The RNA-binding protein Rbm24 is evolutionarily conserved, and its coding gene displays tissue-specific expression in vertebrates. However, the dynamic localization of this protein in different cell lineages remains elusive. We have generated a zebrafish rbm24a-GFP knock-in line in which endogenous Rbm24a is tagged with GFP, allowing the precise monitoring and systematic characterization of its spatiotemporal expression and subcellular localization during development and in the adult. Rbm24a-GFP not only shows strongly restricted expression in a subset of tissues, but also displays cell type- and stage-specific subcellular localization patterns. The protein mainly localizes in the cytoplasm of lens fiber cells and progenitors of sensory hair cells. It undergoes dynamic cytoplasm to nucleus translocation during differentiation of myoblasts and cardiomyoblasts. We further examined the effectiveness of this knock-in line for inhibiting Rbm24a function. Targeted degradation of Rbm24a-GFP using the zGrad system produces phenotypes of zygotic rbm24a mutants or morphants, with defective heart morphogenesis and disrupted cardiac muscle integrity. Therefore, this line will be particularly useful for understanding Rbm24a-GFP dynamic expression and localization changes under homeostasis and pathological conditions. It also enriches the resource of zebrafish knock-in line and provides a convenient tool for functional study of the protein through degron-mediated conditional degradation.

The maternal-to-zygotic transition (MZT) signifies the shift from maternal genome control to zygotic genome control during zygote development. However, the regulatory role of non-coding RNAs (ncRNA), such as micro RNA (miRNA) and long non-coding RNAs (lncRNAs), in plants during MZT remains inadequately studied. Furthermore, the competitive endogenous RNA network (ceRNA), which involves interactions among messenger RNA (mRNA), miRNA, and lncRNA across developmental stages, has not been thoroughly investigated. To address this gap, we conducted an analysis of previously generated RNA-Seq data from egg and zygotes at 14 and 24 h after pollination, and 1 cell embryos of the Columbia ecotype in Arabidopsis thaliana. Through this analysis, we identified over 1900 differentially expressed mRNAs, 80 known lncRNAs, and 300 novel lncRNAs. Additionally, we identified lncRNAs specific to the MZT transition phases and observed differential expression of experimentally validated lncRNAs throughout MZT. Furthermore, we predicted hub-miRNAs across the three distinct stages of zygote development. Overall, our study provides valuable insights into the interactions among mRNA, miRNA, and lncRNA, as well as the differential rewiring patterns of the ceRNA networks during MZT.

How vertebrate skeletal muscle size is regulated and balanced with body size over the life-course is unclear, but is important for human health and quality of life. Muscle growth occurs by increase in myofibre number (hyperplasia) and enlargement of existing fibres (hypertrophy). Fibre enlargement reflects either hypernucleation, an increase in myofibre nuclei, and/or hyperoidy, an increase in nuclear domain size (NDS), the volume of myofibre per myonucleus. Quantitative time lapse imaging of muscle cellularity indicates that myotome growth in early larval zebrafish is dominated by hyperoidy, with lesser contribution by hypernucleation. Addition of small new myofibres makes a quantitatively even smaller contribution to growth. During neonatal mouse muscle growth a distinct balance of different growth mechanisms occurs, but yields quantitively similar hyperoidy. In zebrafish, the number of myofibres and myonuclei continue to increase in the absence of independent feeding, whereas NDS shrinks and whole body growth falters without adequate food intake from 5 days post-fertilisation, despite the continued availability of yolk. The persistent accrual of myonuclei while fibres undergo atrophy in response to starvation we term muscle sparing. Myofibre volume increases more than myofibril content during growth. During atrophy, in contrast, cytoplasmic puncta containing sarcolemmal markers become associated with autophagosomes and lysosomes, and myofibrils fill a larger fraction of the remaining sarcoplasm. These observations lead us to propose a 'shopping bag' hypothesis for myofibre hyperoidy and atrophy, whereby change in sarcolemmal area and myofibre volume (the 'bag') precede, and may be required for, changes in the myofibril content (the 'shopping'). The distinct regulation of three muscle growth mechanisms in developing vertebrate models predict similar controls on human muscle growth which, given the importance of skeletal muscle for whole body metabolic health, are of potential relevance to the developmental origins of human health and disease.

