Fungal extracellular ribotoxins as insecticidal agents

Abstract

Fungal ribotoxins were discovered almost 50 years ago as extracellular ribonucleases (RNases) with antitumoral properties. However, the biological function of these toxic proteins has remained elusive. The discovery of the ribotoxin HtA, produced by the invertebrates pathogen H. thompsonii, revived the old proposal that insecticidal activity would be their long searched function. Unfortunately, HtA is rather singular among all ribotoxins known in terms of sequence and structure similarities. Thus, it was intriguing to answer the question of whether HtA is just an exception or, on the contrary, the paradigmatic example of the ribotoxins function. The work presented uses HtA and -sarcin, the most representative member of the ribotoxins family, to show their strong toxic action against insect larvae and cells.

Characterization of a new toxin from the entomopathogenic fungus Metarhizium anisopliae: the ribotoxin anisoplin

“…Moreover, in order to correlate this effect with ribosome inhibition, RNA from Sf9 cells treated with anisoplin was extracted and analyzed by primer extension ( Figure 3F). Results showed the specific fragment due to ribotoxin cleavage in percentages similar to those obtained with HtA and α-sarcin (Olombrada et al, 2013). Therefore, anisoplin, as a typical ribotoxin, causes cytotoxicity in cultured insect cells by entering into them and specifically cleaving their ribosomes.…”

“…Therefore ribosome recognition seems to be different in these two proteins, at least in terms of efficiency. Important differences in this regard have also been observed in our laboratory between α-sarcin and HtA (Herrero-Galán et al, 2008;Olombrada et al, 2013) (Supplementary Figure 3C and D). Here, however, it cannot be discarded that recombinant anisoplin were less active than its fungal counterpart due to subtle structural disarrangements.…”

“…The specificity here, since they do not interact with any membrane receptor, is the lipid composition of the membrane of the target cell. Recently, it has been shown that ribotoxins display a much higher toxicity against insect than mammalian transformed cells and their insecticidal properties have been described (Olombrada et al, 2013). A similar experiment of protein biosynthesis inhibition with Sf9 insect cells has been performed here ( Figure 3E).…”

“…The main producer is Aspergillus, although the discovery of an entomopathogenic fungus, Hirsutella thompsonii, producing the ribotoxin hirsutellin A (HtA) suggests a wider distribution among fungi as well as their insecticide properties ( Martínez-Ruiz et al, 1999;Herrero-Galán et al, 2008;Olombrada et al, 2013Olombrada et al, , 2014a. Ribotoxins specifically cross some lipid barriers and then efficiently inactivate any kind of ribosome.…”

“…In Aspergillus, they seem to be produced during conidia maturation, most probably as a defense mechanism against predators (Brandhorst and Kenealy, 1992). They have been extensively studied ( Lacadena et al, 2007;García-Ortega et al, 2010;Olombrada et al, 2014a,b) with the most recent studies focused on their antitumor properties conjugated as immunotoxins and their toxicity against insects ( Carreras-Sangrá et al, 2012;Olombrada et al, 2013;Tomé-Amat et al, 2015).…”