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DNA methyltransferase 3b is dispensable for mouse neural crest development.

Jacques-Fricke BT, Roffers-Agarwal J, Gammill LS - PLoS ONE (2012)

Bottom Line: In both neural crest-specific and fully DNMT3b-mutant embryos, cranial neural crest cells exhibited only subtle migration defects, with increased numbers of dispersed cells trailing organized streams in the head.In spite of this, the resulting cranial ganglia, craniofacial skeleton, and heart developed normally when neural crest cells lacked DNMT3b.We conclude that defects in neural crest derivatives in DNMT3b mutant mice reflect a requirement for DNMT3b in lineages such as the branchial arch mesendoderm or the cardiac mesoderm that interact with neural crest cells during formation of these structures.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, Minnesota, United States of America.

ABSTRACT
The neural crest is a population of multipotent cells that migrates extensively throughout vertebrate embryos to form diverse structures. Mice mutant for the de novo DNA methyltransferase DNMT3b exhibit defects in two neural crest derivatives, the craniofacial skeleton and cardiac ventricular septum, suggesting that DNMT3b activity is necessary for neural crest development. Nevertheless, the requirement for DNMT3b specifically in neural crest cells, as opposed to interacting cell types, has not been determined. Using a conditional DNMT3b allele crossed to the neural crest cre drivers Wnt1-cre and Sox10-cre, neural crest DNMT3b mutants were generated. In both neural crest-specific and fully DNMT3b-mutant embryos, cranial neural crest cells exhibited only subtle migration defects, with increased numbers of dispersed cells trailing organized streams in the head. In spite of this, the resulting cranial ganglia, craniofacial skeleton, and heart developed normally when neural crest cells lacked DNMT3b. This indicates that DNTM3b is not necessary in cranial neural crest cells for their development. We conclude that defects in neural crest derivatives in DNMT3b mutant mice reflect a requirement for DNMT3b in lineages such as the branchial arch mesendoderm or the cardiac mesoderm that interact with neural crest cells during formation of these structures.

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DNMT3b-deleted neural crest cells have mild migration defects that recover during cranial gangliogenesis.Migratory neural crest cells and the cranial ganglia they form were visualized by in situ hybridization for Sox10 at E8.5 (A, B), E9.0 (C–H), E9.5 (I–K) and E10 (L–M). (A–H) E8.5–9.0 embryos that were DNMT3b+/fl (A, C; wildtype; n = 6); Wnt1-cre; DNMT3bfl/− (B, D; homozygous deleted in neural crest cells; n = 13), DNMT3b+/− (E, G; whole embryo heterozygous; n = 6), and DNMT3b−/− (F; H; whole embryo homozygous deleted; n = 5). Despite normal migration initially (A, B), dispersed Sox10-positive cells were apparent dorsally, trailing the organized neural crest streams in the branchial arches (ba; arrows in D, F) and eye (e; arrowheads in D, F) in mutant embryos compared to controls (C, E). Subsequently, neural crest cells coalesce to form the trigeminal ganglia (G, H, brackets). (I–N) At E9.5–10, no difference beyond normal embryonic variation was apparent during cranial gangliogenesis between DNMT3b+/fl (n = 24), Sox10-cre; DNMT3bfl/− (n = 6), and Wnt1-cre; DNMT3bfl/− embryos (n = 13). fn, frontonasal region; g, geniculate; n, nodose; o, otic vesicle; p, petrosal; s, somites; t, trigeminal.
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pone-0047794-g004: DNMT3b-deleted neural crest cells have mild migration defects that recover during cranial gangliogenesis.Migratory neural crest cells and the cranial ganglia they form were visualized by in situ hybridization for Sox10 at E8.5 (A, B), E9.0 (C–H), E9.5 (I–K) and E10 (L–M). (A–H) E8.5–9.0 embryos that were DNMT3b+/fl (A, C; wildtype; n = 6); Wnt1-cre; DNMT3bfl/− (B, D; homozygous deleted in neural crest cells; n = 13), DNMT3b+/− (E, G; whole embryo heterozygous; n = 6), and DNMT3b−/− (F; H; whole embryo homozygous deleted; n = 5). Despite normal migration initially (A, B), dispersed Sox10-positive cells were apparent dorsally, trailing the organized neural crest streams in the branchial arches (ba; arrows in D, F) and eye (e; arrowheads in D, F) in mutant embryos compared to controls (C, E). Subsequently, neural crest cells coalesce to form the trigeminal ganglia (G, H, brackets). (I–N) At E9.5–10, no difference beyond normal embryonic variation was apparent during cranial gangliogenesis between DNMT3b+/fl (n = 24), Sox10-cre; DNMT3bfl/− (n = 6), and Wnt1-cre; DNMT3bfl/− embryos (n = 13). fn, frontonasal region; g, geniculate; n, nodose; o, otic vesicle; p, petrosal; s, somites; t, trigeminal.

