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Female-biased expression of long non-coding RNAs in domains that escape X-inactivation in mouse.

Reinius B, Shi C, Hengshuo L, Sandhu KS, Radomska KJ, Rosen GD, Lu L, Kullander K, Williams RW, Jazin E - BMC Genomics (2010)

Bottom Line: Genes located on sex chromosomes were consistently over-represented in all brain regions.Moreover, RNA-DNA combined FISH experiments confirmed the biallelic expression of one such novel domain.At the same time, sexually dimorphic gene expression that is common to several parts of the brain is mostly restricted to the sex chromosomes.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Evolution and Development, EBC, Uppsala University, Sweden. bjorn.reinius@ebc.uu.se

ABSTRACT

Background: Sexual dimorphism in brain gene expression has been recognized in several animal species. However, the relevant regulatory mechanisms remain poorly understood. To investigate whether sex-biased gene expression in mammalian brain is globally regulated or locally regulated in diverse brain structures, and to study the genomic organisation of brain-expressed sex-biased genes, we performed a large scale gene expression analysis of distinct brain regions in adult male and female mice.

Results: This study revealed spatial specificity in sex-biased transcription in the mouse brain, and identified 173 sex-biased genes in the striatum; 19 in the neocortex; 12 in the hippocampus and 31 in the eye. Genes located on sex chromosomes were consistently over-represented in all brain regions. Analysis on a subset of genes with sex-bias in more than one tissue revealed Y-encoded male-biased transcripts and X-encoded female-biased transcripts known to escape X-inactivation. In addition, we identified novel coding and non-coding X-linked genes with female-biased expression in multiple tissues. Interestingly, the chromosomal positions of all of the female-biased non-coding genes are in close proximity to protein-coding genes that escape X-inactivation. This defines X-chromosome domains each of which contains a coding and a non-coding female-biased gene. Lack of repressive chromatin marks in non-coding transcribed loci supports the possibility that they escape X-inactivation. Moreover, RNA-DNA combined FISH experiments confirmed the biallelic expression of one such novel domain.

Conclusion: This study demonstrated that the amount of genes with sex-biased expression varies between individual brain regions in mouse. The sex-biased genes identified are localized on many chromosomes. At the same time, sexually dimorphic gene expression that is common to several parts of the brain is mostly restricted to the sex chromosomes. Moreover, the study uncovered multiple female-biased non-coding genes that are non-randomly co-localized on the X-chromosome with protein-coding genes that escape X-inactivation. This raises the possibility that expression of long non-coding RNAs may play a role in modulating gene expression in domains that escape X-inactivation in mouse.

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Female-biased expression of pairs of coding and non-coding X-linked genes is validated by quantitative RT-PCR. The RNA expression of co-localized pairs of coding and non-coding genes in three tissues, normalized to Actb and Gapdh is shown. Brain (n = 19 females, 19 males), Eye (n = 16 females, 16 males) and Lung (n= 16 females, 16 males). The X-inactivated gene Rps4x is used as a negative control, while Xist is taken as a female-specific control. The heights of the bars represent mean female expression (yellow bars) as relative to mean male expression (blue bars). Error bars signify +/- standard error of the mean. p-values: * ≤ 0.05, ** ≤ 0.01, *** ≤ 0.001 (two-sided unequal variance t-test), ns: not significant, non-significant p-values are given within brackets. The numbers 1-4 above the figure correspond to the four female-biased clusters in Figure 3.
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Figure 4: Female-biased expression of pairs of coding and non-coding X-linked genes is validated by quantitative RT-PCR. The RNA expression of co-localized pairs of coding and non-coding genes in three tissues, normalized to Actb and Gapdh is shown. Brain (n = 19 females, 19 males), Eye (n = 16 females, 16 males) and Lung (n= 16 females, 16 males). The X-inactivated gene Rps4x is used as a negative control, while Xist is taken as a female-specific control. The heights of the bars represent mean female expression (yellow bars) as relative to mean male expression (blue bars). Error bars signify +/- standard error of the mean. p-values: * ≤ 0.05, ** ≤ 0.01, *** ≤ 0.001 (two-sided unequal variance t-test), ns: not significant, non-significant p-values are given within brackets. The numbers 1-4 above the figure correspond to the four female-biased clusters in Figure 3.

Mentions: Quantitative RT-PCR experiments confirmed the female-biased expression of the above mentioned pairs of X-linked coding and non-coding genes in brain, eye and lung tissues (p ≤ 0.05, two-sided unequal variance t-test) (Figure 4). One exception was 2010308F09Rik, not confirmed as female-biased in lung, but evidently female-biased in brain and eye. Female to male fold-changes were in the same range as detected in the microarray experiments, and in the expected range for X-inactivation escapee genes [32]. An X-linked negative control for sex bias, Rps4x, which is inactivated on the silent mouse X-chromosome [33,34] and therefore not expected to show female-biased expression, remained unchanged. A positive control for female-bias, Xist, was highly female-biased (~10 000-fold) as expected.


