The nuclear factor CTCF has been shown to be necessary for the maintenance of genetic imprinting at the mammalian H19/Igf2 locus. MacDonald and colleagues now report in BMC Biology that the mechanisms responsible for maintaining the imprinted state in Drosophila may be evolutionarily conserved and that CTCF may also play a critical role in this process

AC Bell

AT Hark

Chunhui Hou

CJ Schoenherr

HV Crouse

J McGrath

JE Phillips

JL Kermicle

MA Surani

S Kurukuti

Victor G Corces

VK Lloyd

WA MacDonald

BMC Biology

English

PubMed

Springer - Publisher Connector

Insulators and imprinting from flies to mammals

Genetic imprinting is an epigenetic phenomenon that 
results in the expression of certain genes in a parent-of-
origin chromosome-specific manner. Although imprint-
ing was originally discovered in insects [1] and has also 
been described in plants [2] and zebrafish, the pheno-
menon has been most widely studied in mammals [3,4]. 
Imprinting, resulting in the functional non-equivalence 
of the maternal and paternal genomes, affects the 
expression of developmentally important genes in mice 
and humans, and alterations of the process result in 
cancer and various genetic diseases. This is not the case 
in insects; gynogenic and androgenic flies (containing 
only maternal or paternal genomes, respectively) are 
perfectly viable. The molecular mechanisms controlling 
the establishment of imprinting are not fully understood, 
but those involved in the maintenance of the imprinted 
state have been analyzed in detail for some mammalian 
genes, such as the mouse locus that includes the insulin-
like growth factor gene (Igf2) and the RNA gene H19. 
However, understanding of the mechanisms controlling 
imprinting in non-mammalian species has lagged behind 
and it is unclear whether imprinting in insects and 
mammals is a conserved biological process with the same 
underlying molecular mechanisms. In this issue of BMC 
Biology, Lloyd, Meller and colleagues [5] examine the 
potential role of the Drosophila CCCTC binding factor 
(dCTCF) protein in the maintenance of maternal 
imprinting and propose that dCTCF has an evolutionarily 
conserved role in the maintenance of the imprinted state.
All but one case of imprinting described in Drosophila 
is associated with position-effect variegation, in which 
chromosomal rearrangements place genes with visible 
phenotypes close to heterochromatin. MacDonald et al. 
[5] use a Drosophila mini-chromosome in which most of 
the X chromosome is deleted and the garnet gene is placed 
next to the centromeric heterochromatin; the garnet 
protein resembles clathrin and nonclathrin adaptin proteins 
and is similar to the delta subunit of the mammalian 
AP-3 adaptin complex. The rearrangement causes a 
variegated expression of garnet, such that the eyes of the 
adult flies show sectors of expressing and non-expressing 
cells characteristic of heterochromatin-induced silencing. 
Interestingly, this variegated expression is imprinted and 
it is observed in individuals carrying the paternal mini-X-
chromosome, whereas the maternally transmitted copy 
shows normal expression of the garnet gene [6]. This 
observation suggests that the maternally inherited mini-
X-chromosome carries an imprint established in the 
germline that interferes with the somatic spreading of 
heterochromatin silencing in the next generation.
To study whether dCTCF is involved in the differential 
regulation of garnet gene expression in the maternal 
versus paternal chromosomes, the authors [5] examined 
the effect of mutations in the dCTCF gene. Flies carrying 
one mutant copy of dCTCF showed reduced RNA levels 
(30 to 40% of that found in wild type). However, this mild 
reduction is sufficient to significantly compromise 
expression of the garnet gene from the maternal mini-X-
chromosome, leading to a variegated eye color similar to 
that seen from the paternally inherited chromosome. 
This observation suggests that dCTCF is required for the 
non-variegated expression of the garnet gene when it is 
maternally inherited.
Two different processes affect the visually observed 
garnet phenotype: the transcription of the gene under the 
control of regulatory sequences and the spreading of 
heterochromatin silencing - these two components may 
not be easy to separate mechanistically. In the mammalian 
Abstract
The nuclear factor CTCF has been shown to be 
necessary for the maintenance of genetic imprinting 
at the mammalian H19/Igf2 locus. MacDonald 
and colleagues now report in BMC Biology that the 
mechanisms responsible for maintaining the imprinted 
state in Drosophila may be evolutionarily conserved 
and that CTCF may also play a critical role in this 
process.
Insulators and imprinting from flies to mammals
Chunhui Hou and Victor G Corces*
See research article http://www.biomedcentral.com/1741-7007/8/105
CO M M E N TA RY  Open Access
*Correspondence: vcorces@emory.edu 
Department of Biology, Emory University, 1510 Clifton Road NE, Atlanta, GA 30322, 
USA
© 2010 Hou and Corces; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative 
Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and 
reproduction in any medium, provided the original work is properly cited.
Hou and Corces BMC Biology 2010, 8:104 
http://www.biomedcentral.com/1741-7007/8/104
H19/Igf2 locus, direct transcriptional activation can be 
distinguished from indirect effects from the surrounding 
chromatin thanks to information obtained from the 
analysis of the function of CTCF in the spatial organi-
zation of the maternal locus. This organization functions 
to maintain the imprinted expression of H19 and the 
silencing of Igf2 on the maternal chromosome. If CTCF 
binding in the imprinting control region (ICR) is 
abolished, both Igf2 and H19 can be transcribed from the 
maternal allele because of the disruption in the CTCF-
mediated maternal allele chromatin organization; this 
organization prevents the interaction between enhancers 
and the promoter of Igf2. These observations suggest that 
the role of CTCF in the maintenance of imprinting 
involves its ability to mediate interactions that result in a 
specific three-dimensional architecture of the locus. In 
fact, CTCF is not directly involved in the transcription 
activation of the mouse H19 and Igf2 genes [7-10].
Figure 1. Possible models to explain the role of dCTCF in the maintenance of imprinting in Drosophila. In all panels, a hypothetical 
transcription factor (TF) controlling the expression of the garnet gene is shown as a red oval; the garnet gene is represented as a green arrow, 
which is dashed when the gene is subject to silencing by heterochromatin and solid when it is expressed normally; dCTCF is represented by a blue 
oval; nucleosomes are shown as yellow circles; and DNA is in black. (a) dCTCF in the maternal chromosome forms a barrier against the spreading 
of heterochromatin, leading to normal expression of the adjacent gene; presumably, CTCF is not present in the paternal chromosome and 
heterochromatin spreads into the gene. (b) dCTCF, either directly or in combination with other factors, affects the transcription of the garnet gene, 
