ABSTRACT
Bacterial symbionts, such as Wolbachia species, can manipulate the sexual development and reproduction of their insect hosts. For example, Wolbachia infection induces male-specific death in the Asian corn borer Ostrinia furnacalis by targeting the host factor Masculinizer (Masc), an essential protein for masculinization and dosage compensation in lepidopteran insects. Here we identify a Wolbachia protein, designated Oscar, which interacts with Masc via its ankyrin repeats. Embryonic expression of Oscar inhibits Masc-induced masculinization and leads to male killing in two lepidopteran insects, O. furnacalis and the silkworm Bombyx mori. Our study identifies a mechanism by which Wolbachia induce male killing of host progeny.
Subject(s)
Bombyx , Moths , Wolbachia , Male , Animals , Wolbachia/metabolism , Bombyx/genetics , Bombyx/metabolism , Moths/microbiology , Dosage Compensation, Genetic , Insect Proteins/genetics , Insect Proteins/metabolismABSTRACT
Bombyx mori Masculinizer protein (BmMasc) is essential for both masculinization and dosage compensation in B. mori. We previously identified a bipartite nuclear localization signal (NLS) of BmMasc and two essential residues (lysine at 274 [K274] and arginine at 275 [R275]) implicated in its function. Sequence comparison showed the presence of putative NLSs in lepidopteran Masc proteins, but their functional properties and critical residues are unknown. Here we characterized a putative NLS of Ostrinia furnacalis Masc (OfMasc) using B. mori ovary-derived BmN-4 cell line. Deletion and alanine scanning mutagenesis revealed that a putative NLS is required for nuclear localization of OfMasc. However, mutations at both K227 and R228, which correspond to K274 and R275 of BmMasc, respectively, do not greatly abolish the NLS activity. Additional mutagenesis analysis revealed that triple mutations at K227, R228, and K240 almost completely inhibited OfMasc nuclear localization. These results suggest that lepidopteran Masc proteins possess a common functional NLS, but the critical residues for its activity are different. Moreover, we examined the masculinizing activity of OfMasc derivatives and found that nuclear localization is not required for the masculinizing activity of OfMasc. The results from our studies indicate that lepidopteran Masc proteins function in the cytoplasm to drive masculinizing cascade.
Subject(s)
Moths/genetics , Nuclear Localization Signals , Animals , Bombyx/genetics , Cell Line , Insect Proteins/genetics , Insect Proteins/metabolism , Insecta , Nuclear Localization Signals/genetics , Nuclear Localization Signals/metabolismABSTRACT
Leptin is a multi-functional adipokine in vertebrates. The leptin gene and protein are found in many vertebrates; however, the existence of leptin in birds remains controversial. Here we detected leptin-like activity in avian blood using chicken leptin receptor (chLEPR) and green fluorescent protein (GFP)-fused chicken signal transducer and activator of transcription (chSTAT3) co-expressed in CHO-K1 cells (CHO-chLEPR/STAT3). We validated that rat serum specifically induces intranuclear migration of GFP-fused chSTAT3 (GFP-chSTAT3) in CHO-chLEPR/STAT3 cells, but not in CHO-K1 cells expressing GFP-STAT3 (CHO-chSTAT3) before testing the avian blood samples. Blood of chickens (Gallus gallus), wild jungle crows (Corvus macrorhynchos), and carrion crows (Corvus corone) accumulated the GFP signal into nuclei, and frequency varied in each blood sample. Western blotting showed that chicken and crow blood samples specifically phosphorylated GFP-chSTAT3 in the chLEPR-transfected cells. These results indicate that avian blood contains a leptin-like molecule that specifically binds to LEPR, suggesting that the leptin system is conserved across all vertebrate classes.