Worm sperm protein turns sperm on, then keeps rival sperm off

Intestinal roundworm (Ascaris suum)

Ascaris suum adult. Image via Flickr.

I recently wrote about a paper describing a protease in worm seminal fluid that helps activate male sperm (as opposed to hermaphrodite sperm). Now, another group has published a (sort of) follow-up. They found a protease inhibitor that likely blocks the protease (Try-5) found in the first paper, but in a different species of worm. They also show that it has two roles: one in turning sperm on, the other in turning them off. The research was published ahead of print in PNAS January 31, 2012 (See link below for citation).

Nematodes (a type of worm) have sperm that don’t swim, they crawl toward the egg. The most famous nematode is Caenorhabditis elegans, which has two sexes: hermaphrodite and male. Most of these worms are hermaphrodites, but they will happily mate with rare males. In nematodes, sperm have to be activated before they can crawl around and fertilize an egg. In males, activation happens during ejaculation, and relies on seminal fluid proteins, including Try-5. Try-5 is a protease, or at least it looks like one, meaning its role is to chop up other proteins. We still don’t know what Try-5 is chopping up or how that activates sperm, so all we know is that Try-5 is important somehow.

The authors of this paper looked at a different nematode species to understand sperm activation: Ascaris suum (and no, I don’t know how to pronounce that). Ascaris worms have males and females (no hermaphrodites), so there isn’t the issue of hermaphrodite sperm-activating proteins interfering with finding the male ones. By the way, fun fact: While C. elegans are mostly peaceful, soil-dwelling creatures, Ascaris suum are parasites that infect pigs. This isn’t the worm that you can sometimes get from undercooked pork, but it is related. Sorry, I digress…back to the paper.

Ascaris worm sperm crawling around. Image via Florida State University webpage.

The authors started out looking at the membranous organelles (MOs) in the worm’s crawly sperm (I mentioned these in an earlier post, too). These are big blobby things in the sperm that do…something. They’re needed for the sperm’s crawling ability, but it’s not clear what exactly they do. But, there is an antibody that recognizes the MOs so that scientists can study them in the lab. The authors of this paper wanted to know if the antibody, which was made using C. elegans‘ MOs could also recognize MOs from Ascaris. Not only did they find that it could, but they also finally nailed down the exact protein that the antibody is sticking to in the MOs: a protease inhibitor they named As_SRP-1. I’ll just call it SRP.

In pre-activated sperm, SRP is on the surface of the MOs, inside the sperm. When sperm are activated, it moves to the surface of the sperm cell, where it can leave the sperm and go out into the surrounding fluid. Since it was already known that MOs are needed for sperm to crawl, the researchers wanted to know if this depended on SRP. Taking sperm that were happily crawling about, they perfused the sperm with an SRP-specific antibody that would bind to SRP and block its function. The sperm stopped crawling. They next blocked SRP in pre-activated sperm and found that they couldn’t activate. So SRP is needed to activate sperm and keep them moving.

But–hold on a minute. The earlier paper found that a protease (a serine protease, specifically) is needed to activate sperm. Now this paper is saying that a serine protease inhibitor does the same thing? How does that work?

The plot thickens…

Let’s get out of the sperm for a minute and go out into the seminal fluid. Fluid from the vas deferens contains lots of proteins (including proteases), and this fluid is required for sperm activation during ejaculation. The authors referred to this fluid as “sperm activating substance”, or SAS. If you treat pre-activated sperm with SAS, they are activated. Okay, so that’s step 1.

Step 2: SRP moves from the MOs to the outside of the sperm. SRP is required for sperm to activate and crawl–how this connects to step 1 is still unclear.

Step 3: SRP moves into the seminal fluid–the stuff that has all that SAS. And this is the cool part: it turns the SAS into, well, just substance. SRP, the protease inhibitor, blocks the activating ability of the seminal fluid.

The consequence? Any sperm in the vicinity that haven’t already been activated are outta luck. Why? The most obvious possibility was that SRP interacts with (and blocks) some protease, or proteases, in the SAS. So, the researchers broke up the SAS into fractions and looked for the fraction that could activate sperm. This fraction contained a protease that was recognized by SRP. Guess who the protease was? That’s right: Try-5 (or, to be exact, the Ascaris version of it).

The next experiments were pretty cool. The researchers incubated sperm in the SAS fraction that had Try-5. The sperm were activated, as expected. Then, the fluid was re-collected off the activated sperm and added to a second batch of sperm: boom! No more sperm activation.

To show that this effect was because of SRP, they did the experiment again, but added the SRP antibody into the mix when the first batch of sperm were activated–this should bind up all the SRP that gets into the fluid from the activated sperm. After collecting the fluid off the activated sperm again, it was added to a second batch. This time, the sperm were activated! The only difference? No SRP.

This doesn’t prove a direct relationship between Try-5 and SRP, but it makes a pretty convincing case for it. To sum up the paper, I drew you a picture:

But why does SRP block other sperm from activating? The authors speculate that it might play a role in sperm competition. If a female quickly mates with another male or for some reason has unactivated sperm in her uterus, SRP will quickly shut down the activation of the rival sperm. I’m not exactly sure how the timescale works with this. For example, how long does sperm activation actually take? Hopefully, some sperm competition experiments are in the future, using males that don’t make SRP (if someone makes the mutant…).

And now, for the fun behind-the-scenes twist: This paper seems like a follow-up of the C. elegans Try-5 paper, but it actually isn’t. The papers are from different labs and were written at almost the same time, with neither lab knowing about the other’s paper (or so it seems). While the Ascaris paper was still in the review process, the Try-5 paper got published. So each lab showed, in a different way, that Try-5 was really important for sperm activation. I hope that a C. elegans lab (maybe the one from the first paper), will use the SRP results to do some fun genetic experiments with C. elegans and add to the understanding of sperm activation.

Zhao, Y., Sun, W., Zhang, P., Chi, H., Zhang, M., Song, C., Ma, X., Shang, Y., Wang, B., Hu, Y., Hao, Z., Huhmer, A., Meng, F., L’Hernault, S., He, S., Dong, M., & Miao, L. (2012). Nematode sperm maturation triggered by protease involves sperm-secreted serine protease inhibitor (Serpin) Proceedings of the National Academy of Sciences, 109 (5), 1542-1547 DOI: 10.1073/pnas.1109912109

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One thought on “Worm sperm protein turns sperm on, then keeps rival sperm off

  1. Pingback: Sperm are Cool #1: Naked Mole Rats | Molecular Love (and other facts of life)

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