RNA sequencing shows no dosage compensation of the active X-chromosome

Journal name:
Nature Genetics
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Published online


Mammalian cells from both sexes typically contain one active X chromosome but two sets of autosomes. It has previously been hypothesized that X-linked genes are expressed at twice the level of autosomal genes per active allele to balance the gene dose between the X chromosome and autosomes (termed 'Ohno's hypothesis'). This hypothesis was supported by the observation that microarray-based gene expression levels were indistinguishable between one X chromosome and two autosomes (the X to two autosomes ratio (X:AA) ~1). Here we show that RNA sequencing (RNA-Seq) is more sensitive than microarray and that RNA-Seq data reveal an X:AA ratio of ~0.5 in human and mouse. In Caenorhabditis elegans hermaphrodites, the X:AA ratio reduces progressively from ~1 in larvae to ~0.5 in adults. Proteomic data are consistent with the RNA-Seq results and further suggest the lack of X upregulation at the protein level. Together, our findings reject Ohno's hypothesis, necessitating a major revision of the current model of dosage compensation in the evolution of sex chromosomes.

At a glance


  1. Comparison of gene expressions measured by microarray and RNA-Seq.
    Figure 1: Comparison of gene expressions measured by microarray and RNA-Seq6, 11, 12, 13.

    Human liver is considered unless otherwise noted. (a) Estimation variation measured by the fold difference of microarray intensities of two same-target probesets or of RNA-Seq signals from two halves of the same gene. (b) Identical to a, except that mouse liver is considered here. (c) Comparison of the internal consistency of RNA-Seq data and microarray data. The expression differences from one-half of the nucleotides (RNA-Seq) or a probeset (microarray) are shown for 1,000 randomly picked gene pairs each with twofold ± 0.01-fold expression difference from the other half of nucleotides (RNA-Seq) or from the other probeset (microarray). The central bold line shows the median, the box encompasses 50% of data points and the error bars include 90% of data points. (d) Pearson's correlation (r) of microarray and RNA-Seq expression signals (gray) and of RNA-Seq signals from two independent experiments (black). A certain fraction of genes (x axis) with the highest expression according to one of the RNA-Seq datasets are examined. Error bars show 95% confidence intervals estimated by bootstrapping. (e) Microarray consistently underestimates expression differences between genes. The microarray expression differences of 1,000 randomly picked gene pairs each with x-fold (x = 2 ± 0.01, 4 ± 0.02, 8 ± 0.04, 16 ± 0.08, 32 ± 0.16, and 64 ± 0.32) RNA-Seq expression difference are shown. The central bold line shows the median, the box encompasses 50% of data points and the error bars include 90% of data points. (f) Relative liver expressions of 55 mouse genes, measured by RNA-Seq, microarray and qRT-PCR.

  2. Comparisons of RNA-Seq gene expression levels between the X chromosome and autosomes in 12 human tissues and 3 mouse tissues.
    Figure 2: Comparisons of RNA-Seq gene expression levels between the X chromosome and autosomes in 12 human tissues and 3 mouse tissues11, 12, 13, 16.

    (a) The median expression levels of X-linked genes (closed diamonds) and autosomal genes (open circles) are compared. Median expressions of autosomal genes were normalized to 1. Error bars show 95% bootstrap confidence intervals. Sex information is listed in the parantheses after the tissue names (M, male; F, female; NA, unknown). (b) X:AA ratios of median expressions from the human liver when X is compared to individual autosomes. Error bars show 95% bootstrap confidence intervals.

  3. Comparison of RNA-Seq gene expression levels of the X chromosome and autosomes in C. elegans.
    Figure 3: Comparison of RNA-Seq gene expression levels of the X chromosome and autosomes in C. elegans19.

    (a) X:AA expression ratios at four developmental stages estimated by Miller's jackknife method. Error bars show 95% confidence intervals. (b) Gene expression levels of later developmental stages relative to L2. The overall expressions of autosomal genes at different stages relative to L2 are largely the same, with the medians being 0.98, 0.93 and 0.97 for L3/L2, L4/L2 and adult/L2 C. elegans, respectively. X-linked genes show an overall approximate twofold downregulation, with the median relative expressions being 0.71, 0.55 and 0.43 for L3/L2, L4/L2 and adult/L2 C. elegans, respectively.

Accession codes

Referenced accessions

Gene Expression Omnibus


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Author information

  1. These authors contributed equally to this work.

    • Yuanyan Xiong &
    • Xiaoshu Chen


  1. State Key Laboratory of Biocontrol, College of Life Sciences, Sun Yat-sen University, Guangzhou, China.

    • Yuanyan Xiong,
    • Xiaoshu Chen,
    • Zhidong Chen,
    • Xunzhang Wang,
    • Suhua Shi &
    • Xionglei He
  2. School of Mathematics and Computational Science, Sun Yat-sen University, Guangzhou, China.

    • Xueqin Wang
  3. Zhongshan Medical School, Sun Yat-sen University, Guangzhou, China.

    • Xueqin Wang
  4. Department of Ecology and Evolutionary Biology, University of Michigan, Ann Arbor, Michigan, USA.

    • Jianzhi Zhang
  5. State Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China.

    • Xionglei He


X.H. and J.Z. conceived the study. Y.X., X.C. and Z.C. produced data. X.H., X.C., Y.X., J.Z., Xunzhang Wang, S.S. and Xueqin Wang analyzed data. X.H., Xunzhang Wang and S.S. provided reagents. X.H. and J.Z. wrote the paper.

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The authors declare no competing financial interests.

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