Generation of an immortalized mouse embryonic palatal mesenchyme cell line

Fantauzzo, Katherine A.; Soriano, Philippe

doi:10.1371/journal.pone.0179078

Cited by 23 publications

(20 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…To evaluate intracellular pathway activation downstream of wild-type FGFR2 and the FGFR2 F , FGFR2 CPG and FGFR2 FCPG signaling mutant receptors, we generated immortalized E10.5 FNP cell lines from the respective mouse strains also carrying a mutation in Ink4A/Arf to facilitate rapid immortalization of these cells. Similar to a previous study with palatal mesenchymal cells (Fantauzzo and Soriano 2017), we found that expression of facial mesenchyme markers was similar between primary and immortalized FNP cells (iFNPs; Supplementary Figure S4B). We isolated iFNPs for respective genotypes (wt-iFNP, F-iFNP, CPG-iFNP and FCPG-iFNP).…”

Section: Signaling Mutations Disrupt Multiple Intracellular Pathwayssupporting

confidence: 89%

FGF signaling in development beyond canonical pathways

Ray

Mazot

Catela

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

FGFs are key developmental regulators which engage a signal transduction cascade through receptor tyrosine kinases, typically involving ERK1/2, PI3K/AKT, and other effectors. However, it remains unknown if all FGF activities depend on kinase activity or these canonical signal transduction cascades. To address these questions, we generated allelic series of knock-in Fgfr1 and Fgfr2 mouse strains, carrying point mutations that disrupt binding of signaling effectors to the receptors, alone or in combination. We also produced a kinase dead allele of Fgfr2 which broadly phenocopies the null mutant. When interrogated in cranial neural crest cells, point mutations in either receptor revealed discrete functions for signaling pathways in specific craniofacial contexts, but failed to recapitulate the single or double null mutant phenotypes even in their most extensive combination. Furthermore, we found that together these signaling mutations abrogated the established FGF-induced signal transduction pathways, yet certain FGF functions such as cell-matrix and cell-cell adhesion remained unaffected. Our studies establish combinatorial roles of both Fgfr1 and Fgfr2 in development and identify novel kinase-dependent cell adhesion properties of FGF receptors, independent of well-established roles in intracellular signaling.

show abstract

Section: Signaling Mutations Disrupt Multiple Intracellular Pathwayssupporting

confidence: 89%

FGF signaling in development beyond canonical pathways

Ray

Mazot

Catela

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…To do this, primary MEPM cells were dissected from E13.5 control ( Pdgfra +/fl ;Wnt1-Cre +/Tg or Pdgfrb +/fl ;Wnt1-Cre +/Tg ) and conditional knock-out ( Pdgfra fl/fl ;Wnt1-Cre +/Tg or Pdgfrb fl/fl ;Wnt1-Cre +/Tg ) littermate embryos for use in cell growth assays (Figure 8A). Primary MEPM cells are a faithful surrogate for embryonic facial mesenchyme, as wild-type cells uniformly express both PDGFRα and PDGFRβ, as well as numerous additional markers of palatal mesenchyme cells in vivo , and are responsive to stimulation with PDGF-AA, PDGF-BB and PDGF-DD ligand (He and Soriano, 2013; Fantauzzo and Soriano, 2014, 2016, 2017; Vasudevan and Soriano, 2014; Vasudevan et al, 2015). Even after a single day in growth medium containing 10% FBS, control Pdgfrb +/fl ;Wnt1-Cre +/Tg cells (0.1077 ± 0.01233 arbitrary units (AU)) (Figure 8C) had grown about half as much as control Pdgfra +/fl ;Wnt1-Cre +/Tg cells (0.2217 ± 0.07322 AU) (Figure 8B).…”

Section: Resultsmentioning

confidence: 99%

PdgfraandPdgfrbgenetically interact in the murine neural crest cell lineage to regulate migration and proliferation

