Small RNA and mRNA profiling during intersegmental muscle development and programmed cell death in Manduca sexta

PIWI-interacting RNAs (piRNAs) are small single-stranded RNAs that can repress transposon expression via epigenetic silencing and transcript degradation. They have been identified predominantly in the ovary and testis where they serve essential roles in transposon silencing in order to protect the integrity of the germline genome. The potential expression of piRNAs in non-stem somatic cells has been controversial. In the present study we demonstrate the expression of piRNAs derived from both genic and transposon RNAs in the intersegmental muscles (ISMs) from the tobacco hawkmoth Manduca sexta. These piRNAs are abundantly expressed, are ~27 nt long, map antisense to transposons, are oxidation resistant, exhibit a uridine bias at their first nucleotide, and amplify via the canonical ping-pong pathway. An RNA-seq analysis demonstrated that 17 piRNA pathway genes are expressed in the ISMs and are developmentally regulated. The expression levels of piRNAs do not change when the muscles undergo developmentally regulated atrophy, but are repressed when muscles become committed to undergo programmed cell death at the end of metamorphosis. This change in piRNA expression is associated with the targeted repression of several retrotransposons and the induction of certain DNA transposons. Developmental changes piRNAs, piRNA pathway genes, and transposon expression are all regulated by 20-hydroxyecdysone, the steroid that controls the timing of ISM death. Taken together, these data provide compelling evidence for the existence of piRNA in somatic tissues and suggest that they may play a role in developmental processes such as programmed cell death. Analyses of 8 small RNA-seq samples, 1 oxidized small RNA-seq sample, and 21 mRNA-seq samples with 3 replicates from intersegmental muscles during pupal-adult development in Manduca sexta