Basic HTML Version
Poster Full Abstracts - Chromatin and Epigenetics
Poster board number is above title. The first author is the presenter
225
of telomeres as an important chromosomal landmark - whether telomeres organize in the nucleus based on clustering. The four final cell cycles before the
onset of cellularization in the fly development take place at the surface of the embryo. This time-period presents a perfect experimental setting for
simultaneous imaging of many synchronized nuclei through consecutive divisions. To follow telomeres in vivo in this system we live-imaged fluorescently-
labeled telomere capping proteins. We gathered evidence that telomeres do assemble into 4-6 clusters per nucleus during these early divisions. In light of the
long-established findings from S. cerevisiae that yeast interphase telomeres cluster at the nuclear periphery, our results suggest that clustering may be a
feature conserved through evolution. We propose that this behavior is mediated through one of the following mechanisms: 1) pairing of the homologous
chromosomes, 2) self-affinity of homologous telomeric sequences, 3) pairing of telomeres of p and q arms of each chromosome, 4) interactions of telomeric
proteins (with each other or the nuclear envelope). Through fluorescence in situ hybridization (FISH) to a natural telomere sequence as well as to a
specifically-marked telomere, we have so far determined that homology does not play a major role in clustering, eliminating possibilities 1 and 2. Through
further detailed studies we hope to distinguish between the remaining possibilities and elucidate how clustering occurs in the fly.
345C
Role of Drosophila's HKMTs in the recruitment of HP1.
Margarida Figueiredo, Anna-Mia Johansson, Jan Larsson. Department of Molecular Biology,
Umeå University, Umeå, Umeå, Sweden.
The fourth chromosome of Drosophila melanogaster is largely heterochromatic being enriched in satellite repeats and in DNA from transposable elements.
Despite its heterochromatic properties it has a gene density similar to the major chromosome arms. The expression of almost all chromosome 4 genes is fine-
tuned by at least two different proteins: Painting of Fourth (POF) that stimulates transcription and Heterochromatin Protein 1 (HP1) that represses
transcription. It has been shown that POF and HP1 bind interdependently to this chromosome and that the binding and spreading of POF depends on
heterochromatin. POF and HP1 colocalize in exons of active genes and HP1 binds additionally to promoters of the active genes. HP1 is recruited by
H3K9me2 and H3K9mme3, which is performed by the HKMTs SU(VAR)3-9, SETDB1 and G9a. It has been proposed that SU(VAR)3-9 is responsible for
H3K9me in the chromocentre whereas SETDB1 acts mainly on the 4th chromosome. To understand the roles of these HKMTs on the recruitment of HP1 we
have performed ChIP-on-chip and immunostainings experiments in Su(var)3-9, Setdb1, G9a and Pof mutants and analysed the binding of POF, HP1,
H3K9me2 and H3K9me3. We found that POF and SETDB1 are essential for recruiting HP1 to the gene bodies on the 4th chromosome whereas SU(VAR)3-
9 is essential for recruiting HP1 to the centromeric regions of all chromosomes. The region-specific recruitment of HP1 by SETDB1 and SU(VAR)3-9 is
explained by the finding that both these HKMTs are essential for region-specific production of both H3K9me2 and H3K9me3. G9a doesn't seem to be
essential for H3K9me2, H3K9me3 and neither for HP1 recruitment anywhere in the genome. HP1 found in promoters of genes on the 4th and genes on other
chromosomes seems to be independent on all the HKMTs and on both H3K9me2 and H3K9me3. Our results also show that in the absence of SETDB1 or
SU(VAR)3-9 there is a relocalization of HP1 and of both H3K9me2 and H3K9me3 in other places.
346A
Co-ordinate regulation of heterochromatic genes in
Drosophila melanogaster
males.
S.Kiran Koya, Xinxian Deng, Ying Kong, Victoria Meller. Dept of
Biological Sciences, Wayne State University, Detroit, MI.
Drosophila dosage compensation equalizes the expression of X-linked genes between males and females. Dosage compensation is mediated by the MSL
(Male-Specific Lethal) complex. The MSL complex is composed of five proteins (MSL1, MSL2, MSL3, MOF and MLE) and two non-coding RNAs,
roX1
and
roX2
. The
roX
RNAs are essential for normal targeting of the MSL complex along the length of the single male X chromosome. Chromatin modification
by the MSL complex results in a two-fold elevation of X-linked gene expression. Microarray analysis of
roX1 roX2
males revealed decreased expression of
X-linked genes, confirming the role of these RNAs in dosage compensation. Surprisingly, expression from the autosomal heterochromatic genes, including
the entire 4
th
chromosome, is also reduced in
roX1 roX2
males. Expression of these genes is unchanged in
roX1 roX2
females. We used microarray and
quantitative real time PCR to measure gene expression in male larvae mutated for each of the
msl
genes. Our findings revealed that MSL1, MSL3 and MLE,
but not MSL2, are required for heterochromatic gene regulation. This suggests that the MSL proteins participate in two distinct complexes, one that regulates
the male X chromosome, and one that is necessary at heterochromatic genes. We postulate that a complex formed of
roX
and a subset of the MSL proteins
may be necessary for correct establishment of heterochromatin in male embryos. We are using a genetic assay to establish the critical period for
roX
during
development, and are using chromatin immuno-precipitation (ChIP) to measured enrichment of MSL1 and MSL3 proteins at heterochromatic genes during
embryogenesis. Our results suggest a fundamental sex-specific difference in heterochromatin in
D. melanogaster
. My future work will focus on the
molecular mechanism by which this regulation occurs.
347B
Modifiers of X recognition: exploring the secrets of sex chromosome identity.
Debashish Menon, Victoria Meller. Department of Biological Science,
Wayne State University, Detroit, MI.
Dosage compensation modulates expression of an entire chromosome to address the potentially fatal imbalance in X-linked gene dose between males and
females. In male Drosophila, the X chromosome is up-regulated through the activity of the Male Specific Lethal (MSL) complex, consisting of five proteins
and two large, non-coding RNA on the X (
roX
) transcripts (
roX1
and
roX2
). Simultaneous mutation of
roX1
and
roX2
reduces X-localization of the MSL
proteins, lowers X chromosome expression and reduces male survival. The Y chromosome is a potent modifier of the
roX1 roX2
phenotype. Examination of
this effect led us to postulate a role for small RNA in dosage compensation. A directed screen of small RNA pathways revealed one that affects X
chromosome recognition and interacts genetically with
roX1 roX2
mutations. Our findings suggest that small RNA cooperates with the MSL complex to
establish X-chromosome binding. Together these processes may underlie X chromosome identity.
348C
The H3K36 Demethylase KDM4A is a Novel Regulator of Heterochromatin Organization and Dynamics.
Serafin U. Colmenares
1,2
, Sasha Langley
1,2
,
Cameron Kennedy
1,2
, Joel Swenson
1,2
, Irene Chiolo
1,2
, Gary Karpen
1,2
. 1) Genome Dynamics, Life Sciences Division, Lawrence Berkeley National Lab,
Berkeley, CA; 2) Molecular and Cell Biology, University of California, Berkeley, CA.
Heterochromatin comprises 30% of the
Drosophila melanogaster
genome and is characterized by highly-condensed, repetitive DNA enriched for