Upregulation of E93 Gene Expression Acts as the Trigger for Metamorphosis Independently of the Threshold Size in the Beetle Tribolium castaneum

Abstract

Body size in holometabolous insects is determined by the size at which the juvenile larva undergoes metamorphosis to the pupal stage. To undergo larva-pupa transition, larva must reach a critical developmental checkpoint, the threshold size (TS); however, the molecular mechanisms through which the TS cues this transition remain to be fully characterized. Here, we use the flour beetle Tribolium castaneum to characterize the molecular mechanisms underlying entry into metamorphosis. We found that T. castaneum reaches a TS at the beginning of the last larval instar, which is associated with the downregulation of TcKr-h1 and the upregulation of TcE93 and TcBr-C. Unexpectedly, we found that while there is an association between TS and TcE93 upregulation, it is the latter that constitutes the molecular trigger for metamorphosis initiation. In light of our results, we evaluate the interactions that control the larva-pupa transition and suggest alternative models.

TGF-beta/Activin ligand Myoglianin couples muscle growth to the initiation of metamorphosis

“…A previous study has suggested that threshold size in T. castaneum might be regulated by 413 JH (Chafino et al, 2019). However, in that study, the threshold size was identified by starving 414 larvae at various sizes during the final instar and identifying the size above which the larva initiates metamorphosis.…”

“…In this weight range, hypoxia-treated larvae are above the threshold size 272 whereas 40% diet-fed larvae are below the threshold size. We found that myo expression was 273 significantly higher in the muscles of hypoxia-treated larvae than that of 40% diet-fed larvae 274 Previous studies have suggested that a decline in JH may underlie threshold size 281 attainment in other insects (Chafino et al, 2019). To determine whether JH affects threshold size 282 attainment in M. sexta, larvae were placed in hypoxic conditions during the third instar until 283 HCS and then treated with 10 µg methoprene, a JH analog, two days later.…”

“…to the threshold size and demonstrated that it is associated with an increase in E93 expression 100 (Chafino et al, 2019). While the methodology used to determine the threshold size is different 101 from that used in M. sexta and the identified stage may be more consistent with the attainment of 102 irreversible pupal commitment, it is clear that the decision to become a pupa (and hence the 103 threshold size) is already reached 24 hrs after the molt to the final instar.…”

“…In T. castaneum, knockdown of ventral veins lacking 92 (vvl) leads to precocious metamorphosis by suppressing the production of JH (Cheng et al, 93 2014). Recently, the ecdysone response gene, E93, has been shown to be necessary to terminate 94 JH secretion in order to initiate the onset of metamorphosis; knockdown of E93 leads to the 95 induction of supernumerary molts in T. castaneum (Chafino et al, 2019). Chafino et al (2019) 96 demonstrated that in this species, starvation before day 1 of the final instar can induce 97 supernumerary molts whereas starvation after day 1 does not lead to supernumerary molts.…”

“…Chafino et al (2019) 96 demonstrated that in this species, starvation before day 1 of the final instar can induce 97 supernumerary molts whereas starvation after day 1 does not lead to supernumerary molts. 98 Chafino et al (2019) suggested that the mass on the first day of the final instar likely corresponds 99…”