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The transcription factor Nfix is essential for normal brain development.

Campbell CE, Piper M, Plachez C, Yeh YT, Baizer JS, Osinski JM, Litwack ED, Richards LJ, Gronostajski RM - BMC Dev. Biol. (2008)

Bottom Line: On standard lab chow Nfix-/- animals show a decreased growth rate from ~P8 to P14, lose weight from ~P14 to P22 and die at ~P22.A fraction of the animals survive to adulthood and are fertile.The weight loss correlates with delayed eye and ear canal opening and suggests a delay in the development of several epithelial structures in Nfix-/- animals.

View Article: PubMed Central - HTML - PubMed

Affiliation: Dept. of Biochemistry and New York State Center of Excellence in Bioinformatics and Life Sciences, State University of New York at Buffalo, 3435 Main St,, Buffalo, NY 14214, USA. cc59@buffalo.edu

ABSTRACT

Background: The Nuclear Factor I (NFI) multi-gene family encodes site-specific transcription factors essential for the development of a number of organ systems. We showed previously that Nfia-deficient mice exhibit agenesis of the corpus callosum and other forebrain defects; Nfib-deficient mice have defects in lung maturation and show callosal agenesis and forebrain defects resembling those seen in Nfia-deficient animals, while Nfic-deficient mice have defects in tooth root formation. Recently the Nfix gene has been disrupted and these studies indicated that there were largely uncharacterized defects in brain and skeletal development in Nfix-deficient mice.

Results: Here we show that disruption of Nfix by Cre-recombinase mediated excision of the 2nd exon results in defects in brain development that differ from those seen in Nfia and Nfib KO mice. In particular, complete callosal agenesis is not seen in Nfix-/- mice but rather there appears to be an overabundance of aberrant Pax6- and doublecortin-positive cells in the lateral ventricles of Nfix-/- mice, increased brain weight, expansion of the cingulate cortex and entire brain along the dorsal ventral axis, and aberrant formation of the hippocampus. On standard lab chow Nfix-/- animals show a decreased growth rate from ~P8 to P14, lose weight from ~P14 to P22 and die at ~P22. If their food is supplemented with a soft dough chow from P10, Nfix-/- animals show a lag in weight gain from P8 to P20 but then increase their growth rate. A fraction of the animals survive to adulthood and are fertile. The weight loss correlates with delayed eye and ear canal opening and suggests a delay in the development of several epithelial structures in Nfix-/- animals.

Conclusion: These data show that Nfix is essential for normal brain development and may be required for neural stem cell homeostasis. The delays seen in eye and ear opening and the brain morphology defects appear independent of the nutritional deprivation, as rescue of perinatal lethality with soft dough does not eliminate these defects.

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Growth defects in Nfix-/- animals. Delayed weight gain and development in Nfix-/- animals. A) Reduced weight gain in Nfix-/- animals on regular chow. The weights of progeny of Nfix-/+ parents were measured and plotted according to genotype. Note that Nfix-/- animals showed a slower weight gain from ~P6 and died at ~P22 when maintained on standard lab chow. Closed circles, WT (+/+); gray circles, Nfix-/+ (HET); open circles, Nfix-/- (KO). B) Increased weight gain of Nfix-/- animals fed a soft dough diet. The cages containing litters of Nfix-/+ parents were supplemented from P10 with soft transgenic dough. Weight gain and survival were increased substantially compared to non-supplemented litters (e.g. 2A). Symbols are as in panel A. C) Delay in eyelid opening in Nfix-/- mice. Eyelid opening was measured at various times after birth in progeny of Nfix-/+ animals. Nfix-/- animals showed an ~2 day delay in eyelid opening. Closed circles, WT (+/+); closed squares, HET (Nfix-/+); open circles, KO (Nfix-/-). The numbers following the genotype indicate the number of animals analyzed. D) Delay in ear canal opening in Nfix-/- mice. The opening of the ear canal was assessed at various times after birth in progeny of Nfix-/+ animals. Nfix-/- (KO) animals showed an ~4 day delay in ear canal opening. Symbols are as in panel C. The data in panels C and D are from an ~50% mixture of dough-supplemented and non-supplemented litters.
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Figure 2: Growth defects in Nfix-/- animals. Delayed weight gain and development in Nfix-/- animals. A) Reduced weight gain in Nfix-/- animals on regular chow. The weights of progeny of Nfix-/+ parents were measured and plotted according to genotype. Note that Nfix-/- animals showed a slower weight gain from ~P6 and died at ~P22 when maintained on standard lab chow. Closed circles, WT (+/+); gray circles, Nfix-/+ (HET); open circles, Nfix-/- (KO). B) Increased weight gain of Nfix-/- animals fed a soft dough diet. The cages containing litters of Nfix-/+ parents were supplemented from P10 with soft transgenic dough. Weight gain and survival were increased substantially compared to non-supplemented litters (e.g. 2A). Symbols are as in panel A. C) Delay in eyelid opening in Nfix-/- mice. Eyelid opening was measured at various times after birth in progeny of Nfix-/+ animals. Nfix-/- animals showed an ~2 day delay in eyelid opening. Closed circles, WT (+/+); closed squares, HET (Nfix-/+); open circles, KO (Nfix-/-). The numbers following the genotype indicate the number of animals analyzed. D) Delay in ear canal opening in Nfix-/- mice. The opening of the ear canal was assessed at various times after birth in progeny of Nfix-/+ animals. Nfix-/- (KO) animals showed an ~4 day delay in ear canal opening. Symbols are as in panel C. The data in panels C and D are from an ~50% mixture of dough-supplemented and non-supplemented litters.

