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Insect Taiman (Tai) binds to methoprene-tolerant to form a heterodimeric complex, mediating juvenile hormone (JH) signaling to regulate larval development and to prevent premature metamorphosis. Tai also acts as a steroid receptor coactivator of 20-hydroxyecdysone (20E) receptor heterodimer, ecdysone receptor (EcR) and Ultraspiracle (USP), to control the differentiation of early germline cells and the migration of specific follicle cells and border cells in ovaries in several insect species. In holometabolous insects, however, whether Tai functions as the coactivator of EcR/USP to transduce 20E message during larval-pupal transition is unknown. In the present paper, we found that the LdTai mRNA levels were positively correlated with circulating JH and 20E titers in Leptinotarsa decemlineata ; and ingestion of either JH or 20E stimulated the transcription of LdTai. Moreover, RNA interference (RNAi)-aided knockdown of LdTai at the fourth (final) instar stage repressed both JH and 20E signals, inhibited larval growth and shortened larval developing period. The knockdown caused 100% larval lethality due to failure of larval-pupal ecdysis. Under the apolysed larval cuticle, the LdTai RNAi prepupae possessed pupal thorax. In contrast, the process of tracheal ecdysis was uncompleted. Neither JH nor 20E rescued the aforementioned defectives in LdTai RNAi larvae. It appears that Tai mediates both JH and 20E signaling. Our results uncover a link between JH and 20E pathways during metamorphosis in L. decemlineata. [ABSTRACT FROM AUTHOR]

Leptinotarsa decemlineata adults exhibit a season-dependent activity. In spring, post-diapause beetles often fly a long distance from overwintering sites to potato fields. In summer and autumn, the flight ability is sharply reduced. Proline is the main energy substrate of L. decemlineata during flight and proline dehydrogenase (ProDH) catalyzes the first step in proline catabolism. Here we identified a putative LdProDH gene; it had three cDNA isoforms which shared the same 5′UTR and coding region, but differed in the lengths of 3′UTRs (515, 1 092 and 1 242 bp for isoforms-1, -2 and -3, respectively). LdProDH encoded a 616 amino acid protein that showed high sequence similarity to ProDH-like proteins from other insect species. LdProDH was expressed in the third and fourth instars larvae and adults, but not in pupae. Dietary ingestion of bacterially expressed LdProDH-dsRNA by adults significantly decreased its messenger RNA (mRNA) level, and caused an elevation of free proline content in the hemolymph. Further observation revealed that three canonical polyadenylation signals (AATAAA) were tandemly located in the 3′UTR of isoform-3. The first, second and third polyadenylation sites gave rise to isoforms-1, -2 and -3, respectively. Analysis of the genomic DNA uncovered that the three isoforms resulted from alternative polyadenylation. The mRNA level of isoform-1, which expressed at low levels in pre-diapause adults, became abundant in post-diapause beetles. It is indicated that the LdProDH expression is fine-tuned through 3′UTR to control proline catabolism for the season-dependent activity of L. decemlineata adults. [ABSTRACT FROM AUTHOR]

An exploration of novel control strategies for Leptinotarsa decemlineata is becoming more pressing given rapid evolution of insecticide resistance and rise of production loss of potato. Dietary delivery of bacterially expressed double-stranded RNA (dsRNA) is a promising alternative for management. An important first step is to uncover possible RNA-interference (RNAi)-target genes effective against both young and old larvae. Taiman (Tai) is a basic-helix-loop-helix/Per-Arnt-Sim transcription factor that is involved in the mediation of both juvenile hormone (JH) and 20-hydroxyecdysone (20E) signaling. In the present paper, we found that continuous ingestion of ds Tai for three days by third (penultimate)-instar larvae caused approximately 20% larval mortality and 80% pupation failure. The larval lethality resulted from failed cuticle and tracheae shedding, which subsequently reduced foliage consumption and nutrient absorption, and depleted lipid stores. In contrast, pupation failure derived from disturbed JH and 20E signals, and disordered nutrient homeostasis including, among others, inhibition of trehalose metabolism and reduction of chitin content. Knockdown of LdTai caused similar larval lethality and pupation impairment in second and fourth (final) larval instars. Therefore, LdTai is among the most attractive candidate genes for RNAi to control L. decemlineata larvae. Unlabelled Image • DsRNA of LdTaiman (ds Tai) inhibits pupation in 3rd instar larvae. • Ds Tai leads to similar pupation impairment in 2nd and 4th larval instars. • Defects by ds Tai result from disturbed metabolism, and JH and 20E signals. • LdTaiman is a potential target for RNAi to control L. decemlineata larvae. [ABSTRACT FROM AUTHOR]

