Two novel DXZ4-associated long noncoding RNAs show developmental changes in expression coincident with heterochromatin formation at the human (Homo sapiens) macrosatellite repeat
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On the male X and female active X chromosome (Xa), the macrosatellite repeat (MSR) DXZ4 is packaged into constitutive heterochromatin characterized by CpG methylation and histone H3 tri-methylated at lysine-9 (H3K9me3). In contrast, DXZ4 on the female inactive X chromosome (Xi), is packaged into euchromatin, is bound by the architectural protein CCCTC-binding factor, and mediates Xi-specific long-range cis contact with similarly packaged tandem repeats on the Xi. In cancer, male DXZ4 can inappropriately revert to a Xi-like state and other MSRs have been reported to adopt alternate chromatin configurations in response to disease. Given this plasticity, we sought to identify factors that might control heterochromatin at DXZ4. In human embryonic stem cells, we found low levels of 5-hydroxymethylcytosine at DXZ4 and that this mark is lost upon differentiation as H3K9me3 is acquired. We identified two previously undescribed DXZ4 associated noncoding transcripts (DANT1 and DANT2) that are transcribed toward DXZ4 from promoters flanking the array. Each generates transcript isoforms that traverse the MSR. However, upon differentiation, enhancer of Zeste-2 silences DANT1, and DANT2 transcription terminates prior to entering DXZ4. These data support a model wherein DANT1 and/or DANT2 may function to regulate constitutive heterochromatin formation at this MSR.
KeywordsMacrosatellite DXZ4 X chromosome inactivation Human embryonic stem cells Euchromatin and heterochromatin Long noncoding RNA
American Type Culture Collection
DXZ4-associated noncoding transcript, proximal
DXZ4-associated noncoding transcript, distal
Embryoid body out-growths
Enhancer of Zeste 2
Expressed sequence tags
Facioscapulohumeral muscular dystrophy
Histone H3 di-methylated at lysine 4
Histone H3 tri-methyated at lysine 9
Human embryonic stem cells
Human Gene Nomenclature Committee
Long interspersed elements
Long noncoding RNA
Long terminal repeats
Open reading frame
Oxidative bisulfite sequencing
Polycomb repressive complex 2
Quantitative chromatin immunoprecipitation
Rapid amplification of cDNA ends
Reverse transcription PCR
Transcription start sites
Active X chromosome
X chromosome inactivation
Inactive X chromosome
X inactive specific transcript
This work was supported by the National Institutes of Health [GM073120 to B.P.C.] and by a subaward from P01GM085354 to Dr. Stephen Dalton, University of Georgia.
Experiments performed in this manuscript comply with the current laws of the USA.
Conflict of interest
Debbie M. Figueroa declares that she has no conflict of interest.
Emily M. Darrow declares that she has no conflict of interest.
Brian P. Chadwick declares that he has no conflict of interest.
This article does not contain any studies with human or animal subjects performed by any of the authors.
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