During amphibian metamorphosis, the intestine is remodeled from larval to adult form. In this study, we investigated the expression and localization of Xenopus laevis Tead4 (a key transcription factor in the Hippo pathway) in the metamorphosing intestine, examined its interaction, and analyzed its phylogenetic relationships. Quantitative RT-PCR revealed that both tead4.L and tead4.S (homeologs) transcripts are expressed throughout natural and thyroid hormone (TH)-induced metamorphosis, with modest but stage-specific changes, indicating sensitivity to TH. Triple hybridization chain reaction RNA-FISH showed that tead4 expression is broadly distributed in the intestine, with its signal intensity increasing from the metamorphic climax onward, whereas yap1 expression is enriched in the connective and muscle layers. There was a partial overlap between tead4 and yap1 expression, indicating context-dependent co-regulation, and incomplete co-localization of the expressed proteins, suggesting alternative co-factors. Oocyte co-immunoprecipitation assays demonstrated that both Tead4.L and Tead4.S physically associate with Yap1, supporting their role as Hippo pathway effectors. Phylogenetic analysis placed X. laevis Tead4 homeologs within the TEAD2 clade, indicating that the genes annotated as tead4 can more accurately be classified as tead2. These findings demonstrate that Tead4 homeologs function as Yap1 partners and contribute to transcriptional control during amphibian intestinal metamorphosis, highlighting their conserved role as effectors of Hippo signaling.

The use of time-lapse imaging to study neural tube closure in mouse embryos has provided unexpected insights into the complex morphogenetic processes involved. When neural tube closure is disrupted, it leads to neural tube defects (NTDs), which are among the most common structural birth defects in humans, associated with long-term disabilities and death. This review explores the growing body of research on time-lapse imaging experiments conducted in mice, emphasizing discoveries of the dynamic cellular movements and changes that enable neural tube formation. Advances in mouse embryo culture and live imaging techniques have enabled visualization of dynamic cellular movements and shape changes during neural tube formation, allowing researchers to observe abnormal cell behaviors in genetic mouse models with neural tube closure defects. These studies use transgenic reporters, conditional mouse genetics, and various physical and pharmacological interventions to track tissue and cell behavior and elucidate the underlying molecular and biophysical mechanisms as neural folds rise and fuse at the dorsal midline. Observing neural tube closure in real time has led to important findings, including revealing the crucial role of the surface ectoderm in supporting neural fold elevation and fusion. The coordination of apical constriction with cell cycle progression and apoptosis helps shape the neural plate. Analyzing convergent extension shows that oriented neighbor exchanges-requiring planar cell polarity signaling-drive polarized protrusive activity and actomyosin contractility, along with coordinated apical constriction to elevate and bring the neural folds together. Future innovations are expected to improve the measurement of biomechanical forces during neural tube formation and visualization of deep tissues to clarify mechanisms of cranial mesenchyme morphogenesis during cranial neural fold elevation.

Hybridization chain reaction RNA-fluorescent in situ hybridization (HCR RNA-FISH) is a powerful and increasingly used method for visualizing gene expression in cells and tissues. A probe set against polyadenylated RNA (poly(A)) is often used as a positive control for RNA integrity and staining quality. While optimizing this technique in the ovary of the brown anole lizard (Anolis sagrei), we found that the poly(A) probe produced a strikingly specific and intense signal in pyriform cells, a specialized lizard-specific nurse cell type. This staining pattern was found in both whole-mount samples and paraffin sections, suggesting that poly(A) signal intensity can serve as a robust molecular marker for this cell type. The specific and robust signal facilitated segmentation of volumetric data to create the first 3D models of pyriform cells. We also observed unusually diffuse DAPI staining in pyriform cell nuclei, distinguishing them from surrounding granulosa cells, pointing to possible differences in chromatin structure or nuclear organization. Together, these findings highlight the potential of poly(A) probes used in HCR RNA-FISH not only as a technical control, but also as a tool to selectively label specific cell types with high transcriptional activity or storage of abundant poly(A) transcripts.

Developmental biology is one of the fundamental sciences for understanding the basics of life and often intersects with social justice challenges facing society. This article describes an inclusive teaching activity for students and instructors to explore the interface between developmental biology, genetic diversity, and social justice. The instructor and students will choose a recent publication and use it as the basis for exploring the roles of specific genes characterized in autism from educational, emulative, and ethical perspectives. The assignment for students will include a discussion and demonstration of developmental neurobiology and principles of gene function within the nervous system, as well as ethical considerations for how individuals, as well as society as a whole, should consider genetic variations. Two frameworks are introduced for instructors to create an inclusive learning environment, including universal design for learning and multipartiality. Resources and examples are given throughout the article for instructors to use, and a suggested rubric is also provided. A post-activity self-reflection performed by the students will facilitate their own assessment of how the teaching activity has impacted their philosophical and social perspectives on genetic diversity. The short-term goal of the activity is to promote an immediate appreciation of neurodiversity among the participating students, and the long-term goal is to demonstrate the importance of neurodiversity for developing a just society.