Mentions: Initial breedings indicated that mice homozygous or heterozygous for the targeted Conditional Allele (Fig. 1A–D) had no obvious phenotypes (data not shown). Therefore the knockout allele (KO) was generated by Cre-mediated recombination and mice containing the KO allele were analyzed (Fig. 1E). Nfix transcripts extending from exon 1 to exon 3 were detected in both wild-type (WT) and heterozygous Nfix-/+ mice, but no intact transcripts were detected in Nfix-/- mice (Fig. 1E). Since exons 2 and 3 are in different protein reading frames, no functional proteins should be generated from transcripts lacking exon 2. Mice heterozygous for the knockout allele had no obvious phenotype and upon breeding their progeny were born at a Mendelian ratio (1:2:1,+/+:-/+:-/-, 52:127:51, 32 litters). However, by postnatal day 8 (P8) Nfix-/- (KO) animals began to gain less weight per day than their Nfix -/+ (HET) and +/+ (WT) littermates and by P14 the Nfix-/- animals began to lose weight (Fig. 2A). In addition, Nfix-/- animals opened their eyelids and developed open ear canals ~3 days later than +/+ or Nfix-/+ animals (Fig. 2C &2D), suggesting a general delay in epithelial development. In an effort to overcome the weight loss and lethality seen in the Nfix-/- animals, the diet of the Nfix-/- animals was supplemented with a soft dough feed that we showed previously could rescue lethality in Nfic-/- mice that have defects in tooth development. Surprisingly, while no tooth defects have been detected in the Nfix-/- animals, supplementation with soft dough increased their weight gain and they survived longer than animals fed a standard lab diet (Fig. 2B). The alleviation of some of the weight gain defect with soft dough addition indicates possible defects in the digestive tract, food sensing, or eating behavior in Nfix-/- animals.


The transcription factor Nfix is essential for normal brain development.

Campbell CE, Piper M, Plachez C, Yeh YT, Baizer JS, Osinski JM, Litwack ED, Richards LJ, Gronostajski RM - BMC Dev. Biol. (2008)

Growth defects in Nfix-/- animals. Delayed weight gain and development in Nfix-/- animals. A) Reduced weight gain in Nfix-/- animals on regular chow. The weights of progeny of Nfix-/+ parents were measured and plotted according to genotype. Note that Nfix-/- animals showed a slower weight gain from ~P6 and died at ~P22 when maintained on standard lab chow. Closed circles, WT (+/+); gray circles, Nfix-/+ (HET); open circles, Nfix-/- (KO). B) Increased weight gain of Nfix-/- animals fed a soft dough diet. The cages containing litters of Nfix-/+ parents were supplemented from P10 with soft transgenic dough. Weight gain and survival were increased substantially compared to non-supplemented litters (e.g. 2A). Symbols are as in panel A. C) Delay in eyelid opening in Nfix-/- mice. Eyelid opening was measured at various times after birth in progeny of Nfix-/+ animals. Nfix-/- animals showed an ~2 day delay in eyelid opening. Closed circles, WT (+/+); closed squares, HET (Nfix-/+); open circles, KO (Nfix-/-). The numbers following the genotype indicate the number of animals analyzed. D) Delay in ear canal opening in Nfix-/- mice. The opening of the ear canal was assessed at various times after birth in progeny of Nfix-/+ animals. Nfix-/- (KO) animals showed an ~4 day delay in ear canal opening. Symbols are as in panel C. The data in panels C and D are from an ~50% mixture of dough-supplemented and non-supplemented litters.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2414869&req=5

