BmCalpains are involved in autophagy and apoptosis during metamorphosis and after starvation in Bombyx mori

Abstract

Apoptosis and autophagy play crucial roles during Bombyx mori metamorphosis and in response to various adverse conditions, including starvation. Recently, calpain, one of the major intracellular proteases, has been reported to be involved in apoptosis and autophagy in mammals. BmATG5 and BmATG6 have been identified to mediate apoptosis following autophagy induced by 20-hydroxyecdysone and starvation in B. mori. However, B. mori calpains and their functions remain unclear. In this study, phylogenetic analysis of calpains from B. mori, Drosophila melanogaster and Homo sapiens were performed and the results showed distinct close relationships of BmCalpain-A/B with DmCalpain-A/B, BmCalpain-C with DmCalpain-C, and BmCalpain-7 with HsCalpain-7. Then, the expression profiles of BmCalpains were analyzed by quantitative real-time polymerase chain reaction, and results showed that expression of BmCalpain-A/B, BmCalpain-C and BmCalpain-7 was significantly increased during B. mori metamorphosis and induced in the fat body and midgut of starved larvae, which is consistent with the expression profiles of BmAtg5, BmAtg6 and BmCaspase-1. Moreover, the apoptosis-associated cleavage of BmATG6 in Bm-12 cells was significantly enhanced when BmCalpain-A/B and BmCalpain-7 were induced by starvation, and was partially inhibited by the inhibitor of either calpain or caspase, but completely inhibited when both types of inhibitors were applied together. Our results indicated that BmCalpains, including BmCalpain-A/B, -C and -7, may be involved in autophagy and apoptosis during B. mori metamorphosis and after starvation, and may also contribute to the apoptosis-associated cleavage of BmATG6.

Cell death during complete metamorphosis

“…Different Atg proteins have been suggested as molecular links between autophagy and apoptosis in Lepidoptera. In B. mori cell lines, it was recently proposed that the induction of apoptosis, which follows an autophagic response triggered by 20E or starvation, could be mediated by BmAtg5 and BmAtg6 after their cleavage by calpains [78,79]. Although the association of cleaved BmAtg5 to anti-apoptotic Bcl-2 family proteins to trigger cytochrome c release from mitochondria and caspase activation (similarly to that observed in mammals) has not been investigated yet, this mechanism could explain why 20E and/or starvation cause cell death in the larval organs of Lepidoptera during metamorphosis, with autophagy preceding apoptosis [79].…”

Identification of four caspase genes from Spodoptera exigua (Lepidoptera: Noctuidae) and their regulations toward different apoptotic stimulations

“…Apoptosis is an essential mechanism required by the development of insects, especially involved in the histolysis during metamorphosis (Yi et al ., 2018). The detection of stage‐specific expression patterns of SeCasps showed the possible roles these four SeCasps participated in the S. exigua development (Fig.…”

BmCaspase‐8‐like regulates autophagy by suppressing BmDREDD‐mediated cleavage of BmATG6

“…In holometabolous insects, autophagy is coordinated with apoptosis during metamorphosis. Both are required to accomplish PCD in organs that break down during larval-pupal-adult transition and may not form in adults, such as the salivary gland in Drosophila melanogaster and silk glands and midgut in Bombyx mori (Denton et al, 2009;Franzetti et al, 2012;Denton et al, 2013;Montali et al, 2017;Yi et al, 2018). Changes in morphological and biochemical features as well as the differential display of molecular markers demonstrate that starvation or elevated levels of molting hormone, 20hydroxyecdysone (20E), can trigger autophagy prior to or concomitantly with apoptosis (Goncu & Parlak, 2008;Goncu & Parlak, 2009;Liu et al, 2015;Romanelli et al, 2016), suggesting the sequential activation of PCDrelated molecules and a crosstalk between autophagy and apoptotic pathways.…”