Female-biased expression of long non-coding RNAs in domains that escape X-inactivation in mouse.

Reinius B, Shi C, Hengshuo L, Sandhu KS, Radomska KJ, Rosen GD, Lu L, Kullander K, Williams RW, Jazin E - BMC Genomics (2010)

Female-biased expression of pairs of coding and non-coding X-linked genes is validated by quantitative RT-PCR. The RNA expression of co-localized pairs of coding and non-coding genes in three tissues, normalized to Actb and Gapdh is shown. Brain (n = 19 females, 19 males), Eye (n = 16 females, 16 males) and Lung (n= 16 females, 16 males). The X-inactivated gene Rps4x is used as a negative control, while Xist is taken as a female-specific control. The heights of the bars represent mean female expression (yellow bars) as relative to mean male expression (blue bars). Error bars signify +/- standard error of the mean. p-values: * ≤ 0.05, ** ≤ 0.01, *** ≤ 0.001 (two-sided unequal variance t-test), ns: not significant, non-significant p-values are given within brackets. The numbers 1-4 above the figure correspond to the four female-biased clusters in Figure 3.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3091755&req=5

Figure 4: Female-biased expression of pairs of coding and non-coding X-linked genes is validated by quantitative RT-PCR. The RNA expression of co-localized pairs of coding and non-coding genes in three tissues, normalized to Actb and Gapdh is shown. Brain (n = 19 females, 19 males), Eye (n = 16 females, 16 males) and Lung (n= 16 females, 16 males). The X-inactivated gene Rps4x is used as a negative control, while Xist is taken as a female-specific control. The heights of the bars represent mean female expression (yellow bars) as relative to mean male expression (blue bars). Error bars signify +/- standard error of the mean. p-values: * ≤ 0.05, ** ≤ 0.01, *** ≤ 0.001 (two-sided unequal variance t-test), ns: not significant, non-significant p-values are given within brackets. The numbers 1-4 above the figure correspond to the four female-biased clusters in Figure 3.
Mentions: Quantitative RT-PCR experiments confirmed the female-biased expression of the above mentioned pairs of X-linked coding and non-coding genes in brain, eye and lung tissues (p ≤ 0.05, two-sided unequal variance t-test) (Figure 4). One exception was 2010308F09Rik, not confirmed as female-biased in lung, but evidently female-biased in brain and eye. Female to male fold-changes were in the same range as detected in the microarray experiments, and in the expected range for X-inactivation escapee genes [32]. An X-linked negative control for sex bias, Rps4x, which is inactivated on the silent mouse X-chromosome [33,34] and therefore not expected to show female-biased expression, remained unchanged. A positive control for female-bias, Xist, was highly female-biased (~10 000-fold) as expected.

Bottom Line: Genes located on sex chromosomes were consistently over-represented in all brain regions.Moreover, RNA-DNA combined FISH experiments confirmed the biallelic expression of one such novel domain.At the same time, sexually dimorphic gene expression that is common to several parts of the brain is mostly restricted to the sex chromosomes.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Evolution and Development, EBC, Uppsala University, Sweden. bjorn.reinius@ebc.uu.se

ABSTRACT

Background: Sexual dimorphism in brain gene expression has been recognized in several animal species. However, the relevant regulatory mechanisms remain poorly understood. To investigate whether sex-biased gene expression in mammalian brain is globally regulated or locally regulated in diverse brain structures, and to study the genomic organisation of brain-expressed sex-biased genes, we performed a large scale gene expression analysis of distinct brain regions in adult male and female mice.

Results: This study revealed spatial specificity in sex-biased transcription in the mouse brain, and identified 173 sex-biased genes in the striatum; 19 in the neocortex; 12 in the hippocampus and 31 in the eye. Genes located on sex chromosomes were consistently over-represented in all brain regions. Analysis on a subset of genes with sex-bias in more than one tissue revealed Y-encoded male-biased transcripts and X-encoded female-biased transcripts known to escape X-inactivation. In addition, we identified novel coding and non-coding X-linked genes with female-biased expression in multiple tissues. Interestingly, the chromosomal positions of all of the female-biased non-coding genes are in close proximity to protein-coding genes that escape X-inactivation. This defines X-chromosome domains each of which contains a coding and a non-coding female-biased gene. Lack of repressive chromatin marks in non-coding transcribed loci supports the possibility that they escape X-inactivation. Moreover, RNA-DNA combined FISH experiments confirmed the biallelic expression of one such novel domain.

Conclusion: This study demonstrated that the amount of genes with sex-biased expression varies between individual brain regions in mouse. The sex-biased genes identified are localized on many chromosomes. At the same time, sexually dimorphic gene expression that is common to several parts of the brain is mostly restricted to the sex chromosomes. Moreover, the study uncovered multiple female-biased non-coding genes that are non-randomly co-localized on the X-chromosome with protein-coding genes that escape X-inactivation. This raises the possibility that expression of long non-coding RNAs may play a role in modulating gene expression in domains that escape X-inactivation in mouse.

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