Insect Bacterial Symbiont-Mediated Vitellogenin Uptake into Oocytes To Support Egg Development

Abstract

Many insect species, such as aphids, leafhoppers, planthoppers, and whiteflies harbor obligate bacterial symbionts that can be transovarially transmitted to offspring through the oocytes of female insects. Whether obligate bacterial symbionts can carry important molecules/resources to the embryos to support egg development is still unknown. Here, we show that the vitellogenin (Vg) precursor of rice leafhopper Nephotettix cincticeps is biosynthesized by the fat body, secreted into the hemolymph and subsequently cleaved into the 35- and 178-kDa subunits, whereas only the 178-kDa subunit is taken up by the leading end of oocytes in a receptor-dependent manner or moves into the posterior pole of the terminal oocyte in association with obligate bacterial symbiont “Candidatus Nasuia deltocephalinicola” (hereafter Nasuia) in a receptor-independent manner. Furthermore, the 178-kDa Vg subunit can directly interact with a surface channel molecule (porin) on the envelope of Nasuia, allowing Vg to enter bacterial cytoplasm. Thus, Vg can hitchhike the ancient oocyte entry path of Nasuia, the common obligate symbiont of leafhoppers. Knocking down a Nasuia growth-related protein expression or treatment with porin antibody strongly prevents the ability of Nasuia to carry Vgs into oocytes and impair insect egg development. Nasuia-carried Vgs provide at least 20% of the total Vgs in the developing eggs. We anticipate that the bacterial symbiont-mediated Vg uptake into oocytes to support efficient egg development may be a common pattern shared by many insects. IMPORTANCE Many insects harbor obligate bacterial symbionts that can be vertically transmitted to offspring by female insects through eggs. Here, we report that leafhopper vitellogenin (Vg) recognizes and binds a surface channel molecule (porin) on the envelope of obligate bacterial symbiont Nasuia, which potentially induces the opening of porin channels for Vg to access the cytoplasm of Nasuia. Thus, Vg can exploit bacterial symbionts as the independent carriers into the oocytes. Such Nasuia-carried Vg contents support efficient insect egg development. Thus, our findings indicate that insects have evolved strategies to exploit the symbionts for carrying additional Vgs to guarantee optimal insect reproduction.

Roles of Bacterial Symbionts in Transmission of Plant Virus by Hemipteran Vectors

“…Consequently, Sodalis and Arsenophonus can directly enter the cytoplasms of obligate bacteria to ensure their simultaneous transmission through insect generations (Michalik et al, 2014;Kobiałka et al, 2016). In some leafhoppers, Vg is incorporated into obligate symbionts, after which bacterial symbionts are exploited as independent carriers into oocytes (Mao et al, 2020). In conclusion, some insect components and microorganisms in insects can use obligate symbiotic bacteria as transporters and hitchhike the existing pathways to facilitate the entry of bacterial symbionts into insect oocytes.…”

The Long and Winding Road for Symbiont and Yolk Protein to Host Oocyte

“…In this context, a totally unexpected phenomenon was recently reported by Mao et al (16) in the rice green leafhopper Nephotettix cincticeps. The insect is obligatorily associated with dual genome-reduced endosymbiotic bacteria, Sulcia and Nasuia, in paired bacteriomes in the abdomen (17).…”

“…Mao et al's (16) finding is particularly intriguing in the following respects. First, this study uncovers a previously unrecognized route of Vg transport to oocytes in insect development.…”

“…Whether Nasuia-allied endosymbiont lineages, such as Zinderia and Vidania (18), also incorporate Vg or not should be inspected in future studies. Finally, I point out that although Mao et al (16) suggest that Vg-porin interaction activates the porin channel to open and allow Vg to enter the Nasuia cytoplasm, this claim seems dubious on the ground that porins have threshold sizes of transportable molecules, which are generally Downloaded from no more than 600 Da (20). Since Vg in Nephotettix oocytes is 178 kDa in size, the idea of porin-mediated Vg transfer into Nasuia cells should be critically reconsidered.…”

“…Sulcia and Nasuia jointly synthesize essential amino acids for their leafhopper hosts and have cospeciated with them over evolutionary time, with Sulcia being more ancient than Nasuia (18). Using thorough histological, biochemical, and molecular genetic approaches, Mao et al (16) demonstrated that (i) both Sulcia and Nasuia enter the posterior region of developing oocytes at the vitellogenic stage, when Vg is actively taken up by the germarium and then transported to the oocyte via a nutritive cord, (ii) in addition to the Vg localization spanning from germarium through nutritive cord to ooplasm, a considerable amount of Vg is detected in the posterior region of the oocytes, where Vg signals colocalize with Nasuia cells, (iii) confocal imaging and immunoelectron microscopy confirm that, strikingly, Vg is present inside the oocyte-invading Nasuia cells, (iv) RNA interference knockdown of VgR does not affect invasion of Nasuia-Vg complex into oocytes, uncovering that the Nasuia-associated Vg incorporation into oocytes is independent of the conventional VgR-mediated Vg transport mechanism, (v) yeast two-hybrid assay and glutathione Stransferase pulldown assay showed interaction between Vg and Nasuia's porin, the major bacterial outer membrane channel protein, (vi) experimental suppression of Nasuia infection to oocytes results in reduced ovarial Vg, (vii) blocking of Nasuia's porin by antibody injection causes reduced Vg titer in the oocytes and suppressed egg development and hatching, (viii) the Nasuia-Vg colocalization is also observed in other leafhopper species, and (ix) based on these results, it is proposed that the porin-mediated incorporation of Vg into the oocyte-infecting Nasuia cells constitutes another Vg transport system in addition to the conventional VgR-mediated Vg transport system in the leafhoppers.…”