Tissues comprise not only cells, but also extracellular matrix (ECM), which is assembled from some of the most abundant molecules in animals. ECM composition and organization underlie the tissue architecture of most organs by design and is thus optimized for their principal physiologic functions. While the structural role of ECM is widely accepted, its regulatory role is often overlooked. We identified a crucial role for ECM proteoglycans aggrecan and versican and the proteases which degrade them in ensuring dimorphism of the umbilical arteries and vein in relation to rapid closure of the umbilical arteries at birth. Another study which sought to define the roles of proteases in the vasculature, unexpectedly uncovered a requirement for ADAMTS9, a secreted protease, in trimming pericellular versican in the late-gestation myometrium. This activity was found to be necessary for priming the myometrium for sufficiently strong uterine contraction at parturition. These studies and others showed that fine tuning of umbilical vascular and myometrial ECM is crucial for survival of the species and illustrate how regulatory processes of the ECM can modulate smooth muscle cell phenotype. In addition to the outcomes, the underlying narrative illustrates how a mix of curiosity, happenstance, exploration of tangents, interdisciplinary collaboration and generous sharing of resources is rewarding and has a significant role in scientific discoveries.

Nuclear receptors are iteratively deployed during neural crest development, from pre-induction through differentiation stages. NR2F1 and NR2F2 in particular have been proposed as broad regulators of early neural crest gene expression in mammals, but the timing, extent, and redundancy of their developmental requirement has remained unclear, as Nr2f1 and Nr2f2 single mouse mutants present only minimal craniofacial phenotypes. Here we report the dynamic expression patterns of Nr2f1 and Nr2f2 in the mouse cranial neural crest from specification through post-migratory stages. Combined conditional knockout of both Nr2f1 and Nr2f2 in the neural crest with Wnt1-Cre or Pax3 caused severe midfacial clefting, loss of the maxilla and palate, and hypoplasticity of all other facial skeletal elements except the distal mandible. These perinatal phenotypes were rooted in a major shortage of pharyngeal arch mesenchyme at mid-gestation. This in turn traced to a deficiency of migrating neural crest cells, first evident in the trailing part of the first arch migratory stream at embryonic day 8.75. RNAseq at a slightly earlier stage revealed downregulation of many migratory neural crest genes, including a possible direct target, the phospholipase Plcg2. These findings reveal a vital requirement for NR2F1/2 within the later-forming cranial neural crest.

Cell migration is an important process underlying animal embryonic body patterning, organogenesis, and diseases like metastatic cancer. Acetylcholine (ACh) signaling plays a key role in the migration of various cell types and cancer cells, yet in vivo studies are lacking. We investigated the function of nicotinic ACh receptors (nAChRs) on the migration of a gonadal leader cell, the linker cell (LC). During C. elegans male gonadogenesis, the LC migrates posteriorly along the ventral body wall, following a path that runs parallel and adjacent to the ACh-releasing ventral nerve cord (VNC). Excess ACh reoriented the polarity of the LC from posterior-facing to anterior-facing through an intermediate stage of facing the ventral body wall. nAChRs, which are expressed by both the VNC and LC, were required for the LC reversal response. The specific combination of subunits of the pentameric nAChR produced different reversal responses, with acr-16(-) and lgc-9(-) mutants inhibiting and acr-15(-) promoting reversals. LC reversal in response to excess ACh also required the L1 cell adhesion molecule (L1CAM), SAX-7, which is expressed by both VNC and LC. We propose that an increase in ACh signaling in the VNC and LC promotes stronger SAX-7 mediated adhesion of the LC to the ventral body wall, causing the LC to change directions from posterior to ventral facing.

Metamorphosis is a key event in development that is conserved in many marine organisms. Ciona intestinalis type A induces metamorphosis through the settlement of papillae onto the substrate. The papilla consists of collocytes (CCs), primary sensory neurons (PSNs), and axial columnar cells (ACCs), but it remains unclear whether PSNs alone can induce metamorphosis. Manipulating single neurons is crucial for elucidating the neural network system that drives metamorphosis. In this study, we developed an optogenetic system in which ChrimsonR, a red-shifted mutant of channelrhodopsin, was expressed exclusively in PSNs, enabling metamorphosis to be induced by light stimulation. A Ciona-optimized self-cleaving peptide, T2A, was used to co-express the Ca indicator GCaMP6s, allowing us to monitor neural activity during light stimulation. Activation of PSNs alone induced a series of metamorphic events, including epidermal backward movement, mesenchymal cell extravasation, and tail regression. Furthermore, we confirmed that metamorphosis proceeded to the juvenile stage. Metamorphosis was induced even with intermittent light stimulation, and the total stimulation time required for its initiation was approximately 6 min. The optogenetic system developed in this study may significantly contribute to elucidating the link between neuronal function and metamorphosis at the single-cell level.