antagonizing the spreading of heterochromatin and overcoming its silencing effect. (c) An alternative explanation that involves the formation of a 
loop between a dCTCF site adjacent to the heterochromatin and a second site somewhere else in the genome. The garnet gene and its regulatory 
sequences are located inside of the loop, which protects the gene against heterochromatin silencing. The models in (a,c) are conceptually similar 
but mechanistically different and the latter is more in line with observations in mammals.
Paternal chromosome Maternal chromosome(a)
(b)
(c)
Subject to silencing Expressed normally
TF TF
TF TF
TF TF
C
T
C
F
C
T
C
F
C
TC
F
C
TC
F
C
T
C
F
Hou and Corces BMC Biology 2010, 8:104 
http://www.biomedcentral.com/1741-7007/8/104
Page 2 of 3
In Drosophila, the primary effect of the maternal 
imprint is to inhibit the spread of heterochromatin-
induced silencing (that is, silencing that turns euchro-
matin into a more compact state that limits the access of 
transcription factors to the genes). The finding that the 
garnet gene is poorly expressed from the paternally 
transmitted mini-X-chromosome and is not affected by 
reduced dCTCF expression [5] suggests that hetero-
chromatini zation is an effective gene silencing mecha-
nism. Expression of the garnet gene in the maternally 
derived mini-X-chromosome could then be accomplished 
by the establishment of a distinct barrier to the spread of 
heterochromatin (Figure 1a) or by a direct effect of 
dCTCF on garnet transcription that indirectly antago-
nizes heterochromatin spreading (Figure 1b). In the latter 
case, an epigenetically transmitted increased expression 
of garnet in the maternal germline would be the imprint 
that inhibits heterochromatin spreading in the somatic 
cells of the progeny.
After showing a role for dCTCF in the maintenance of 
the maternal imprint, MacDonald et al. [5] explored the 
possibility of a similar function for this protein in the 
establishment of the imprinted state in the maternal 
germline. This process is poorly understood, not only in 
insects but also in mammals. Although the exact nature 
of the proteins involved and how they function in the 
establishment of genetic imprinting during gametogenisis 
are unknown, some candidate proteins have been 
excluded from a direct role in this process. For example, 
it is now clear that CTCF is not necessary for the 
establishment of imprinting in the mouse H19/Igf2 locus. 
Given the functional conservation of CTCF as an insu-
lator protein between flies and mammals, it is interesting 
to ask whether this is also the case in Drosophila. 
MacDonald et al. [5] found that expression of the garnet 
gene is not subject to heterochromatin-induced silencing 
in a mini-X-chromosome inherited from females hetero-
zy gous for mutations in the dCTCF gene. This obser-
vation suggests that dCTCF is not required for the 
establishment of the maternal imprint. However, the 
question remains as to whether further reduction in the 
levels of dCTCF in the maternal germline may actually 
show an effect on this process. For example, it is possible 
that dCTCF expres sion during oogenesis is much higher 
than in somatic cells, and that the small reduction in 
dCTCF levels in heterozygous mutant females is not 
sufficient to affect the establishment of the imprint. 
Alterna tively, there may be other mechanisms that can 
prevent the spreading of heterochromatin during 
gametogenesis independent of dCTCF. Such a mecha-
nism could use other insulator proteins or alternative 
processes to prevent hetero chromatin spreading.
CTCF has multiple roles in chromatin organization and 
gene regulation that derive from its ability to mediate 
intra- and inter-chromosome interactions [11]. Given 
that the ability of CTCF to organize chromatin resulting 
from its insulator function is the basis for its role in 
transcriptional regulation and genetic imprinting, many 
of these functions are probably evolutionarily conserved 
from flies to humans. In vertebrates, CTCF has been 
shown to act as an enhancer-blocking insulator and to 
function by creating intra- and inter-chromosomal loops, 
but CTCF does not seem to form barrier insulators, 
which seem to function by recruiting chromatin remodel-
ing proteins that act by covalently modifying histones. 
Although such a distinction has not yet been made in 
Drosophila, it is possible that the role of dCTCF in 
maintaining the imprinted state is related to the ability of 
this protein to mediate interactions that create a chroma-
tin domain insulated from heterochromatin silenc ing 
(Figure  1c). The findings of MacDonald et al. [5] agree 
with this idea. Additional analysis of the molecular 
mecha nism of imprinting in Drosophila will shed new 
light not only on the understanding of the mechanisms 
controlling this process, but also on the understanding of 
the evolutionary conservation of CTCF function.
Acknowledgements
We thank Ashley M Bushey for her critical reading and helpful comments on 
the manuscript. Work in the authors’ laboratory is supported by Public Health 
Service Award GM35463 from the National Institutes of Health.
Published: 30 July 2010
References
1. Crouse HV: The controlling element in sex chromosome behavior in Sciara. 
Genetics 1960, 45:1429-1443.
2. Kermicle JL: Dependence of the R-mottle aleurone phenotype in maize on 
mode of sexual transmission. Genetics 1970, 66:69-85.
3. McGrath J, Solter D: Completion of mouse embryogenesis requires both 
the maternal and paternal genomes. Cell 1984, 37:179-183.
4. Surani MA, Barton SC, Norris ML: Development of reconstituted mouse 
eggs suggests imprinting of the genome during gametogenesis. Nature 
1984, 308:548-550.
5. MacDonald WA, Menon D, Bartlett NJ, Sperry GE, Rasheva V, Meller V, Lloyd 
VK: The Drosophila homolog of the mammalian imprint regulator, CTCF, 
maintains the maternal genomic imprint in Drosophila melanogaster. BMC 
Biol 2010, 8:105
6.  Lloyd VK, Sinclair DA, Grigliatti TA: Genomic imprinting and position-effect 
variegation in Drosophila melanogaster. Genetics 1999, 151:1503-1516.
7. Kurukuti S, Tiwari VK, Tavoosidana G, Pugacheva E, Murrell A, Zhao Z, 
Lobanenkov V, Reik W, Ohlsson R: CTCF binding at the H19 imprinting 
control region mediated maternally inherited higher-order chromatin 
conformation to restrict enhancer access to Igf2. Proc Natl Acad Sci USA 
2006, 103:10684-10689.
8. Bell AC, Felsenfeld G: Methylation of a CTCF-dependent boundary controls 
imprinted expression of the Igf2 gene. Nature 2000, 405:482-485.
9. Hark AT, Schoenherr CJ, Katz DJ, Ingram RS, Levorse JM, Tilghman SM: CTCF 
mediated methylation-sensitive enhancer-blocking activity at the 
H19/Igf2 locus. Nature 2000, 405:486-489.
10. Schoenherr CJ, Levorse JM, Tilghman SM: CTCF maintains differential 
methylation at the Igf2/H19 locus. Nat Genet 2003, 33:66-69.
11. Phillips JE, Corces VG: CTCF: Master weaver of the genome. Cell 2009, 
137:1194-1211.
doi:10.1186/1741-7007-8-104
Cite this article as: Hou C, Corces VG: Insulators and imprinting from flies to 
mammals. BMC Biology 2010, 8:104.
Hou and Corces BMC Biology 2010, 8:104 
http://www.biomedcentral.com/1741-7007/8/104
Page 3 of 3


