The olfactory epithelium (OE) has the remarkable capability to constantly replace olfactory receptor neurons (ORNs) due to the presence of neural stem cells (NSCs). For this reason, the OE provides an excellent model to study neurogenesis and neuronal differentiation. In the present work, we induced neuronal degeneration in the OE of Xenopus laevis larvae by bilateral axotomy of the olfactory nerves. We found that axotomy induces specific- neuronal death through apoptosis between 24 and 48h post-injury. In concordance, there was a progressive decrease of the mature-ORN marker OMP until it was completely absent 72h post-injury. On the other hand, neurogenesis was evident 48h post-injury by an increase in the number of proliferating basal cells as well as NCAM-180- GAP-43+ immature neurons. Mature ORNs were replenished 21 days post-injury and the olfactory function was partially recovered, indicating that new ORNs were integrated into the olfactory bulb glomeruli. Throughout the regenerative process no changes in the expression pattern of the neurotrophin Brain Derivate Neurotrophic Factor were observed. Taken together, this work provides a sequential analysis of the neurodegenerative and subsequent regenerative processes that take place in the OE following axotomy. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 77: 1308-1320, 2017.

The molecular events driving specification of the kidney have been well characterized. However, how the initial kidney field size is established, patterned, and proportioned is not well characterized. Lhx1 is a transcription factor expressed in pronephric progenitors and is required for specification of the kidney, but few Lhx1 interacting proteins or downstream targets have been identified. By tandem-affinity purification, we isolated FRY like transcriptional coactivator (Fryl), one of two paralogous genes, fryl and furry (fry), have been described in vertebrates. Both proteins were found to interact with the Ldb1-Lhx1 complex, but our studies focused on Lhx1/Fry functional roles, as they are expressed in overlapping domains. We found that Xenopus embryos depleted of fry exhibit loss of pronephric mesoderm, phenocopying the Lhx1-depleted animals. In addition, we demonstrated a synergism between Fry and Lhx1, identified candidate microRNAs regulated by the pair, and confirmed these microRNA clusters influence specification of the kidney. Therefore, our data shows that a constitutively-active Ldb1-Lhx1 complex interacts with a broadly expressed microRNA repressor, Fry, to establish the kidney field.

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Significance CHARGE syndrome is a complex developmental disorder caused by mutations in CHD7 (chromodomain helicase DNA-binding protein-7). We identified Caenorhabditis elegans chd-7 in a screen for suppressors of dauer formation, an alternative larval stage that develops under harsh environmental conditions. We found chd-7 regulates tumor growth factor-β (TGF-β) signaling pathways both for dauer diapause and for development of the cuticle, a specialized extracellular matrix. In frog embryos, Chd7 promotes Col2a1 expression, which is necessary and sufficient to prevent CHARGE features. These studies establish a conserved role for Chd7 from worms to vertebrates in regulating the TGF-β signaling pathway. Genetic dissection of chd-7 ’s role in C. elegans may help to define the molecular and cellular events that contribute to CHARGE syndrome.

Gastrulation is a key event in animal embryogenesis during which the germ layers precursors are rearranged and the embryonic axes are established. Cell polarization is essential during gastrulation driving asymmetric cell division, cell movements and cell shape changes. Furry (Fry) gene encodes an evolutionarily conserved protein with a wide variety of cellular functions mostly related to cell polarization and morphogenesis in invertebrates. However, little is known about its function in vertebrate development. Here we show that in Xenopus, Fry participates in the regulation of morphogenetic processes during gastrulation. Using morpholino knock-down, we demonstrate a role of Fry in blastopore closure and dorsal axis elongation. Loss of Fry function drastically affects the movement and morphological polarization of cells during gastrulation, in addition to dorsal mesoderm convergent extension, responsible for head-to-tail elongation. Finally, we demonstrate a functional interaction between Fry and NDR1 kinase, providing evidence of an evolutionarily conserved complex required for morphogenesis.

CHARGE syndrome is a complex developmental disorder caused by mutations in the chromodomain helicase DNA-binding protein7 (CHD7) and characterized by retarded growth and malformations in the heart and nervous system. However, despite the public health relevance of this disorder, relevant targets of CHD7 that relate to disease pathology are still poorly understood. Here we report that chd-7, the nematode ortholog of CHD7, is required for dauer morphogenesis, lifespan determination, and stress response. Genetic epistasis placed chd-7 in the TGF-β pathway. Consistent with our discoveries, we found chd-7 to be allelic to scd-3, a previously identified dauer suppressor from the TGF-β pathway. Interestingly, DAF-12 transcriptionally upregulated chd-7, which is necessary to repress daf-9 for execution of the dauer program. Transcriptomic analysis comparing chd-7 defective and normal dauers showed enrichment of collagen genes, consistent with a conserved role for the TGF-β pathway in expression of the extracellular matrix. To validate a conserved function for chd-7 in vertebrates, we used Xenopus laevis embryos, an established model to study craniofacial development. Morpholino mediated knockdown of Chd7 led to embryonic lethality, a reduction in col2a1 mRNA levels and craniofacial defects in tadpoles. Both lethality and malformations were partially rescued in Chd7-depleted embryos by over-expression of col2a1. We suggest that pathogenic features of CHARGE syndrome caused by Chd7 mutations, such as craniofacial malformations, result from the reduction of collagen levels. These studies establish C. elegans as an amenable animal model to study the etiology of the developmental defects associated with pathogenic Chd7.