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The tissue-specific lncRNA Fendrr is an essential regulator of heart and body wall development in the mouse
The histone modifying complexes PRC2 and TrxG/MLL play pivotal roles in determining the activation state of genes controlling pluripotency, lineage commitment, and cell differentiation. Long non-coding RNAs (lncRNAs) can bind to either complex, and some have been shown to act as modulators of PRC2 or TrxG/MLL activity. Here we show that the lateral mesoderm-specific lncRNA Fendrr is essential for proper heart and body wall development in the mouse. Embryos lacking Fendrr displayed upregulation of several transcription factors controlling lateral plate or cardiac mesoderm differentiation, accompanied by a drastic reduction in PRC2 occupancy along with decreased H3K27 trimethylation and/or an increase in H3K4 trimethylation at their promoters. Fendrr binds to both the PRC2 and TrxG/MLL complexes, suggesting that it acts as modulator of chromatin signatures that define gene activity. Thus, our work identifies a lncRNA that plays an essential role in fine-tuning the regulatory networks which control the fate of lateral mesoderm derivatives
Die Rolle des Transkriptionsfaktors BRACHYURY in der Regulierung der Chromatinmodifikationen in frĂŒhen Mesodermzellen
The gene Brachyury, T, encodes for the founding member of the T-box family of
transcription factors and was previously shown to be crucial for the proper
specification and development of the mesodermal germ layer during mouse
embryogenesis. In addition to their ability to directly activate and repress
transcription by binding to T-box binding sites within target genes, T-box
factors were also shown to modulate gene expression by their ability to modify
the chromatin environment at target genes through a physical interaction with
the MLL/Trithorax family of H3K4 methyltransferases, H3K27 demethylases, and
BRG1, a component of the SWI/SNF chromatin remodeling complex of enzymes. This
interaction is important for the function of most T-box family members, as
multiple mutations in T-box factors which are predicted to disrupt this
interaction are found in human patients with congenital developmental
disorders. I generated a point mutation within the endogenous T locus of mouse
embryonic stem cells, creating an amino acid substitution (Y88A) in the T
protein, to determine whether the interaction of T with histone-modifying
enzymes is necessary for proper T function during mesodermal development.
Mouse embryos expressing the mutant TY88A allele resemble T knockout embryos
and lose the ability to maintain T expression, suggesting that the Y88 residue
is necessary for proper T function and for maintenance of T expression itself.
Gene expression profiling of T-positive mesodermal cells from embryos
expressing the TY88A allele revealed that 60% of dysregulated genes were
unique to the TY88A mutant when compared to T knockout embryos, suggesting
that the amino acid substitution disrupts a critical function of T separate
from the maintenance of T expression. Furthermore, gene expression profiling
corroborated previous data that T may play a role in hemangioblast commitment,
and uncovered a role for T in mammalian erythrocyte specification and
development. Genome-wide histone modification profiling in T-positive cells
differentiated into mesoderm in vitro revealed that functional interaction
with H3K4 methyltransferases and H3K27 demethylases was not disrupted in the
TY88A mutant, however, H3K27 acetylation at T binding sites was decreased.
Overall, the data in this thesis suggest that the Y88 residue in T is
necessary for proper T function during mesodermal specification, and is
necessary in the control of H3K27 acetylation at target loci in early
mesodermal cells.Brachyury, T, wurde als erstes Gen, das fĂŒr einen T-Box Transkriptionsfaktor
kodiert identifiziert und begrĂŒndet daher diese Genfamilie. Brachyury ist
essentiell fĂŒr die Spezifizierung und Entwicklung des mesodermalen Keimblattes
wÀhrend der Embryonalentwicklung. T-Box Faktoren binden sogenannte T-Box
Zielsequenzen in der genomischen DNA und können dadurch in der NÀhe gelegene
Gene aktivieren oder reprimieren. DarĂŒberhinaus konnte gezeigt werden, dass
T-Box Faktoren durch die Bindung von Proteinkomplexen der MLL/Trithorax
Familie der H3K4 Histonmethyltransferasen, H3K27 Histondemethylasen und dem
