Feedback regulation of RTK signaling in development.

Precise regulation of the amplitude and duration of receptor tyrosine kinase (RTK) signaling is critical for the execution of cellular programs and behaviors. Understanding these control mechanisms has important implications for the field of developmental biology, and in recent years, the question of how augmentation or attenuation of RTK signaling via feedback loops modulates development has become of increasing interest. RTK feedback regulation is also important for human disease research; for example, germline mutations in genes that encode RTK signaling pathway components cause numerous human congenital syndromes, and somatic alterations contribute to the pathogenesis of diseases such as cancers. In this review, we survey regulators of RTK signaling that tune receptor activity and intracellular transduction cascades, with a focus on the roles of these genes in the developing embryo. We detail the diverse inhibitory mechanisms utilized by negative feedback regulators that, when lost or perturbed, lead to aberrant increases in RTK signaling. We also discuss recent biochemical and genetic insights into positive regulators of RTK signaling and how these proteins function in tandem with negative regulators to guide embryonic development

Modeling craniofacial and skeletal congenital birth defects to advance therapies.

Craniofacial development is an intricate process of patterning, morphogenesis, and growth that involves many tissues within the developing embryo. Genetic misregulation of these processes leads to craniofacial malformations, which comprise over one-third of all congenital birth defects. Significant advances have been made in the clinical management of craniofacial disorders, but currently very few treatments specifically target the underlying molecular causes. Here, we review recent studies in which modeling of craniofacial disorders in primary patient cells, patient-derived induced pluripotent stem cells (iPSCs), and mice have enhanced our understanding of the etiology and pathophysiology of these disorders while also advancing therapeutic avenues for their prevention

Bent bone dysplasia syndrome reveals nucleolar activity for FGFR2 in ribosomal DNA transcription

Fibroblast growth factor receptor 2 (FGFR2) promotes osteoprogenitor proliferation and differentiation during bone development, yet how the receptor elicits these distinct cellular responses remains unclear. Analysis of the FGFR2-skeletal disorder bent bone dysplasia syndrome (BBDS) demonstrates that FGFR2, in addition to its canonical signaling activities at the plasma membrane, regulates bone formation from within the nucleolus. Previously, we showed that the unique FGFR2 mutations that cause BBDS reduce receptor levels at the plasma membrane and diminish responsiveness to extracellular FGF2. In this study, we find that these mutations, despite reducing canonical signaling, enhance nucleolar occupancy of FGFR2 at the ribosomal DNA (rDNA) promoter. Nucleolar FGFR2 activates rDNA transcription via interactions with FGF2 and UBF1 by de-repressing RUNX2. An increase in the nucleolar activity of FGFR2 in BBDS elevates levels of ribosomal RNA in the developing bone, consequently promoting osteoprogenitor cell proliferation and decreasing differentiation. Identifying FGFR2 as a transcriptional regulator of rDNA in bone unexpectedly reveals a nucleolar route for FGF signaling that allows for independent regulation of osteoprogenitor cell proliferation and differentiation

The Roles of RNA Polymerase I and III Subunits Polr1c and Polr1d in Craniofacial Development and in Zebrafish Models of Treacher Collins Syndrome

<div><p>Ribosome biogenesis is a global process required for growth and proliferation of all cells, yet perturbation of ribosome biogenesis during human development often leads to tissue-specific defects termed ribosomopathies. Transcription of the ribosomal RNAs (rRNAs) by RNA polymerases (Pol) I and III, is considered a rate limiting step of ribosome biogenesis and mutations in the genes coding for RNA Pol I and III subunits, <i>POLR1C</i> and <i>POLR1D</i> cause Treacher Collins syndrome, a rare congenital craniofacial disorder. Our understanding of the functions of individual RNA polymerase subunits, however, remains poor. We discovered that <i>polr1c</i> and <i>polr1d</i> are dynamically expressed during zebrafish embryonic development, particularly in craniofacial tissues. Consistent with this pattern of activity, <i>polr1c</i> and <i>polr1d</i> homozygous mutant zebrafish exhibit cartilage hypoplasia and cranioskeletal anomalies characteristic of humans with Treacher Collins syndrome. Mechanistically, we discovered that <i>polr1c</i> and <i>polr1d</i> loss-of-function results in deficient ribosome biogenesis, Tp53-dependent neuroepithelial cell death and a deficiency of migrating neural crest cells, which are the primary progenitors of the craniofacial skeleton. More importantly, we show that genetic inhibition of <i>tp53</i> can suppress neuroepithelial cell death and ameliorate the skeletal anomalies in <i>polr1c</i> and <i>polr1d</i> mutants, providing a potential avenue to prevent the pathogenesis of Treacher Collins syndrome. Our work therefore has uncovered tissue-specific roles for <i>polr1c</i> and <i>polr1d</i> in rRNA transcription, ribosome biogenesis, and neural crest and craniofacial development during embryogenesis. Furthermore, we have established <i>polr1c</i> and <i>polr1d</i> mutant zebrafish as models of Treacher Collins syndrome together with a unifying mechanism underlying its pathogenesis and possible prevention.</p></div

Analysis of NCC development in <i>polr1c</i> and <i>polr1d</i> mutant embryos.

<p>(A-H) <i>sox10</i> expression at 12 hpf and (I-P) <i>foxd3</i> expression at 14 hpf reveal relatively normal patterns of early cranial NCC specification and migration in <i>polr1c</i>^-/- and <i>polr1d</i>^-/- embryos (black arrows). (Q-X) In contrast, <i>dlx2</i> expression at 36 hpf reveals slightly diminished domains of activity in mutant embryos, particularly with respect to the posterior pharyngeal arches, which is suggestive of fewer mature NCC colonizing the pharyngeal arches. White arrows indicate pharyngeal arches 1 and 2. Scale bar = 200 μm.</p

Craniofacial cartilage development is disrupted in <i>polr1c</i>^-/- and <i>polr1d</i>^-/- mutant embryos.

<p>(A-C) Alcian blue staining reveals cranial cartilage in 5 dpf <i>polr1c</i>^-/- and <i>polr1d</i>^-/- mutant embryos is hypoplastic compared to controls. (D-F) The jaws of mutant embryos are smaller overall, with noticeable differences in the size of Meckel’s cartilage, the palatoquadrate, and ceratohyal elements. (G-I) Staining of the viscerocranium reveals smaller cartilage elements derived from each of the pharyngeal arches in mutant embryos, most notably the ceratobranchials, as well as altered polarity of the ceratohyal. (J-L) Staining of the neurocranium reveals hypoplasia of the ethmoid plate. Abbreviations: M, Meckel’s cartilage; pq, palatoquadrate; ch, ceratohyal; cb, ceratobranchial; ep, ethmoid plate; pch, parachordal. Scale bar = 200 μm.</p

<i>tp53</i> inhibition ameliorates cartilage anomalies in <i>polr1d</i>^-/-mutant embryos in a dosage-dependent manner.

<p>(A-D) Alcian blue staining of cartilage in an allelic series of <i>polr1d</i> and <i>tp53</i> mutant embryos. Dosage-dependent improvement in cartilage development is particularly noticeable in the jaw (E-H), elements of the viscerocranium (I-L), and more specifically the ceratohyal (G-L). Abbreviations: M, Meckel’s cartilage; pq, palatoquadrate; ch, ceratohyal; cb, ceratobranchial. Scale bar = 200 μm.</p