title: Determination of dosage compensation of the mammalian X chromosome by RNA-seq is dependent on analytical approach.
abstract: BACKGROUND
  An enduring question surrounding sex chromosome evolution is whether effective hemizygosity in the heterogametic sex leads inevitably to dosage compensation of sex-linked genes, and whether this compensation has been observed in a variety of organisms. Incongruence in the conclusions reached in some recent reports has been attributed to different high-throughput approaches to transcriptome analysis. However, recent reports each utilizing RNA-seq to gauge X-linked gene expression relative to autosomal gene expression also arrived at diametrically opposed conclusions regarding X chromosome dosage compensation in mammals.
  
  
  RESULTS
  Here we analyze RNA-seq data from X-monosomic female human and mouse tissues, which are uncomplicated by genes that escape X-inactivation, as well as published RNA-seq data to describe relative X expression (RXE). We find that the determination of RXE is highly dependent upon a variety of computational, statistical and biological assumptions underlying RNA-seq analysis. Parameters implemented in short-read mapping programs, choice of reference genome annotation, expression data distribution, tissue source for RNA and RNA-seq library construction method have profound effects on comparing expression levels across chromosomes.
  
  
  CONCLUSIONS
  Our analysis shows that the high number of paralogous gene families on the mammalian X chromosome relative to autosomes contributes to the ambiguity in RXE calculations, RNA-seq analysis that takes into account that single- and multi-copy genes are compensated differently supports the conclusion that, in many somatic tissues, the mammalian X is up-regulated compared to the autosomes.
authors.affiliation: Department of Molecular and Cell Biology, University of Connecticut, 354 Mansfield Road, Storrs, CT 06235, USA.
authors.forename: Nathaniel K
authors.initials: NK
authors.lastname: Jue
authors.forename: Michael B
authors.initials: MB
authors.lastname: Murphy
authors.forename: Seth D
authors.initials: SD
authors.lastname: Kasowitz
authors.forename: Sohaib M
authors.initials: SM
authors.lastname: Qureshi
authors.forename: Craig J
authors.initials: CJ
authors.lastname: Obergfell
authors.forename: Sahar
authors.initials: S
authors.lastname: Elsisi
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Foley
authors.forename: Rachel J
authors.initials: RJ
authors.lastname: O'Neill
authors.forename: Michael J
authors.initials: MJ
authors.lastname: O'Neill
mesh_terms: D000818:Animals; D004303:Dosage Compensation, Genetic; D005260:Female; D020869:Gene Expression Profiling; D015723:Gene Library; D006801:Humans; D051379:Mice; D058977:Molecular Sequence Annotation; D017423:Sequence Analysis, RNA / Q000379:methods*; D013223:Statistics as Topic; D014960:X Chromosome / Q000235:genetics*