Think of neurons like cities, which need to be able to transport goods and services throughout themselves. In cities this is done on highways by cargo trucks. In neurons it’s done on microtubules by motor proteins. To direct traffic, cities have signals like lights and road signs, while in neurons these signals are thought to come in the form of post-translational modifications.
These modifications on the microtubules are hypothesized to ensure that cargo in these neurons gets to the right place at the right time. The lab of assistant professor Jill Wildonger at the University of Wisconsin–Madison Department of Biochemistry studies these modifications and their impact on the transport of cargo. New research from the Wildonger Lab led by postdoctoral scholar Brian Jenkins investigated a particular post-translational modification, using a new approach to find that researchers’ assumptions about the modification might not have been as correct as once thought. Their research was recently on the cover of the Journal of Cell Science.
The particular post-translational modification they studied occurs on the microtubule at lysine 40 (K40). The modification is called lysine acetylation, which means a chemical combo called an acetyl group is added to the end of the amino acid lysine on tubulin (microtubules are made from proteins called tubulin). This modification occurs on the inside of microtubules, which are little tubes, rather than the outside where transport occurs. This acetylation was thought to be essential for proper transport in a neuron and hence survival of an organism.
“Our main finding, which was kind of surprising, is that when you get rid of the K40 acetylation, the flies we study were perfectly happy,” says Jenkins, who is now a postdoc at the Jungers Center for Neurosciences Research at Oregon Health and Science University. “It was assumed if you really messed up transport that the flies would be very unhealthy or die but that wasn’t what we found. Researchers in this field might need to reevaluate how important this particular modification is.”
While not serious, the researchers did find some other interesting effects due to the lack of microtubule acetylation. In one example, the dendrites of the neurons were over branched. Dendrites are the signal-receiving piece of the neuron that branches out to receive stimuli, and over branching can cause issues. It wasn’t previously thought that microtubule acetylation affected the dendrites so the finding was novel and intriguing, Jenkins says.
Something else they found was decreased lysosome transport. Lysosomes are an organelle commonly known as the “trash can of the cell,” responsible for keeping the cell healthy and recycling and degrading proteins and other molecules. They saw some of these potentially negative effects but the flies were still healthy, showing how resilient the cells can be and how this modification might not be as essential as researchers thought.
“Overall it was interesting how we found that this acetylation did affect transport, although not seriously, because this acetylation takes place on the inside of the microtubule and the transport occurs on the outside,” Jenkins says. “A big unanswered question in this field is the purpose of this modification and this research helps us get at an answer.”
Their experiments took a unique genetic approach that allowed them to test the effect of microtubule acetylation in a more unbiased manner than other methods. Using a form of genome engineering, the researchers made a specific change to part of the microtubule, rather than using a drug to manipulate the acetylation or inhibiting an enzyme involved in the acetylation process. It’s possible those alternative methods have unknown effects on other proteins in the cell so while they are good approaches, a genetic method directly targeting microtubules may give better results, the researchers say.
The Wildonger Lab uses fruit fly larvae in their experiments and images the sensory neurons in the peripheral nervous system, analogous to those nerves in human skin that sense heat, cold, and other stimuli. Wildonger explains that the use of fruit flies is a great model for studying neuronal microtubules.
“Mice are furry animals so it’s difficult to visualize transport and microtubule dynamics within neurons in live mice,” she says. “It’s much easier in the fruit fly model to visualize individual neurons, so that coupled with the genetic editing tools we have in fruit flies makes for a very powerful system to experiment on.”
The implications of their work are spread across basic research and neurodegenerative disease. Jenkins says the work of understanding the basic mechanisms and functions of microtubules and post-translational modifications is extremely useful and this research gets at those questions.
He and Wildonger also add that in some neurodegenerative diseases like Parkinson’s or Huntington’s, other researchers have shown that a manipulation to increase microtubule acetylation helps fix some issues with cargo transport experienced by the neurons.
“While a change in these modifications might not cause the disease or directly fix it, it’s possible that changes in microtubule acetylation may make a disease worse or, hopefully, hold the potential to relieve some symptoms,” says Wildonger. “That’s driven a lot of the interest in this particular modification. We aren’t directly addressing that question but we are working to put together parts of that underlying puzzle.”
She and Jenkins say being on the cover of the journal’s December issue was an honor and nice recognition of their unique experimental approach and findings. While the post-translational modification they studied serves as a traffic signal for motor proteins on the microtubule highways, it’s possible this particular sign isn’t completely essential for the neuronal city to keep humming along.
“This work is the first time anyone has used a gene editing approach to directly mutate the microtubules to see the effects in a live animal,” Wildonger says. “It was exciting to be recognized on the cover with a beautiful image highlighting Brian’s work. And, we got to shed some light on this particular modification and how those in this field might need to change the way we think about it.”