A human YAC transgene rescues craniofacial and neural tube development in PDGFRalpha knockout mice and uncovers a role for PDGFRalpha in prenatal lung growth

Development ◽  
2000 ◽  
Vol 127 (21) ◽  
pp. 4519-4529
Author(s):  
T. Sun ◽  
D. Jayatilake ◽  
G.B. Afink ◽  
P. Ataliotis ◽  
M. Nister ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Neural Crest ◽  
Knockout Mice ◽  
Yeast Artificial Chromosome ◽  
Neural Crest Cells ◽  
Artificial Chromosome ◽  
Regulatory Elements ◽  
Vital Role ◽  
Lung Growth ◽  
Oligodendrocyte Progenitors

The platelet-derived growth factor alpha-receptor (PDGFRalpha) plays a vital role in the development of vertebrate embryos, since mice lacking PDGFRalpha die in mid-gestation. PDGFRalpha is expressed in several types of migratory progenitor cells in the embryo including cranial neural crest cells, lung smooth muscle progenitors and oligodendrocyte progenitors. To study PDGFRalpha gene regulation and function during development, we generated transgenic mice by pronuclear injection of a 380 kb yeast artificial chromosome (YAC) containing the human PDGFRalpha gene. The YAC transgene was expressed in neural crest cells, rescued the profound craniofacial abnormalities and spina bifida observed in PDGFRalpha knockout mice and prolonged survival until birth. The ultimate cause of death was respiratory failure due to a defect in lung growth, stemming from failure of the transgene to be expressed correctly in lung smooth muscle progenitors. However, the YAC transgene was expressed faithfully in oligodendrocyte progenitors, which was not previously observed with plasmid-based transgenes containing only upstream PDGFRalpha control sequences. Our data illustrate the complexity of PDGFRalpha genetic control, provide clues to the location of critical regulatory elements and reveal a requirement for PDGF signalling in prenatal lung growth, which is distinct from the known requirement in postnatal alveogenesis. In addition, we found that the YAC transgene did not prolong survival of Patch mutant mice, indicating that genetic defects outside the PDGFRalpha locus contribute to the early embryonic lethality of Patch mice.

scholarly journals In VitroSystem for Differentiating Pluripotent Neural Crest Cells into Smooth Muscle Cells

1998 ◽  
Vol 273 (11) ◽  
pp. 5993-5996 ◽  
Author(s):  
Mukesh K. Jain ◽  
Matthew D. Layne ◽  
Masafumi Watanabe ◽  
Michael T. Chin ◽  
Mark W. Feinberg ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Neural Crest ◽  
Neural Crest Cells ◽  
Muscle Cells

scholarly journals Functional Analysis of the DXPas34Locus, a 3′ Regulator of Xist Expression

1999 ◽  
Vol 19 (12) ◽  
pp. 8513-8525 ◽  
Author(s):  
E. Debrand ◽  
C. Chureau ◽  
D. Arnaud ◽  
P. Avner ◽  
E. Heard
Keyword(s):  
X Chromosome ◽  
Yeast Artificial Chromosome ◽  
Es Cells ◽  
Artificial Chromosome ◽  
Antisense Transcription ◽  
Regulatory Elements ◽  
X Inactivation ◽  
X Chromosomes ◽  
Inactivation Center ◽  
Controlling Element

ABSTRACT X inactivation in female mammals is controlled by a key locus on the X chromosome, the X-inactivation center (Xic). The Xic controls the initiation and propagation of inactivation in cis. It also ensures that the correct number of X chromosomes undergo inactivation (counting) and determines which X chromosome becomes inactivated (choice). The Xist gene maps to the Xic region and is essential for the initiation of X inactivation in cis. Regulatory elements of X inactivation have been proposed to lie 3′ toXist. One such element, lying 15 kb downstream ofXist, is the DXPas34 locus, which was first identified as a result of its hypermethylation on the active X chromosome and the correlation of its methylation level with allelism at the X-controlling element (Xce), a locus known to affect choice. In this study, we have tested the potential function of theDXPas34 locus in Xist regulation and X-inactivation initiation by deleting it in the context of largeXist-containing yeast artificial chromosome transgenes. Deletion of DXPas34 eliminates both Xistexpression and antisense transcription present in this region in undifferentiated ES cells. It also leads to nonrandom inactivation of the deleted transgene upon differentiation. DXPas34 thus appears to be a critical regulator of Xist activity and X inactivation. The expression pattern of DXPas34 during early embryonic development, which we report here, further suggests that it could be implicated in the regulation of imprintedXist expression.

scholarly journals Regulation of the myeloid-cell-expressed human gp91-phox gene as studied by transfer of yeast artificial chromosome clones into embryonic stem cells: suppression of a variegated cellular pattern of expression requires a full complement of distant cis elements.

1997 ◽  
Vol 17 (4) ◽  
pp. 2279-2290 ◽  
Author(s):  
L L Lien ◽  
Y Lee ◽  
S H Orkin
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Yeast Artificial Chromosome ◽  
Embryonic Stem ◽  
Myeloid Cells ◽  
Artificial Chromosome ◽  
Regulatory Elements ◽  
Chromatin Domain ◽  
Open Chromatin ◽  
Cis Elements

Identifying the full repertoire of cis elements required for gene expression in mammalian cells (or animals) is challenging, given the moderate sizes of many loci. To study how the human gp91-phox gene is expressed specifically in myeloid hematopoietic cells, we introduced yeast artificial chromosome (YAC) clones and derivatives generated in yeast into mouse embryonic stem cells competent to differentiate to myeloid cells in vitro or into mouse chimeras. Fully appropriate regulation was recapitulated with a 130-kb YAC containing 60 and 30 kb of 5' and 3' flanking sequences, respectively. Immunodetection of human gp91-phox protein revealed uniform expression in individual myeloid cells. The removal of upstream sequences led to decreased overall expression which reflected largely a variegated pattern of expression, such that cells were either "on" or "off," rather than pancellular loss of expression. The proportion of clones displaying marked variegation increased with progressive deletion. DNase I mapping of chromatin identified two hypersensitive clusters, consistent with the presence of multiple regulatory elements. Our findings point to cooperative interactions of complex regulatory elements and suggest that the presence of an incomplete set of elements reduces the probability that an open chromatin domain (or active transcriptional complex) may form or be maintained in the face of repressive influences of neighboring chromatin.

scholarly journals Preotic neural crest cells contribute to coronary artery smooth muscle involving endothelin signalling

2012 ◽  
Vol 3 (1) ◽  
Author(s):  
Yuichiro Arima ◽  
Sachiko Miyagawa-Tomita ◽  
Kazuhiro Maeda ◽  
Rieko Asai ◽  
Daiki Seya ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Coronary Artery ◽  
Neural Crest ◽  
Neural Crest Cells
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