This study employs an eco-friendly approach to synthesize superparamagnetic iron oxide (FeO) nanoparticles (SPIO) using Aegle Marmelos (A. Marmelos) pulp extract as a surfactant as well as a reducing agent. The pulp extract from A. marmelos is medicinally employed to treat cholera, diabetes, skin infections, earaches, blood purification, and heart problems. Further, the XRD and TEM analyses confirmed the formation of SPIO nanoparticles with a cubic structure and crystallite sizes ranging from 5 to 12 nm. The FE-SEM showed that the SPIO displayed a uniform distribution with quasi-spherical morphology. FTIR evaluation directed the existence of iron-oxygen (Fe-O) bonds, while XPS analysis confirmed iron (Fe) in both +3 and + 2 oxidation states. SQUID studies verify the superparamagnetic nature of the material, with a magnetization (Ms) of 42.02 emu/g. Following characterization, the hyperthermia performance and specific loss power (SLP) of SPIO nanoparticles were systematically explored to assess their dependency on concentration, frequency, and the alternating magnetic field (AC field). These SPIO nanoparticles exhibit excellent hyperthermia proficiency (42-45 °C), with SLP values of 153.48 and 40.33 W/g at concentrations of 1 mg/mL in aqueous media (DI HO) and ethylene glycol media (E.G.), respectively, under an AC field (400 A). Furthermore, different concentrations of SPIO were tested for acute toxicity using a static renewal bioassay method. The results indicate non-toxic behavior towards vital organs such as the ovaries, gills, liver, heart, kidneys, brain, and muscles of the benthopelagic fish Cirrhinusmrigala. These findings highlight the potential of the SPIO nanoparticles as biocompatible for magnetic hyperthermia applications (MHT). These newly developed SPIO nanoparticles are suitable for deployment in the medical field, as they exhibit remarkable performance in the treatment of MHT when exposed to an AC field.

From inception, the environment plays a critical role, first in the development of an organism, and later, as adults, in how an organism through homeostatic processes may undergo changes in response to external exposures or insults. Exposure during early development can impact future susceptibility to diseases, and adult exposure has been observed to cause epigenetic changes that can be passed down to generations. This review aims to address the dual aspects of environmental response; 1) at the level of an organism, focusing on how an organism adapts or responds to environmental toxins (yin), and 2) at the cellular level, examining how a single cell develops into a complete organism (yang). Bridging the gap between these two forms of exposure, the intrinsic at level of a cell and the extrinsic at the level of an organism, and their response is becoming increasingly critical due to changes in our current exposure landscapes and the consequent need to understand the etiologies behind the rise in various developmental and diseased states.

During kidney formation, segmented epithelial tubules and blood vessels develop within a heterogeneous and progressively patterned stroma. By E18.5, the murine renal stroma exhibits several transcriptionally and spatially distinct populations, including specialized stromal cells associated with the vasculature, termed mural cells. However, the precise contributions of stromal progenitor lineages to this stromal heterogeneity, as well as the dynamics of renal mural cell investment, remain unclear. Previous studies have described stromal progenitors in the developing cortex that transiently express the transcription factor Foxd1, as well as stromal progenitors in the ureter that express Tbx18, and have shown that both are capable of giving rise to renal stromal cells, including vascular mural cells. Here, we use pulse induction of Tbx19CreERT2 at different timepoints to elucidate the contribution of the Tbx18 population to stromal patterning. We show that the Tbx18-lineage, when induced at E12.5, gives rise to arterial mural cells, without ever progressing through a Foxd1+ cortical stromal progenitor state. These arterial mural cells are only transiently present along arteries during development, ultimately contributing instead to peritubular capillaries. When traced post-natally, the Tbx18-lineage gives rise to pericytes, which are enriched in S3-segment-associated, Cxcl14-enriched stroma in the inner cortex. We show that these pericytes arise directly from arterial mural cells seen earlier during development. These data help clarify a small portion of the complicated lineage relationships of renal stromal progenitors and their contribution to the kidney vascular-associated mural cells.