Chromatinremodellierungs-Komplex BRG1, einem Mitglied der SWI/SNF Familie, die
GenaktivitĂ€t beeinflussen können. Diese Interaktion ist fĂŒr die physiologische
Funktion der T-Box Faktoren sehr wichtig, so sind mehrere Mutationen im
humanen Genom bekannt, die AminosÀureaustausche in T-Box Faktoren zur Folge
haben, die diese Interaktionen verhindern und damit bei betroffenen Patienten
zu schweren Symptomen fĂŒhren. Um festzustellen wie essentiell die Bindung von
chromatinmodifizierenden Proteinkomplexen durch T-Box Faktoren ist, wurde das
Genom von embryonale Stammzellen der Maus so verĂ€ndert, dass es fĂŒr ein T
Protein mit einem AminosÀureaustausch (Y88A) kodiert, welches diese
Interaktion nicht mehr zulassen sollte. Die sich aus diesen verÀnderten
Stammzellen entwickelnden Mausembryonen weisen morphologisch dieselben
PhÀnotypen wie T Null Mutanten auf. Des weiteren wird Brachyury-Expression in
den TY88A Mutanten initiiert, aber nicht aufrecht erhalten, was zeigt, dass
Y88 wichtig fĂŒr die normale Funktion von T ist, und T wichtig fĂŒr die
Aufrechterhaltung der Expression seiner selbst. Der Vergleich von
Genexpressionsprofilen zwischen T positiven, mesodermalen Zellen aus dem
frĂŒhen Embryo von TY88A und T Null Mutanten zeigt, dass 60% der deregulierten
Gene spezifisch nur in TY88A Mutanten dereguliert sind. Den meisten dieser
Gene kann eine Funktion in der Entwicklung des HĂ€mangioblasten, einer
Population multipotenter Stammzellen aus der Endothelzellen und Blutzellen
hervorgehen, zugeordnet werden. Die genomweite Analyse und Vergleich von in
vitro gewonnenen mesodermalen T positiven Zellen der TY88A Mutante zeigt
ausserdem, dass H3K4- und H3K27-Methylierung nicht aberrant ist. Jedoch ist
die H3K27- Acetylierung an Loci an denen T direkt bindet deutlich reduziert.
Zusammengenommen zeigen die hier gewonnen Daten, dass die ĂŒber die AminosĂ€ure
Y88 vermittelte Interaktion von BRACHYURY mit anderen Komplexen essentiell ist
fĂŒr bestimmte Funktionen von BRACHYURY wĂ€hrend der Mesodermentwicklung und
dass diese Interaktion wichtig ist fĂŒr die Acetylierung von Chromatin an von
BRACHYURY gebundenen genomischen Regionen
The Tissue-Specific lncRNA Fendrr Is an Essential Regulator of Heart and Body Wall Development in the Mouse
The histone-modifying complexes PRC2 and TrxG/MLL play pivotal roles in determining the activation state of genes controlling pluripotency, lineage commitment, and cell differentiation. Long noncoding RNAs (lncRNAs) can bind to either complex, and some have been shown to act as modulators of PRC2 or TrxG/MLL activity. Here we show that the lateral mesoderm-specific lncRNA Fendrr is essential for proper heart and body wall development in the mouse. Embryos lacking Fendrr displayed upregulation of several transcription factors controlling lateral plate or cardiac mesoderm differentiation, accompanied by a drastic reduction in PRC2 occupancy along with decreased H3K27 trimethylation and/or an increase in H3K4 trimethylation at their promoters. Fendrr binds to both the PRC2 and TrxG/MLL complexes, suggesting that it acts as modulator of chromatin signatures that define gene activity. Thus, we identified an lncRNA that plays an essential role in the regulatory networks controlling the fate of lateral mesoderm derivatives
The tissue-specific IncRNA Fendrr is an essential regulator of heart and body wall development in the mouse.
The histone-modifying complexes PRC2 and TrxG/MLL play pivotal roles in determining the activation state of genes controlling pluripotency, lineage commitment, and cell differentiation. Long noncoding RNAs (lncRNAs) can bind to either complex, and some have been shown to act as modulators of PRC2 or TrxG/MLL activity. Here we show that the lateral mesoderm-specific lncRNA Fendrr is essential for proper heart and body wall development in the mouse. Embryos lacking Fendrr displayed upregulation of several transcription factors controlling lateral plate or cardiac mesoderm differentiation, accompanied by a drastic reduction in PRC2 occupancy along with decreased H3K27 trimethylation and/or an increase in H3K4 trimethylation at their promoters. Fendrr binds to both the PRC2 and TrxG/MLL complexes, suggesting that it acts as modulator of chromatin signatures that define gene activity. Thus, we identified an lncRNA that plays an essential role in the regulatory networks controlling the fate of lateral mesoderm derivatives