic action against insect cells. The determination of high resolution structures, the characterization of a large number of mutants, and the toxicity assays against different cell lines have been the tools used for the study of the mechanism of action of ribotoxins at the molecular level. The aim of this review is to serve as a compilation of the facts that allow identification of HtA as a paradigmatic example of the insecticidal function of fungal ribotoxins.

Mitochondrial genome, comparative analysis and evolutionary insights into the entomopathogenic fungus Hirsutella thompsonii

“…As long‐term reliance of mite control on chemical acaricides has caused resistance and public concerns of residues in agroproducts, exploiting biopesticides has been suggested as sustainable alternatives to conventional pesticides (Kumar, ). H. thompsonii is reported to produce phomalactone (Krasnoff and Gupta, ) as well as the protein hirsutellin (Herrero‐Galán et al ., ; ), which are toxic to a variety of agricultural pests. The fungus is safe to human, mammals, predators and parasites, and has been one of the few fungal species that were commercially exploited as bioinsecticides.…”

Minimized natural versions of fungal ribotoxins show improved active site plasticity

“…α-Sarcin, restrictocin, Aspf1, and hirsutellin A (HtA) are the most exhaustively characterized ribotoxins [10,11,[19][20][21][22][23][24][25][26], but many others have been identified and partially characterized in different fungal species [27][28][29][30][31][32][33][34][35]. Most of them show a high degree of conservation, with similar sizes and amino acid sequence identities above 85% [1,18,33].…”

Involvement of loop 5 lysine residues and the N-terminal β-hairpin of the ribotoxin hirsutellin A on its insecticidal activity

“…It is well known the dramatic effect of HtA on the inhibition of in vivo S. frugiperda cells protein biosynthesis (Castaño-Rodríguez et al 2015;Herrero-Galán et al 2013;Olombrada et al 2013). Therefore, this insect cellular line was chosen to evaluate the toxic effect of the mutants studied when employed against intact cells.…”

“…Most of them show a high degree of conservation with sequence identities above 60% and even higher than 85% in many instances. The only exception known so far is hirsutellin A (HtA), another extracellular RNase produced by an invertebrate pathogen, the fungus Hirsutella thompsonii, which has been demonstrated to be a ribotoxin even though it only displays about 25% sequence identity with the previously known members of the family (Figure 1) (Boucias et al 1998;Herrero-Galán et al 2013;Herrero-Galán et al 2008; Martínez-Ruiz et al 1999b;Olombrada et al 2014a). HtA is considerably smaller than the rest of ribotoxins known but still contains the same elements of periodic secondary structure and an identical structural arrangement of the active site residues (Figure 1) (Herrero-Galán et al 2012a; HerreroGalán et al 2012b;Herrero-Galán et al 2008; Martínez-Ruiz et al 1999b; Viegas et al 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 HtA is indeed a well-known insecticidal protein (Boucias et al 1998;Liu et al 1995).…”