Mentions: In order to compare the onset and extent of cre recombinase expression in cranial neural crest cells in Wnt1-cre and Sox10-cre lines side by side, we crossed them to the R26R cre-dependent lacZ reporter line [23]. Wnt1-cre was active as early as 3 somites (3 s), when low levels of cre expression were detectable in the anterior neural plate (Fig. 3A, arrowhead). This expression increased at 5 s (Fig. 3B, arrowhead), and by 11 s, Wnt1-cre expression was apparent in the forebrain, midbrain, and migratory neural crest cells in the first branchial arch stream (Fig. 3C, arrowhead). Premigratory neural crest cells in the hindbrain and trunk also expressed Wnt1-cre at this stage (Fig. 3C, arc; inset). The pattern of ß-galactosidase activity in Wnt1-cre embryos at 11s included the Wnt1 expression domain (Fig. 4D) plus migratory neural crest cells, in which Wnt1-cre had been expressed when these cells were in the neural folds (arrow).


DNA methyltransferase 3b is dispensable for mouse neural crest development.

Jacques-Fricke BT, Roffers-Agarwal J, Gammill LS - PLoS ONE (2012)

DNMT3b-deleted neural crest cells have mild migration defects that recover during cranial gangliogenesis.Migratory neural crest cells and the cranial ganglia they form were visualized by in situ hybridization for Sox10 at E8.5 (A, B), E9.0 (C–H), E9.5 (I–K) and E10 (L–M). (A–H) E8.5–9.0 embryos that were DNMT3b+/fl (A, C; wildtype; n = 6); Wnt1-cre; DNMT3bfl/− (B, D; homozygous deleted in neural crest cells; n = 13), DNMT3b+/− (E, G; whole embryo heterozygous; n = 6), and DNMT3b−/− (F; H; whole embryo homozygous deleted; n = 5). Despite normal migration initially (A, B), dispersed Sox10-positive cells were apparent dorsally, trailing the organized neural crest streams in the branchial arches (ba; arrows in D, F) and eye (e; arrowheads in D, F) in mutant embryos compared to controls (C, E). Subsequently, neural crest cells coalesce to form the trigeminal ganglia (G, H, brackets). (I–N) At E9.5–10, no difference beyond normal embryonic variation was apparent during cranial gangliogenesis between DNMT3b+/fl (n = 24), Sox10-cre; DNMT3bfl/− (n = 6), and Wnt1-cre; DNMT3bfl/− embryos (n = 13). fn, frontonasal region; g, geniculate; n, nodose; o, otic vesicle; p, petrosal; s, somites; t, trigeminal.
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pone-0047794-g004: DNMT3b-deleted neural crest cells have mild migration defects that recover during cranial gangliogenesis.Migratory neural crest cells and the cranial ganglia they form were visualized by in situ hybridization for Sox10 at E8.5 (A, B), E9.0 (C–H), E9.5 (I–K) and E10 (L–M). (A–H) E8.5–9.0 embryos that were DNMT3b+/fl (A, C; wildtype; n = 6); Wnt1-cre; DNMT3bfl/− (B, D; homozygous deleted in neural crest cells; n = 13), DNMT3b+/− (E, G; whole embryo heterozygous; n = 6), and DNMT3b−/− (F; H; whole embryo homozygous deleted; n = 5). Despite normal migration initially (A, B), dispersed Sox10-positive cells were apparent dorsally, trailing the organized neural crest streams in the branchial arches (ba; arrows in D, F) and eye (e; arrowheads in D, F) in mutant embryos compared to controls (C, E). Subsequently, neural crest cells coalesce to form the trigeminal ganglia (G, H, brackets). (I–N) At E9.5–10, no difference beyond normal embryonic variation was apparent during cranial gangliogenesis between DNMT3b+/fl (n = 24), Sox10-cre; DNMT3bfl/− (n = 6), and Wnt1-cre; DNMT3bfl/− embryos (n = 13). fn, frontonasal region; g, geniculate; n, nodose; o, otic vesicle; p, petrosal; s, somites; t, trigeminal.