Crossref

Corces VG: CTCF: Master weaver of the genome. Cell

CTCF binding at the H19 imprinting control region mediated maternally inherited higher-order chromatin conformation to restrict enhancer access to Igf2. Proc Natl Acad Sci USA

Dependence of the R-mottle aleurone phenotype in maize on mode of sexual transmission. Genetics

Felsenfeld G: Methylation of a CTCF-dependent boundary controls imprinted expression of the Igf2 gene. Nature

Genomic imprinting and position-effect variegation in Drosophila melanogaster. Genetics

ML: Development of reconstituted mouse eggs suggests imprinting of the genome during gametogenesis. Nature

Solter D: Completion of mouse embryogenesis requires both the maternal and paternal genomes. Cell

The controlling element in sex chromosome behavior in Sciara. Genetics

Tilghman SM: CTCF maintains differential methylation at the Igf2/H19 locus. Nat Genet

Tilghman SM: CTCF mediated methylation-sensitive enhancer-blocking activity at the H19/Igf2 locus. Nature

VK: The Drosophila homolog of the mammalian imprint regulator, CTCF, maintains the maternal genomic imprint in Drosophila melanogaster.

https://bmcbiol.biomedcentral.com/track/pdf/10.1186/1741-7007-8-104?site=bmcbiol.biomedcentral.com

Insulators and imprinting from flies to mammals

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