Figure 2: Growth defects in Nfix-/- animals. Delayed weight gain and development in Nfix-/- animals. A) Reduced weight gain in Nfix-/- animals on regular chow. The weights of progeny of Nfix-/+ parents were measured and plotted according to genotype. Note that Nfix-/- animals showed a slower weight gain from ~P6 and died at ~P22 when maintained on standard lab chow. Closed circles, WT (+/+); gray circles, Nfix-/+ (HET); open circles, Nfix-/- (KO). B) Increased weight gain of Nfix-/- animals fed a soft dough diet. The cages containing litters of Nfix-/+ parents were supplemented from P10 with soft transgenic dough. Weight gain and survival were increased substantially compared to non-supplemented litters (e.g. 2A). Symbols are as in panel A. C) Delay in eyelid opening in Nfix-/- mice. Eyelid opening was measured at various times after birth in progeny of Nfix-/+ animals. Nfix-/- animals showed an ~2 day delay in eyelid opening. Closed circles, WT (+/+); closed squares, HET (Nfix-/+); open circles, KO (Nfix-/-). The numbers following the genotype indicate the number of animals analyzed. D) Delay in ear canal opening in Nfix-/- mice. The opening of the ear canal was assessed at various times after birth in progeny of Nfix-/+ animals. Nfix-/- (KO) animals showed an ~4 day delay in ear canal opening. Symbols are as in panel C. The data in panels C and D are from an ~50% mixture of dough-supplemented and non-supplemented litters.
Mentions: Initial breedings indicated that mice homozygous or heterozygous for the targeted Conditional Allele (Fig. 1A–D) had no obvious phenotypes (data not shown). Therefore the knockout allele (KO) was generated by Cre-mediated recombination and mice containing the KO allele were analyzed (Fig. 1E). Nfix transcripts extending from exon 1 to exon 3 were detected in both wild-type (WT) and heterozygous Nfix-/+ mice, but no intact transcripts were detected in Nfix-/- mice (Fig. 1E). Since exons 2 and 3 are in different protein reading frames, no functional proteins should be generated from transcripts lacking exon 2. Mice heterozygous for the knockout allele had no obvious phenotype and upon breeding their progeny were born at a Mendelian ratio (1:2:1,+/+:-/+:-/-, 52:127:51, 32 litters). However, by postnatal day 8 (P8) Nfix-/- (KO) animals began to gain less weight per day than their Nfix -/+ (HET) and +/+ (WT) littermates and by P14 the Nfix-/- animals began to lose weight (Fig. 2A). In addition, Nfix-/- animals opened their eyelids and developed open ear canals ~3 days later than +/+ or Nfix-/+ animals (Fig. 2C &2D), suggesting a general delay in epithelial development. In an effort to overcome the weight loss and lethality seen in the Nfix-/- animals, the diet of the Nfix-/- animals was supplemented with a soft dough feed that we showed previously could rescue lethality in Nfic-/- mice that have defects in tooth development. Surprisingly, while no tooth defects have been detected in the Nfix-/- animals, supplementation with soft dough increased their weight gain and they survived longer than animals fed a standard lab diet (Fig. 2B). The alleviation of some of the weight gain defect with soft dough addition indicates possible defects in the digestive tract, food sensing, or eating behavior in Nfix-/- animals.

Bottom Line: On standard lab chow Nfix-/- animals show a decreased growth rate from ~P8 to P14, lose weight from ~P14 to P22 and die at ~P22.A fraction of the animals survive to adulthood and are fertile.The weight loss correlates with delayed eye and ear canal opening and suggests a delay in the development of several epithelial structures in Nfix-/- animals.

View Article: PubMed Central - HTML - PubMed

Affiliation: Dept. of Biochemistry and New York State Center of Excellence in Bioinformatics and Life Sciences, State University of New York at Buffalo, 3435 Main St,, Buffalo, NY 14214, USA. cc59@buffalo.edu

ABSTRACT

Background: The Nuclear Factor I (NFI) multi-gene family encodes site-specific transcription factors essential for the development of a number of organ systems. We showed previously that Nfia-deficient mice exhibit agenesis of the corpus callosum and other forebrain defects; Nfib-deficient mice have defects in lung maturation and show callosal agenesis and forebrain defects resembling those seen in Nfia-deficient animals, while Nfic-deficient mice have defects in tooth root formation. Recently the Nfix gene has been disrupted and these studies indicated that there were largely uncharacterized defects in brain and skeletal development in Nfix-deficient mice.

Results: Here we show that disruption of Nfix by Cre-recombinase mediated excision of the 2nd exon results in defects in brain development that differ from those seen in Nfia and Nfib KO mice. In particular, complete callosal agenesis is not seen in Nfix-/- mice but rather there appears to be an overabundance of aberrant Pax6- and doublecortin-positive cells in the lateral ventricles of Nfix-/- mice, increased brain weight, expansion of the cingulate cortex and entire brain along the dorsal ventral axis, and aberrant formation of the hippocampus. On standard lab chow Nfix-/- animals show a decreased growth rate from ~P8 to P14, lose weight from ~P14 to P22 and die at ~P22. If their food is supplemented with a soft dough chow from P10, Nfix-/- animals show a lag in weight gain from P8 to P20 but then increase their growth rate. A fraction of the animals survive to adulthood and are fertile. The weight loss correlates with delayed eye and ear canal opening and suggests a delay in the development of several epithelial structures in Nfix-/- animals.

Conclusion: These data show that Nfix is essential for normal brain development and may be required for neural stem cell homeostasis. The delays seen in eye and ear opening and the brain morphology defects appear independent of the nutritional deprivation, as rescue of perinatal lethality with soft dough does not eliminate these defects.

Show MeSH
Related in: MedlinePlus