Long

Fantauzzo

2020

Preprint

Self Cite

View full text Add to dashboard Cite

Cranial neural crest cells (cNCCs) are migratory, multipotent cells that originate from the forebrain to the hindbrain and eventually give rise to the bone and cartilage of the frontonasal skeleton, among other derivatives. Signaling through the two members of the platelet-derived growth factor receptor (PDGFR) family of receptor tyrosine kinases, alpha and beta, plays critical roles in the cNCC lineage to regulate craniofacial development during murine embryogenesis. Further, the PDGFRs have been shown to genetically interact during murine craniofacial development at mid-to-late gestation. Here, we examined the effect of ablating both Pdgfra and Pdgfrb in the murine NCC lineage on earlier craniofacial development and determined the cellular mechanisms by which the observed phenotypes arose. Our results confirm a genetic interaction between the two receptors in this lineage, as phenotypes observed in an allelic series of mutant embryos often worsened with the addition of conditional alleles. The defects observed here were shown to stem from reduced cNCC stream size and aberrant cNCC directional migration, as well as decreased proliferation of the facial mesenchyme upon combined decreases in PDGFRα and PDGFRβ signaling. Importantly, we found that PDGFRα plays a predominant role in cNCC migration whereas PDGFRβ primarily contributes to proliferation of the facial mesenchyme. Our findings provide insight into the distinct mechanisms by which PDGFRα and PDGFRβ signaling regulate cNCC activity and subsequent craniofacial development in the mouse embryo.

show abstract

“…This remodeling is likely initiated by the medial edge epithelium of the palatal shelf, but driven by the palate mesenchyme. While primary MEPM cells have been used for testing the effect of various inhibitory compounds, growth factors, and gene-expression changes (Fantauzzo and Soriano 2014;Gao et al 2019;Iyyanar and Nazarali 2017;Jiang et al 2017;Liu et al 2014;Vasudevan and Soriano 2014), only a few studies have investigated the migration of MEPM cells (Fantauzzo and Soriano 2017;Gao et al 2019;He and Soriano 2013), and none, to our knowledge, have examined if they can migrate collectively. We identify the presence of stream formation in MEPM cells, a behavior generally considered to be a feature of collective movement (Czirok et al 2013;Scarpa and Mayor 2016;Szabo et al 2010).…”

Section: Discussionmentioning

confidence: 99%

SPECC1L-deficient palate mesenchyme cells show speed and directionality defect

Goering

Isai

Hall

et al. 2019

Preprint

View full text Add to dashboard Cite

Clefts of the lip and/or palate (CL/P) are common anomalies that occur in 1/800 live births. Pathogenic SPECC1L variants identified in patients with rare atypical clefts and syndromic CL/P suggest the gene plays a primary role in face and palate development. We have generated Specc1l gene-trap (Specc1l cGT ) and truncation (Specc1l DC510 ) alleles that cause embryonic or perinatal lethality, respectively.Specc1l cGT/DC510 compound mutants show delayed and abnormal palatal shelf elevation at E14.5. By E15.5, the mutant shelves do elevate and fuse, however, the palatal rugae form abnormally. Palatogenesis requires extensive mesenchymal remodeling, especially during palatal shelf elevation. We posit that this remodeling involves collective movement of neural crest-derived palatal mesenchyme cells. Live time-lapse microscopy was performed to visualize in vitro wound-repair assays with wildtype and SPECC1L-deficient primary mouse embryonic palatal mesenchyme (MEPM) cells. SPECC1L-deficient MEPM cells consistently showed delayed closure in wound-repair assays. To evaluate which features of cellular movement were responsible, we performed automated particle image velocimetry (PIV) and manual cell tracking. The analyses revealed that both cell speed and directionality are disrupted in SPECC1L-deficient cells compared to controls.To determine if primary MEPM cells can move collectively, we assayed stream formation, which is a hallmark of collective movement. Indeed, MEPM cultures displayed correlated movement of neighboring cells. Importantly, correlation length was reduced in SPECC1Ldeficient cultures, consistent with a role for SPECC1L in collective migration.Furthermore, we demonstrated that activation of the PI3K-AKT pathway with the 740Y-P small molecule can rescue the wound-closure delay in SPECC1L-deficient MEPM cells. 3Cell tracking analyses showed that this rescue was due to both increased speed and improved directionality. Altogether, our data showed a novel role for SPECC1L in guided movement through modulation of PI3K-AKT signaling.

show abstract

Generation of an immortalized mouse embryonic palatal mesenchyme cell line

Cited by 23 publications

References 44 publications

FGF signaling in development beyond canonical pathways

FGF signaling in development beyond canonical pathways

PdgfraandPdgfrbgenetically interact in the murine neural crest cell lineage to regulate migration and proliferation

SPECC1L-deficient palate mesenchyme cells show speed and directionality defect

Contact Info

Product

Resources

About