Dosage compensation of the X chromosome and inverse effect on the autosomes in RNASeq analysis of triple X metafemales compared to normal females. James A. Birchler¹, Lin Sun¹, Adam Johnson¹, Jilong Li², Jianlin Cheng². 1) Division Biological Sci; 2) Department of Computer Sci, Univ Missouri, Columbia, MO.

   An RNAseq experiment was conducted to examine global gene expression in larval metafemales, normal females and normal males. Triplicate biological replicates were used to determine the number of sequencing reads per gene in each genotype. Then, a ratio distribution analysis was conducted using bins of 0.05. The distribution of the X chromosome for metafemales compared to normal females was largely centered around a ratio of 1.0 representing dosage compensation in metafemales as first noted by Stern (1960). A minor peak was centered near 1.5 representing a subset of genes that exhibited a dosage effect of the X chromosome. For the autosomes, the major peak was centered near 0.67 representing an inverse dosage effect compared to normal females. A minor peak was present near 1.0 representing no change. Another minor peak centered near 0.44, which is the inverse of an inverse effect, which has previously been found in segmental trisomic experiments in flies. Phenotypic validation was conducted by examining the eye color intensity of a mini-white reporter on the X chromosome and the autosomes. One copy of the X linked reporter had the weakest eye color in metafemales; one copy in normal females showed more color and one copy males exhibited the strongest intensity. This continuum illustrates that each gene copy on the X has the lowest expression in metafemales, increases in normal females and is greatest in males conforming to an inverse relationship with the dosage of the X chromosome. The autosomal reporter showed the lowest expression in metafemales, was greater in females and, as is commonly known, the greatest expression was found in males. These results are consistent with an inverse dosage component to dosage compensation.