The haplolethality paradox of the wupA gene in Drosophila
Haplolethals (HL) are regions of diploid genomes that in one dose are fatal for the organism. Their biological meaning is obscure, given that heterozygous loss-of-function mutations will result in dominant lethality and, consequently, should be under strong negative selection. In addition, their biological nature is under debate: a chromosomal structure that is essential for the expression of several genes (regulatory hypothesis), or the strict dosage requirement of one particular gene (functional hypothesis). We report the first in depth study of an haplolethal region, the one associated to the Drosophila gene wings up A (wupA). It encodes 13 transcripts (A-M) that yield 11 protein isoforms (A-K) of Troponin I (TnI). They are functionally diverse in their control of muscle contraction, cell polarity and cell proliferation. Isoform K can transfer to the nucleus where it increases the transcription of the cell proliferation related genes CDK2, CDK4, Rap and Rab5. The nuclear translocation of isoform K is prevented by the co-expression of A or B isoforms, which illustrates isoform interactions. The corresponding dominant lethal mutations (DL) result from DNA rearrangements, clustered in the intron between exons 7-8, and affect the genomic organization of all transcripts. The joint elimination of isoforms C, F, G and H, however, do not cause DL phenotypes. Genetically driven expression of single isoforms rescue neither DL nor any of the mutants known in the gene, suggesting that normal function of wupA requires the properly regulated expression of specific combinations, rather than single, TnI isoforms along development. We conclude that the HL function at wupA results from the combined haploinsufficiency of a large set of TnI isoforms. The qualitative and quantitative normal expression of which, requires the chromosomal integrity of the wupA genomic region. Since all fly TnI isoforms are encoded in the same gene, its HL condition becomes unavoidable. Author summaryMost species contain two copies of their genetic endowment, each received from their progenitors. If one of the duplicated genes is non-functional, due to either null mutations or genomic loss, the remaining gene copy may supply enough product as to cover the requirements for normal function or, alternatively, may reflect the insufficiency through a visible phenotype. In rare occasions, however, mutation in one copy is so deleterious that causes lethality, usually at early stages of development. These so called "haplolethal regions", exist across species and represent an evolutionary paradox since they should have been subject to intense negative selection. The inherent difficulties to study haplolethals have precluded their study so far. Here, we analyzed the case of one of the five haplolethal regions of Drosophila, the one associated to the Troponin I encoding gene wupA, by measuring the transcriptional effects of mutations and chromosomal rearrangements affecting this gene. The data show that haplolethality results from the combined insufficiency of a large number of Troponin I isoforms, which are functionally specialized, show interference and require the integrity of the native chromatin structure for their quantitatively regulated expression. These features unveil novel aspects of gene expression and, possibly, on evolutionary gene splitting.