Receptor-directed delivery of dsRNA into crustacean oocytes

Oviparous animals are characterized by an elaborated process of yolk production and packaging in the oocytes. The major yolk protein (vitellin) is usually produced as vitellogenin (Vg) outside the egg and internalized into oocytes by receptor-mediated endocytosis. In our crustacean model the prawn M. rosenbergii, like many other crustacean species, Vg is produced in the hepatopancreas. Upon arrival to the oocytes, the Vg-receptor (VgR) interacts with distinct regions of the Vg and internalizes it to form yolk-droplets. One distinctive characteristic of the VgR family is their role in massive internalization of lipoproteins. It is for that reason we predicted that Vg endocytosis could be used as an effective tool for high throughput delivery into the oocyte. Indeed, a specific Vg-derived peptide was found to be capable to enter specifically into oocytes both in vitro and in vivo. Contrary to the above, a control peptide was not able to enter the oocytes. Upon injection into vitellogenic females, only dsRNA conjugated to the Vg-derived peptide was capable of reducing the expression of the corresponding gene in the embryos and larvae of the injected females, while females injected with the dsRNA conjugated to the control peptide revealed no silencing. Shrimp and prawn aquaculture has become a significant worldwide industry, with an annual market volume of above $57 billion worldwide. The market is in need for fast growing, sustainable, vaccinated populations. A single treated female would produce thousands of affected embryos with the desired trait. Therefore, a delivery tool of dsRNA for short term silencing or CRISPR-CAS9 complex for gene editing, or vaccination by small molecules directly into the oocytes could make a huge impact on future aquaculture.