The vitellogenin receptor functionality of the migratory locust depends on its phosphorylation by juvenile hormone

Vitellogenin receptor (VgR) plays a pivotal role in ovarian vitellogenin (Vg) uptake and vertical transmission of pathogenic microbes and Wolbachia symbionts. However, the regulatory mechanisms of VgR action as an endocytic receptor and translocation from oocyte cytoplasm to the membrane remain poorly understood. Here, by using the migratory locust Locusta migratoria as a model system, we report that juvenile hormone (JH) promotes VgR phosphorylation at Ser1361 in the second EGF-precursor homology domain. A signaling cascade including GPCR, PLC, extracellular calcium, and PKC-ι is involved in JH-stimulated VgR phosphorylation. This posttranslational regulation is a prerequisite for VgR binding to Vg on the external surface of the oocyte membrane and subsequent VgR/Vg endocytosis. Acidification, a condition in endosomes, induces VgR dephosphorylation along with the dissociation of Vg from VgR. Phosphorylation modification is also required for VgR recycling from oocyte cytoplasm to the membrane. Additionally, VgR phosphorylation and its requirement for Vg uptake and VgR recycling are evolutionarily conserved in other representative insects including the cockroach Periplaneta americana and the cotton bollworm Helicoverpa armigera. This study fills an important knowledge gap of low-density lipoprotein receptors in posttranslational regulation, endocytosis, and intracellular recycling.

RNAi Reveals Role of Insulin-Like Androgenic Gland Hormone 2 (IAG2) in Sexual Differentiation and Growth in Hermaphrodite Shrimp

Insulin-like androgenic gland hormone (IAG) is the most widely known hormone that regulates sexual differentiation in crustaceans. Previously, a transcriptome study described two transcripts of IAGs (Lvit-IAG1 and Lvit-IAG2) in the peppermint shrimp Lysmata vittata, a species characterized by a rare reproductive system of protandric simultaneous hermaphroditism (PSH). Herein, we explored the function of Lvit-IAG2 via RNA interference (RNAi) experiments, and then compared the functional differences between the two IAGs. We demonstrated that Lvit-IAG2 positively regulated the expression of crustacean hyperglycemic hormone (Lvit-CHH) in eyestalk ganglion but exhibited no significant effect on the expression of gonad-inhibiting hormone (Lvit-GIHs) and crustacean female sex hormone (Lvit-CFSHs). Besides, Lvit-IAG2 gene knockdown caused degeneration in appendices masculinae (AM) and suppressed germ cells at the secondary spermatocyte stage. Moreover, silencing the Lvit-IAG2 gene impeded ovarian development, including smaller previtellogenic oocytes, and lower expression of vitellogenin (Lvit-Vg) gene in hepatopancreas and vitellogenin receptor (Lvit-VgR) gene in the ovarian region. Notably, silencing the Lvit-IAG2 gene impeded individual growth of the species. Collectively, findings from this study demonstrate that Lvit-IAG2 and Lvit-IAG1 coordinatively function to modulate sexual differentiation in L. vittata; meanwhile, Lvit-IAG2 stimulates the growth of the PSH species.

20E-mediated regulation of BmKr-h1 by BmKRP promotes oocyte maturation

Background Krüppel homolog 1 (Kr-h1) is a critical transcription factor for juvenile hormone (JH) signaling, known to play a key role in regulating metamorphosis and adult reproduction in insects. Kr-h1 can also be induced by molting hormone 20-hydroxyecdysone (20E), however, the underlying mechanism of 20E-induced Kr-h1 expression remains unclear. In the present study, we investigated the molecular mechanism of Kr-h1 induction by 20E in the reproductive system of a model lepidopteran insect, Bombyx mori. Results Developmental and tissue-specific expression analysis revealed that BmKr-h1 was highly expressed in ovaries during the late pupal and adult stages and the expression was induced by 20E. RNA interference (RNAi)-mediated depletion of BmKr-h1 in female pupae severely repressed the transcription of vitellogenin receptor (VgR), resulting in the reduction in vitellogenin (Vg) deposition in oocytes. BmKr-h1 specifically bound the Kr-h1 binding site (KBS) between − 2818 and − 2805 nt upstream of BmVgR and enhanced the transcription of BmVgR. A 20E cis-regulatory element (CRE) was identified in the promoter of BmKr-h1 and functionally verified using luciferase reporter assay, EMSA and DNA-ChIP. Using pull-down assays, we identified a novel transcription factor B. mori Kr-h1 regulatory protein (BmKRP) that specifically bound the BmKr-h1 CRE and activated its transcription. CRISPR/Cas9-mediated knockout of BmKRP in female pupae suppressed the transcription of BmKr-h1 and BmVgR, resulting in arrested oogenesis. Conclusion We identified BmKRP as a new transcription factor mediating 20E regulation of B. mori oogenesis. Our data suggests that induction of BmKRP by 20E regulates BmKr-h1 expression, which in turn induces BmVgR expression to facilitate Vg uptake and oogenesis.

The direct regulation of Aalbdsx on AalVgR is indispensable for ovarian development in Aedes albopictus

BACKGROUNDAedes albopictus is an important vector with an extensive worldwide distribution. Only female mosquitoes play a significant role in the transmission of pathogens. Doublesex (dsx) is a central nexus gene in the insect somatic sex determination hierarchy.RESULTSIn this study, we characterized the full-length sex-specific splicing forms of the Ae. albopictus dsx gene (Aalbdsx). Then, we identified 15 direct target genes of DSX in adult females using digital gene expression (DGE) combined with qPCR following a chromatin immunoprecipitation assay (ChIP) with specific DSX antibodies. The knockdown of Aalbdsx suppressed ovarian development, and the transcript levels of the Aalbdsx target vitellogenin receptor (VgR) gene decreased, whereas vitellogenin (Vg) expression showed an increase in the fat body. Genes in the major Vg regulatory pathway were also upregulated. Our results suggest that the effects of Aalbdsx RNAi on ovarian development are exerted mainly via VgR rather than Vg.CONCLUSIONThe results of our study not only provide a reference for the further elucidation of the sex determination cascade and comparative analyses of dsx target interactions in mosquitoes but also reveal potential molecular targets for application to the development of sterile male mosquitoes to be released for vector control.

How Crucial is the Functional Pit Organ for the Varroa Mite?

Olfaction as well as gustation, are essential for animal survival, allowing behavioral modulation according to environmental input. We focused our study on an obligate ecto-parasitic mite of honey bees, the Varroa destructor Anderson and Trueman (Parasitiformes, Mesostigmata, Varroidae). By mechanically blocking the main olfactory organ on Varroa forelegs by varnishing with nail polish, we were able to show that other sensory organs cannot significantly compensate chemosensory abilities required for mite’s host selection, identification as well as reproduction. In fact, we found that mites with blocked forelegs had a significantly lower ability to reach a host bee than those with varnished idiosoma and unvarnished control. Furthermore, fewer foreleg blocked mites were feeding on the nurse bees and their reproduction in the brood cells was significantly impaired. The inhibition of reproduction was also reflected in altered expression levels of vitellogenin and vitellogenin receptor genes in foreleg-blocked mites.