Unravelling the secrets of a long life

A new study in Evol­u­tion Let­ters has invest­ig­ated the genet­ic basis of long life in the fruit fly Dro­so­phila melano­gaster. We quizzed the lead­er of the research, Pro­fess­or Thomas Flatt, on his group’s find­ings and what they mean for our under­stand­ing of ageing.

EL: Your study set out to identi­fy the mech­an­isms by which longer life evolves. What were your expect­a­tions going into this research?

TF: Since the 1980s a lot has been learned about the genes and the molecu­lar path­ways that affect age­ing and longev­ity, most prom­in­ently genes involved in insulin sig­nalling, but also in oth­er path­ways. We there­fore had the clear – but in hind­sight, naïve – expect­a­tion that we would find pro­nounced evolved genet­ic changes in some of the key path­ways that had been dis­covered in mutant screens and trans­gen­ic ana­lyses. While it was clear that we would not identi­fy the kind of (typ­ic­ally dele­ter­i­ous) large-effect muta­tions that mutant screens have uncovered, we expec­ted to find more subtle genet­ic changes in some of these well recog­nised ‘longev­ity’ genes and path­ways. When I star­ted plan­ning this pro­ject with my col­leagues my vis­ion was that this exper­i­ment would allow us to learn more about the role of nat­ur­al, stand­ing genet­ic vari­ation in insulin sig­nalling and how it affects lifespan. Although we did find some changes in a few well-known genes and path­ways asso­ci­ated with insulin sig­nalling, we were rather sur­prised to find a lot of changes in genes whose link to lifespan was unknown. Per­haps most sur­pris­ingly, we found an enrich­ment of evolved changes in immune genes. While it intu­it­ively makes sense that immune func­tion is involved in age­ing, and while a lot of cur­rent mech­an­ist­ic research is study­ing ‘immuno-sen­es­cence’ and ‘inflamm-aging’, we were non­ethe­less intrigued that our ana­lyses turned up immunity genes as one of the ‘top’ mech­an­isms under­ly­ing evol­u­tion­ary changes in lifespan: these immune genes are not tra­di­tion­ally known to play an import­ant role in lifespan. Since the 2000s there have been a few papers here and there sug­gest­ing that immune genes in flies can affect lifespan, but for the most part the con­nec­tion between lifespan and immunity remains poorly under­stood to this day. Our obser­va­tion, com­bined with read­ing papers about sim­il­ar lifespan selec­tion exper­i­ments by Kim Hughes’ lab at Flor­ida State and Trudy Mackay’s group at North Car­o­lina State, promp­ted us to explore the role of immunity in lifespan.

EL: You used lines of Dro­so­phila selec­ted for late-life fer­til­ity. Can you tell us more about these selec­tion lines, and why they were ideal for test­ing your ideas?

TF: The lines we used were first pub­lished in 1984 in a now fam­ous paper by Leo Luck­in­bill, our coau­thor Bob Ark­ing, and col­leagues. At the same time Michael Rose pub­lished a back-to-back paper in Evol­u­tion on an inde­pend­ent but qual­it­at­ively sim­il­ar selec­tion exper­i­ment. These two papers were ground-break­ing – they showed for the first time that lifespan can be made to evolve in the labor­at­ory. To study the genet­ics under­ly­ing the evol­u­tion of lifespan we had to rely on such ‘labor­at­ory evol­u­tion’ lines rather than long-lived mutants from mutant screens, because large-effect mutants are very unlikely to reflect the sort of genet­ic vari­ants that are segreg­at­ing in nat­ur­al pop­u­la­tions i.e. the sub­strate of evol­u­tion­ary change. The prob­lem is that in many organ­isms it is very hard (and time-con­sum­ing!) to pro­duce long-lived indi­vidu­als via select­ive breed­ing – this is most eas­ily done in rel­at­ively short-lived organ­isms. The fruit fly is optim­al for this pur­pose. The advant­age of the lines we used is that they have been main­tained under selec­tion for over 35 years, thus allow­ing suf­fi­cient time for pro­found evol­u­tion­ary genet­ic changes to be detect­able with gen­ome sequen­cing. In short­er selec­tion exper­i­ments, detect­ing adapt­ive genet­ic changes is often a chal­lenge because the evolved gen­om­ic pat­terns may not be that clear-cut yet.

EL: What tech­niques did you use to unravel the genet­ic mech­an­isms at play?

TF: The tech­nic­al work involved was fun but also a lot of hard work, espe­cially for two first authors on the paper, Daniel Fabi­an and Kath­rin Garschall, two former PhD stu­dents. Although we already had the evolved flies (from Bob Ark­ing), so did not have to per­form a labor­at­ory evol­u­tion exper­i­ment from scratch, we had to use a wide array of meth­ods to nail down the genet­ic mech­an­isms: next-gen­er­a­tion sequen­cing; bioin­form­at­ics, stat­ist­ics, and com­bin­at­or­ics; infec­tion assays with dif­fer­ent patho­gens; meas­ure­ments of gene expres­sion; trans­gen­ic con­structs (RNAi) to knock down the expres­sion of can­did­ate genes; and finally assays to meas­ure the lifespan of flies. Our ambi­tion was not to just per­form a pop­u­la­tion gen­om­ic ana­lys­is of the selec­tion lines, which would have been quite quick, but also to exam­ine and val­id­ate some of the physiolo­gic­al and genet­ic mech­an­isms that we dis­covered via gen­om­ics. To make things even more chal­len­ging, my lab moved three times through­out the course of this work: from Vienna to Lausanne to Fri­bourg – in the end it took us almost 7 years to com­plete this project.

flatt flies.jpg
Pop­u­la­tion cages of fruit flies in the Flatt lab. Photo cred­it: Guil­laume Murat.

