Numerous studies have shown increased testis expression of retrogenes that have moved out of the X chromosome in D. melanogaster[1–3, 7, 8]. Those findings are associated with several evolutionary hypotheses in which autosomal male-biased genes have been favoured by natural selection
[1, 13–19]. However, the recent study of Metta and Schlötterer
 found no evidence of male-biased expression among retrogenes for which the parental copy has been lost. On the contrary, the genes analyzed have mostly female-biased or unbiased expression
. As those genes also show the excessive movement out of the X chromosome, Metta and Schlötterer
 suggested that such a trend is an intrinsic property of retrogenes in Drosophila and not part of an adaptive process.
The segmental dataset used by Metta and Schlötterer
 did not show the same proportion of testis-biased expressed genes observed in the entire dataset of retrogenes in which the parental gene was subsequently lost
. Thus it is clear that the segmental dataset used by Metta and Schlötterer was not representative of the entire dataset of retrogenes for which the parental copy has been lost and the authors therefore took this as evidence against selection-based hypotheses
In addition, statistical analysis of gene movement and sex chromosome evolution can only be performed using tissue-specific expression profiles across species, particularly male gonads
[1–3, 6, 7, 9, 20]. However, such studies are complicated in cases where the parental copy has degenerated or has been lost. In those instances, movements of parent and retrogenes can only be inferred using genomic comparisons and phylogenetic inference between different Drosophila species
[7, 8, 21, 23]. Unfortunately, expression data derived from gonad analysis do not yet exist for all genomic sequenced Drosophila species (only whole-body expression data has been assembled in
Although a previous study of whole-body expression analysis successfully detected the non-random chromosomal distribution of sex-biased genes
, it failed to recover the known extensive male-biased expression obtained using tissue-specific data in D. melanogaster. That means whole-body expression analyses lack the statistical power needed to detect the tissue-specific basis of retrogene movement out of the X chromosome
[7, 8] probably due to the smaller sample size of this dataset in comparison to genome-wide analyses. In a previous study
, we approached this problem by using a conservative analysis of gene movement in D. melanogaster for which gonad expression data are available
[7, 24]. Although the number of retrogenes was too small to conduct a statistical test, it was possible to show that X-linked parental genes for which the corresponding retrogene had moved to the autosomes were generally under-expressed in testis in agreement with sexual antagonism, MSCI and dosage compensation models
. Thus, hypotheses concerning the generality of retrogene movements from the X (with or without parental genes) cannot be tested with existing expression data. We must await the acquisition of appropriate tissue-specific expression data from across the Drosophila clade.
However, we were able to show that there is an association of sex-biased expression with movement out of the X chromosome within the group of retrogenes analyzed by Metta and Schlötterer
. First, using D. melanogaster gonad data from FlyAtlas
, we found the X-linked parental genes tend to be more up regulated in ovaries than retrogenes located in the autosomes. Second, autosomal genes tend to more expressed in meiotic cells of the testis in comparison to X-linked genes. Those results are in agreement with the hypothesis that autosomal regions provide a favourable environment for male-expression
[1, 13–19, 31].
Nevertheless, it is important to notice that even if the tissue-specific data across the Drosophila clade provides evidence for reduced testis-biased expression of retrogenes without parental genes compared to that of retrogenes with parental copies, it will not necessarily rule out MSCI, sexual antagonism, meiotic drive and dosage compensation models
[1, 13–19]. The current sex-biased expression of retrogenes without parental gene does not necessary reflects expression levels when duplication occurred. In this model of retrotransposition, it is reasonable to assume that before the parental gene is lost, the retrogene would either complement the parental gene’s function, or undergo neo- or sub-functionalization
. Only after degeneration of the parental copy could selection favour mutations in the retrogene that gradually restore the parental function
. Therefore, for the selection-driven hypothesis, male-biased expression is only expected by the time the inter-chromosome movements have occurred.
In addition, there are several other lines of evidence supporting hypotheses that predict excessive gene movement off the X chromosome is driven by natural selection. First, the excessive gene movement out of the X chromosome is not exclusively found in retrogenes. Genes created by DNA-based mechanisms also show excessive out-of-the-X movement, which suggest that natural selection, rather than mutation processes intrinsic to retrotransposition, played an essential role in distributing male-biased genes
[7, 8]. Second, chicken and silkworm, which have ZW sex determining systems, also present association between sex-bias gene expression and chromosomal gene movement. In those cases, a symmetrical pattern to the XY sex determining system is observed: genes that move out of the Z chromosome tend to be ovary-biased expressed
[32, 33]. Therefore the phenomenon is not dependent on mutational processes intrinsic to the testis expression and therefore is more likely to be driven by natural selection. Third, a recent population genomic analysis of the copy number variants of Drosophila retrogenes found that there are more fixed than polymorphic retrogenes originating on the X chromosome, which provided direct and strong population genetic evidence for the positive selection hypotheses
. Fourth, it worth mentioning that several autosomal retrogenes that moved out of the Drosophila X chromosome showing clear testis-specific functions have been indentified and extensively described. Examples of those genes are Drosophila nuclear transport factor-2-related (Dntf-2r), Rcd-1 related (Rcd-1r) and gasket (gskt),