Periodic Reporting for period 2 - TOX-ANT (Toxin-antidote selfish elements in animals: from gene drive to speciation)
Okres sprawozdawczy: 2021-09-01 do 2023-02-28
Selfish genetic elements are regions of the genome that promote their own survival while being neutral or even harmful to their host. For instance, they can create multiple copies of themselves (transposons), bias the segregation of alleles during meiosis (meiotic drivers), or—in an extreme scenario—kill individuals that do not inherit them. This genetic conflict induced by selfish elements underlie numerous cases of hybrid dysgenesis, sterility, and genetic incompatibility in the wild. Although selfish elements are a universal feature of genomes, we still know surprisingly little about their molecular diversity, abundance in nature, and their contribution to evolution and disease.
We and others recently discovered a largely uncharacterized class of selfish genes, toxin-antidote elements (TAs), in the nematode C. elegans. TAs are genetic dyads that subvert the laws of Mendelian segregation by killing non-carrier individuals. Since their discovery in nematodes, TAs have now been described in fungi, insects, and plants. However, to date, their molecular underpinnings in eukaryotes remain largely unknown, which severely limits our understanding of their amazing ability to spread in natural populations. It is precisely their “gene drive” activity that has served as the inspiration for the design of synthetic drive elements that control vector-borne diseases, such as malaria and dengue. Homing endonuclease, microRNA and CRISPR/Cas9-based drive elements can spread genes that cause sterility or lethality in mosquitos. However, the rapid evolution of resistant alleles that prevents the spread of these elements significantly precludes utilizing these strategies. To overcome this critical barrier, we must decipher the molecular mechanisms used by natural selfish elements. The results from this ERC Starting Grant will aid in designing more efficient and specific synthetic drive elements.
Specific objectives
1. To dissect the molecular mechanisms underlying an animal toxin-antidote element.
2. To identify and characterize TA elements in diverse nematode species.
3. To screen for toxin-antidote elements in medaka, a vertebrate model organism.
We and others recently discovered a largely uncharacterized class of selfish genes, toxin-antidote elements (TAs), in the nematode C. elegans. TAs are genetic dyads that subvert the laws of Mendelian segregation by killing non-carrier individuals. Since their discovery in nematodes, TAs have now been described in fungi, insects, and plants. However, to date, their molecular underpinnings in eukaryotes remain largely unknown, which severely limits our understanding of their amazing ability to spread in natural populations. It is precisely their “gene drive” activity that has served as the inspiration for the design of synthetic drive elements that control vector-borne diseases, such as malaria and dengue. Homing endonuclease, microRNA and CRISPR/Cas9-based drive elements can spread genes that cause sterility or lethality in mosquitos. However, the rapid evolution of resistant alleles that prevents the spread of these elements significantly precludes utilizing these strategies. To overcome this critical barrier, we must decipher the molecular mechanisms used by natural selfish elements. The results from this ERC Starting Grant will aid in designing more efficient and specific synthetic drive elements.
Specific objectives
1. To dissect the molecular mechanisms underlying an animal toxin-antidote element.
2. To identify and characterize TA elements in diverse nematode species.
3. To screen for toxin-antidote elements in medaka, a vertebrate model organism.
During my postdoctoral work, my colleagues and I discovered the sup-35/pha-1 TA in C. elegans. To our surprise, the antidote, pha-1, was originally described over 25 year ago by Ralf Schnabel who proposed that pha-1 was an essential gene necessary for embryonic development, as pha-1 mutants fail to develop a proper pharynx, the feeding organ of worms. Our experiments challenged this view. Our results indicated instead that this mutant phenotype directly arose from the toxic effect of the linked gene sup-35, not the direct consequence of lack of pha-1 function. Our current understanding how sup-35/pha-1 subverts the laws of Mendelian segregation remains limited. We have identified four main components: sup-35 (toxin), pha-1 (antidote), and two toxic cofactors: sup-36 and sup-37. We hypothesize that the specificity in the activity and expression of sup-36 and sup-37 may determine the pharyngeal phenotypes of pha-1 mutants and that the toxic complex generates “fake” kinetochores that disrupt chromosome segregation. We recently attained a breakthrough in our research when we were able to simultaneously express and purify in both bacterial and insect cells a protein complex consisting of SUP-35, SUP-36, SUP-37, and also additionally a large complex of SUP-35 and PHA-1. Our preliminary results indicate that these complexes are suitable for cryo-EM reconstruction and we expect to solve their structure in the next year.
To better understand the prevalence of TAs in animals and gain insights into their molecular mechanisms, my team and I searched for TAs in two other nematode species, C. briggsae and C. tropicalis. We mapped one TA element in C. briggsae and five distinct TA elements segregating in a single cross between two C. tropicalis isolates, leading to a striking degree of intraspecific incompatibility—over 70% of their F2 hybrid progeny are affected (3). Unlike all previously known elements, four of the five elements in C. tropicalis, as well as the element in C. briggsae, do not cause embryonic lethality, but instead target post-embryonic development.
Our newest discoveries show that rather than being isolated instances, toxin-antidote elements are a ubiquitous class of selfish elements, reveal novel mechanisms of gene drive, and point to a larger role for these elements in genome evolution in animals
To better understand the prevalence of TAs in animals and gain insights into their molecular mechanisms, my team and I searched for TAs in two other nematode species, C. briggsae and C. tropicalis. We mapped one TA element in C. briggsae and five distinct TA elements segregating in a single cross between two C. tropicalis isolates, leading to a striking degree of intraspecific incompatibility—over 70% of their F2 hybrid progeny are affected (3). Unlike all previously known elements, four of the five elements in C. tropicalis, as well as the element in C. briggsae, do not cause embryonic lethality, but instead target post-embryonic development.
Our newest discoveries show that rather than being isolated instances, toxin-antidote elements are a ubiquitous class of selfish elements, reveal novel mechanisms of gene drive, and point to a larger role for these elements in genome evolution in animals
We have now characterized from a genetic standpoint over a dozen toxin-antidote elements across diverse nematode species. Our work has led to a better understanding not only of the evolutionary origin and biology of these elements but also has taught us some unexpected lessons. For instance, TAs have proven extremely useful to study the molecular basis of some key questions, such as how organisms discriminate “self” from “non-self” transcripts in the germline or how giant viral-like mobile elements mediate evolutionary innovation.