EL: A key find­ing of your work was that the evol­u­tion long life is under­pinned by the evol­u­tion of immunity. Why do you think these two things are linked?

TF: Based on our obser­va­tions, I think that the long-lived flies we have stud­ied might have evolved an improved age-depend­ent reg­u­la­tion of their immune sys­tem, a reg­u­la­tion that allows them to fight off infec­tions bet­ter while at the same time redu­cing the costs asso­ci­ated with deploy­ing the immune sys­tem. We were intrigued to see that at old age, short-lived flies up-reg­u­late immune effect­ors but are worse at fight­ing off infec­tions, where­as old long-lived flies tend to down-reg­u­late these effect­ors, yet are bet­ter cap­able of fight­ing infec­tions. This sug­gests to us that long-lived flies have evolved reduced immuno-sen­es­cence and might suf­fer less from immun­o­path­o­logy. Thus, we strongly believe that the evol­u­tion of an improved immune sys­tem con­trib­utes to the evol­u­tion of long life. To what extent this might be a gen­er­al phe­nomen­on remains to be seen.

EL:  How com­par­able do you think the genet­ic mech­an­isms you’ve iden­ti­fied in Dro­so­phila will be to those in oth­er anim­als, par­tic­u­larly those with longer lifespans such as large vertebrates?

TF: A prime example is the insulin sig­nalling path­way men­tioned above: this path­way seems to have evol­u­tion­ar­ily con­served effects on age­ing and lifespan across many organ­isms, from flies and nem­at­ode worms to humans. From work in the nem­at­ode worm C. eleg­ans we know that long-lived worm mutants in the insulin sig­nalling path­way are more res­ist­ant to patho­gen­ic infec­tions. Sim­il­arly, in people who live to over 100 years, cer­tain gene vari­ants in the insulin sig­nalling path­way have been asso­ci­ated with excep­tion­al longev­ity. Yet, we did not really find a lot of changes in this path­way, as men­tioned above – instead we found lots of changes in immune genes. With regard to these immune mech­an­isms, there are at least three argu­ments that make me believe that humans and flies might not be so dif­fer­ent from each oth­er. In humans it is clear from lon­git­ud­in­al stud­ies that the age­ing pro­cess often leads to a chron­ic pro-inflam­mat­ory state and that this can con­trib­ute sig­ni­fic­antly to multi-mor­bid­ity, dis­ab­il­ity, frailty and death, This is some­what sim­il­ar to aged and frail flies, which have prob­lems fight­ing off infec­tions, suf­fer from immun­osen­es­cence, and show signs con­sist­ent with chron­ic inflam­ma­tion. Secondly, there is evid­ence that healthy, long-lived people have improved anti-inflam­mat­ory responses – some­thing sim­il­ar might explain why the long-lived flies we have stud­ied are bet­ter cap­able of deal­ing with infec­tions when they are old. Third, the path­way we have stud­ied in our paper – Toll sig­nalling – plays an evol­u­tion­ar­ily con­served role in immunity across spe­cies: one of our coau­thors (Bruno Lemaitre) was the first to dis­cov­er the role of this path­way in immunity in flies and sub­sequent work then showed that Toll sig­nalling also plays a role in ver­teb­rate immunity. So I do think that some of the insights gained in flies might – at least to some extent – be applic­able to oth­er sys­tems, poten­tially even to humans. But obvi­ously much more work needs to be done before we can draw gen­er­al conclusions.

EL: What would you say are the big unanswered ques­tions in this field? Where do you see your own work head­ing next?

TF: First, we do not know yet wheth­er our obser­va­tions are gen­er­al, so more stud­ies in oth­er organ­isms are needed. Second, the molecu­lar mech­an­isms under­ly­ing our obser­va­tions still need to be worked out in detail – how can down-reg­u­la­tion of immune effect­ors at old age actu­ally improve real­ised immunity and lifespan? How does down-reg­u­la­tion of immune genes affect lifespan mech­an­ist­ic­ally, i.e. what oth­er path­ways and inter­ac­tions are involved in these effects? How does this reg­u­la­tion work in dif­fer­ent tis­sues across the body? Is there a dir­ect con­nec­tion between fer­til­ity and immunity? At the moment, we are ana­lys­ing gen­ome-wide gene expres­sion data in the long-lived fly lines to bet­ter under­stand the physiolo­gic­al changes in these flies. But we are work­ing on a lot of oth­er stuff too. We are study­ing chro­mo­somal inver­sions and ‘supergenes’ and how these affect fit­ness traits, as well as per­form­ing a com­pre­hens­ive ana­lys­is of genet­ic vari­ation in flies on the European con­tin­ent (where flies arrived first 10–15 k years ago when they migrated out of Africa) with an inter­na­tion­al team of col­leagues (the Dros­EU con­sor­ti­um). We are also per­form­ing tran­scrip­tom­ic ana­lyses of the trade-off between repro­duc­tion and somat­ic main­ten­ance. So we are not going to be bored for a long time!

 

Thomas Flatt is Pro­fess­or of Evol­u­tion­ary Bio­logy at the Depart­ment of Bio­logy, Uni­ver­sity of Fri­bourg, Switzer­land. The ori­gin­al study is freely avail­able to read and down­load here.