Identification Of Tribolium Dosage Compensation-related Genes And RNAi Analysis

Dosage compensation mechanism is crucial for the balance expression of X chromosome genes, which ensures the protein or enzyme encoded by the X chromosome to be equal or almost equal expression amounts between males and females. However, different organisms have evolved distinct dosage compensation strategies, and so far three kinds of dosage compensation strategies among organisms have been reported. The first strategy is that the single male X chromosome expression is doubly activated; the second one is to inactivate one female X chromosome by leaving both sexes with one active allele; and the third one is to reduce the expression to half activity in both X chromosomes of the female. The study of dosage compensation will be useful to reveal the mechanism of regulation of X-linked genes as well as the evolution and the differentiation progress of the sex chromosome, and it can also contribute to illustrate mutation and distortion of sex chromosome.Tribolium castaneum is not only a seriously harmful grain pest, but also a representative insect for Coleoptera and a new model insect. And T. castaneum is a sole insect with efficient systemic RNA interference (RNAi) as nematodes. So T. castaneum is optimal model organism knock out a specific gene activity during development. So far, the study of dosage compensation mechanism just only one article based on Microrray sequencing method, proposed that the T. castaneum Xmale/Amale is1, Xmale/Xfemale is0.79, Xfemale/Afemale is1.53(X:X chromosome, A:Autosomes), instead of inperfect dosage compensation with Xmale/Xfemale, Xmale/Amale and Xfemale/Afemal all about1, so Tribolium have a inperfect dosage compensation. To further explore the Tribolium dosage compensation effect, we identified the Drosophila dosage compensation key genes homologous genes and explored approach their function using RNAi in Tribolium castaneum.Thus, we have identified and cloned5genes, msl2, msl3a&b (msl3a on Chr6and msl3b on Chr10), mof and mle, from Tribolium castaneum; they were named as Tcmsl2, Tcmsl3a&b, Tcmof and Tcmle. Phylogenetic analysis revealed that MOF is highly conserved in eukaryotes, with three conserved domains, but it was lost in birds. And MOF may be functional differentiation in insects. Real time quantitative PCR results revealed that the Tcmsl2was higher expressed in female than in male from pupa to adult; the Tcmsl3a&b was higher expressed in female than in male in early adult, and Tcmsl3a&b higher expressed in early pupa and late adult; Tcmof was highest expressed in the early embryo stage, Tcmof and Tcmle did not show any differential expressions between males and females in T. castaneum. RNAi of Tcmof caused79.1%insects were arrested at the eclosion procedure and the rest either died in the larval stage or just after eclosion. Parental RNAi suggested that the suppression of Tcmof will completely reduced the fertility of T. castaneum, which could rescued by reciprocal cross with wild type female but not male. In addition to the Tcmle gene knowdown the expression in late larvae by RNAi, the late adult mortality is slightly larger than the control group; RNAi of the others genes Tcmsl2、Tcmsl3a&b and Tcmle, the growth, development and reproduction of the red flour beetle almost no effect.Therefore, these results illustrate that the dosage compensation mechanism in T. castaneum is not similarly to Drosophila melanogaster. Even it has dosage compensation in T. castaneum, but the Tcmsl2, Tcmsl3a&b, Tcmof, and Tcmle4genes may not involved in dosage compensation. And our results illustrated that the mof gene is essential for the growth, development and fertility in T. castaneum, and MOF is probable involved in non-specific lethal comlex with widely regulate the gene expression of sex chromosome and autosomal in female and male. This study helps to reveal the X-linked gene regulation mechanism, evolution and sex chromosome differentiation process, as well as the explanatory chromosomal aberrations mechanism. The study also laid the foundation for future to research dosage compensation of other species and other epigenetic.