Under covering the mystery of hybrid animals

In animals that reproduce by external fertilization (like sea urchins), species-specificity is vital so

that the egg of one species recognizes and fuses with the sperm of the same species.

Consequently, there are not one but several steps during fertilization that require species-

specific recognition for fertilization to proceed. These steps include sperm attraction, initiation of

the acrosomal reaction at the jelly coat, and sperm-egg binding. Interestingly, in mammals that

undergo internal fertilization, there are rare examples of interbreeding between species,

including mules (horse x donkey) and zonkey (zebra x donkey

Consider the following scenario

You are a developmental biologist on vacation in Florence, Italy. Talking to a local while having a gelato, you hear of an animal reserve just outside of the city where a curious animal was born. The animal has a head of a donkey, and the stripes of a zebra on its body and legs! You instantly guess it must be a zebroid, a hybrid between a zebra and another member of the horse family. Like any curious biologist, you drag your partner to the animal reserve to check out this curious zebroid.

When you arrive at the animal reserve, you immediately spot Ippo the zebroid running around her pen, and you go to talk to the caretakers at the reserve. The caretakers tell you that the zebras are kept in a fenced-in pen next to an area where an endangered species of donkey from the Amiata region are kept. One day Martin the zebra jumped the fence and mated with Giada the donkey. Twelve months later, Giada gave birth to Ippo!

Ippo has been such a sensation at the animal reserve that ticket sales to the park have soared. Hearing that you are a biologist, the owner strides over and asks you, how likely is a hybrid event like this to happen again? Could they produce other combinations of animals successfully to keep profits up? Though you are concerned about the ethical implications, your curiosity about how animals can mate to form hybrids is too much to resist. You can't wait to return to your laboratory in Middletown and study this question.

Back in the lab, you begin to read about the mechanism of fertilization. You learn that Deno is a protein expressed on the acrosomal process of mammalian sperm (deno means 'bind' in Greek). In their discussion section, the authors of the paper speculate that Deno may recognize glycoproteins in the zona pellucida, though no evidence is provided in the paper. The protein sequence of Deno is 99% identical between horse, donkey and zebra, and 70% identical between horse, human, mouse and rabbit.

Question 1

Test the hypothesis that Deno is a likely candidate for the recognition of sperm and egg. How would you design an experiment to test this hypothesis?

Question 2

What would you predict to find from your proposed experiment?