• Knockdown of LdHR38 inhibits ecdysteroidogenesis in third instar larvae. • Silencing LdHR38 intensify the expression of early ecdysone genes but represses LdFTZ-F1. • Depletion of LdHR38 impairs larval-pupal ecdysis. • Silencing LdHR38 at the fourth instar causes similar but less severe defects. In insects, nuclear receptors (NRs) including EcR (NR1H1), USP (NR2B4), E75 (NR1D3), HR3 (NR1F), HR4 (NR6) and FTZ-F1 (NR5A3) mediate the 20-hydroxyecdysone (20E) signaling cascade to play a critical role during larval metamorphosis. In this present paper, we focused on hormone receptor 38 (HR38) in Leptinotarsa decemlineata , the only insect homolog of the NR4A subclass. RNA interference (RNAi) of LdHR38 in the penultimate (third) instar larvae reduced the expression of an ecdysteroidogenesis gene and declined the titer of 20E. Knockdown of LdHR38 intensified the expression of LdUSP , LdE75 , LdE74 , LdE93 , LdBroad and LdHR3 , whereas repressed the transcription of LdFTZ-F1. Disruption of 20E signaling inhibited chitin biosynthesis in the larval cuticle. Approximately 25% of the LdHR38 RNAi larvae died, around 40% of the resultant larvae remained as prepupae or become deformed pupae. The body surface of the HR38 depleted abnormal prepupae and pupae looked wet, just like the cuticle being covered with a layer of liquid. Moreover, the increase of larval mortality, and the impairment of pupation and emergence exhibited dose-dependent manners. Furthermore, silencing LdHR38 at the final (fourth) instar caused similar but less severe impairment of pupation. Dietary supplement with 20E for the third instar larvae did not rescue the high larval death and only slightly alleviated the low pupation rate in the LdHR38 RNAi hypomorphs. Accordingly, we propose that HR38 is necessary for tune of ecdysteroidogenesis and for mediation of 20E signaling during metamorphosis in L. decemlineata. [ABSTRACT FROM AUTHOR]

Trehalose is proposed to serve multiple physiological roles in insects. However, its importance remains largely unconfirmed. In the present paper, we knocked down either a trehalose biosynthesis gene (trehalose-6-phosphate synthase, LdTPS ) or each of three degradation genes (soluble trehalases LdTRE1a , LdTRE1b or membrane-bound LdTRE2 ) in Leptinotarsa decemlineata by RNA interference (RNAi). Knockdown of LdTPS decreased trehalose content and caused larval and pupal lethality. The LdTPS RNAi survivors consumed a greater amount of foliage, obtained a heavier body mass, accumulated more glycogen, lipid and proline, and had a smaller amount of chitin compared with the controls. Ingestion of trehalose but not glucose rescued the food consumption increase and larval mass rise, increased survivorship, and recovered glycogen, lipid and chitin to the normal levels. In contrast, silencing of LdTRE1a increased trehalose content and resulted in larval and pupal lethality. The surviving LdTRE1a RNAi hypomorphs fed a smaller quantity of food, had a lighter body weight, depleted lipid and several glucogenic amino acids, and contained a smaller amount of chitin. Neither trehalose nor glucose ingestion rescued these LdTRE1a RNAi defects. Silencing of LdTRE1b caused little effects. Knockdown of LdTRE2 caused larval death, increased trehalose contents in several tissues and diminished glycogen in the brain-corpora cardiaca-corpora allata complex (BCC). Feeding glucose but not trehalose partially rescued the high mortality rate and recovered glycogen content in the BCC. It seems that trehalose is involved in feeding regulation, sugar absorption, brain energy supply and chitin biosynthesis in L. decemlineata larvae. [ABSTRACT FROM AUTHOR]