Molecular cloning and functional characterization of hybrid male inviability loci between nematodes Caenorhabditis briggsae and C. nigoni

Postzygotic hybrid incompatibility (HI) is a phenomenon where hybrids experience reduced viability or fertility due to genetic incompatibilities between closely related species. One common form of HI is hybrid male inviability (HMI), where hybrid males die before reaching sexual maturity. This prevents gene flow and maintains the genetic identity of each species. Despite its prevalence, the genes responsible for HMI remain largely unknown. Identification of these genes or elements is critical for understanding the evolutionary processes driving speciation.

Cloning HMI loci is challenging, as it requires a species pair that can mate and produce inviable hybrid males between wild type parents, but produce viable hybrid males at least conditionally to enable genetic mapping. Additionally, conventional genetic mapping techniques may not be directly applicable due to little conservation of the loci involved. Therefore, specialized methods and resources need to be developed for mapping HMI loci.

A species pair that meets these criteria is the nematode species Caenorhabditis nigoni, which is dioecious (having separate sexes), and its close relative, C. briggsae, which is hermaphroditic (having both male and female reproductive organs). Both species develop into males with a single X chromosome and females/hermaphrodites with two X chromosomes. F1 hybrid males from C. nigoni fathers are inviable, while those from C. briggsae fathers are sterile. However, the absence of viable hybrid males has posed a significant challenge in mapping and cloning the genes or elements responsible for HMI between these two species. Fortunately, in our previous study, we found that the HMI could be rescued when C. nigoni fathers carried an introgression fragment either from the right arm of chromosome II or from the left arm of chromosome IV of C. briggsae, which opens up a promising opportunity for us to map and clone the specific genes or elements responsible for HMI.

Given that the genotypes of hybrid males from reciprocal crosses differ only in the origin of X chromosome, we hypothesize that a negative epistatic interaction exisits between the C. briggsae X chromosome and a C. nigoni autosomal gene or element, leading to HMI. We propose to test the hypothesis by establishing the molecular identity of the C. nigoni autosomal genes or elements responsible for HMI using the HMI rescuing strains through targeted recombination with CRISPR/Cas9, as we previously demonstrated. The findings from the proposed studies are expected to provide mechanistic insights into HMI and the evolutionary processes driving speciation.