Mentions: In order to compare the onset and extent of cre recombinase expression in cranial neural crest cells in Wnt1-cre and Sox10-cre lines side by side, we crossed them to the R26R cre-dependent lacZ reporter line [23]. Wnt1-cre was active as early as 3 somites (3 s), when low levels of cre expression were detectable in the anterior neural plate (Fig. 3A, arrowhead). This expression increased at 5 s (Fig. 3B, arrowhead), and by 11 s, Wnt1-cre expression was apparent in the forebrain, midbrain, and migratory neural crest cells in the first branchial arch stream (Fig. 3C, arrowhead). Premigratory neural crest cells in the hindbrain and trunk also expressed Wnt1-cre at this stage (Fig. 3C, arc; inset). The pattern of ß-galactosidase activity in Wnt1-cre embryos at 11s included the Wnt1 expression domain (Fig. 4D) plus migratory neural crest cells, in which Wnt1-cre had been expressed when these cells were in the neural folds (arrow).

Bottom Line: In both neural crest-specific and fully DNMT3b-mutant embryos, cranial neural crest cells exhibited only subtle migration defects, with increased numbers of dispersed cells trailing organized streams in the head.In spite of this, the resulting cranial ganglia, craniofacial skeleton, and heart developed normally when neural crest cells lacked DNMT3b.We conclude that defects in neural crest derivatives in DNMT3b mutant mice reflect a requirement for DNMT3b in lineages such as the branchial arch mesendoderm or the cardiac mesoderm that interact with neural crest cells during formation of these structures.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, Minnesota, United States of America.

ABSTRACT
The neural crest is a population of multipotent cells that migrates extensively throughout vertebrate embryos to form diverse structures. Mice mutant for the de novo DNA methyltransferase DNMT3b exhibit defects in two neural crest derivatives, the craniofacial skeleton and cardiac ventricular septum, suggesting that DNMT3b activity is necessary for neural crest development. Nevertheless, the requirement for DNMT3b specifically in neural crest cells, as opposed to interacting cell types, has not been determined. Using a conditional DNMT3b allele crossed to the neural crest cre drivers Wnt1-cre and Sox10-cre, neural crest DNMT3b mutants were generated. In both neural crest-specific and fully DNMT3b-mutant embryos, cranial neural crest cells exhibited only subtle migration defects, with increased numbers of dispersed cells trailing organized streams in the head. In spite of this, the resulting cranial ganglia, craniofacial skeleton, and heart developed normally when neural crest cells lacked DNMT3b. This indicates that DNTM3b is not necessary in cranial neural crest cells for their development. We conclude that defects in neural crest derivatives in DNMT3b mutant mice reflect a requirement for DNMT3b in lineages such as the branchial arch mesendoderm or the cardiac mesoderm that interact with neural crest cells during formation of these structures.

Show MeSH
Related in: MedlinePlus