Amphibian metamorphosis is tightly regulated by thyroid hormone (TH). During this process, most larval epithelial cells in the Xenopus laevis intestine undergo apoptosis, whereas a small population dedifferentiates into adult epithelial stem cells. They subsequently proliferate and differentiate to form a trough-crest epithelial architecture similar to the mammalian crypt-villus axis. We have previously identified a number of TH-responsive genes likely involved in this intestinal remodeling. Here, we focus on one such gene, folate receptor 4 (folr4). We examined the spatiotemporal expression of folr4.L by using quantitative RT-PCR and in situ hybridization chain reaction (HCR) and found that folr4.L expression is highly upregulated during both natural and TH-induced metamorphosis. Interestingly, in the epithelium at the climax of metamorphosis, folr4.L is specifically expressed in the proliferating and/or differentiating adult epithelial cells located adjacent to proliferating adult stem cells, which express intestinal stem cell marker leucine-rich repeat-containing G protein-coupled 5 (lgr5). Moreover, we identified a TH response element (TRE) in the folr4.L promoter that binds to the heterodimer of TH receptor (TR) and 9-cis retinoic acid receptor (RXR) in vitro and mediates T3-dependent transcriptional activation in vivo. Phylogenetic analysis suggested that X. laevis Folr4.L may be more closely related to riboflavin binding protein (Rfbp) than mammalian FOLR4. These findings suggest that TH-induced Folr4.L might be involved in the development of adult intestinal epithelium.

Perivascular fibroblasts (PVFs) are a cell type associated with large diameter blood vessels in the brain and spinal cord parenchyma and leptomeninges. PVFs have previously defined roles in injury and neuroinflammatory diseases and predicted roles in supporting neurovascular function. The temporal dynamics of PVF development in the pre- and postnatal cerebral cortex have recently been described, however the molecular mechanisms that underly PVF development have not been identified. PVFs express both platelet-derived growth factor receptors (PDGFRs), PDGFRα and PDGFRβ. Here we investigate the role of PDGF signaling in PVF development. We use immunohistochemistry and RNA transcript detection methods to show developmental expression of PDGFRs by PVFs and examine distribution of PDGF ligand expression in the brain. We show that postnatal deletion of PDGFRα in fibroblasts using the Col1a2-CreERT mouse line impairs PVF coverage of cerebral vessels at postnatal day 10. Perivascular macrophages, a cell type previously shown to co-develop with PVFs, have impaired cerebral vessel coverage in conditional mutants that is similar to PVF coverage defects. This work establishes a requirement for PDGFRα signaling in PVF development and may shed light upon the potential pathways that are over-activated in PVFs in injury and disease contexts.

Developmental biology stands at a crossroads. While some have suggested the field is in decline, we, early-career developmental biologists, see an era of renewal driven by conceptual expansion, technical innovation, and cross-disciplinary integration. In this Commentary, we reflect on discussions from a 2024 workshop of postdoctoral scholars from across North America, outlining shared challenges and opportunities that will shape the field's future. We argue that the perceived crisis in developmental biology stems not from a lack of relevance, but from a narrow definition that overlooks its broader reach, from embryogenesis to regeneration, stem cell biology, aging, and environmental responsiveness. We highlight how emerging model organisms, single-cell systems, and advances in imaging and genomics now enable comparative and mechanistic insights across the tree of life. To sustain this progress, we call for renewed investment in basic research, structural reforms to support early-career scientists, and accessible community-driven resources for emerging model systems. Finally, we emphasize the importance of public engagement, equitable mentorship, and acknowledgment of the field's complex history to foster an inclusive and resilient scientific community. Together, these efforts can reprogram the trajectory of developmental biology and secure its central place in understanding the origins and dynamics of life.

In many species, the differential localization of RNAs along the animal-vegetal axis is established during oogenesis. The resulting asymmetry is essential for axis formation, germ layer patterning, and cell fate determination, especially in fish and amphibians. In recent years, research in this field has focused mainly on zebrafish, which raises the question about the conservation of localization processes across all teleost species. Although extant teleost species utilize meroblastic cleavage only, there are extreme differences in their oocyte size. These differences are fundamentally linked to each species' life history. Some have rapid embryonic development, while embryos of other species, like salmonids, take weeks to develop. This might have consequences on the spatial distribution of biomolecules during oogenesis and their relocalization during early embryogenesis. Yet, our knowledge is based on data from small-sized oocyte species with rapid development only (e.g. zebrafish). In this study, we performed a spatially resolved TOMO-seq method on early embryos of the rainbow trout, a species characterized by prolonged embryonic development and large oocytes, and compared it with zebrafish. We revealed that the maternal pre-patterned localization of transcripts can be disrupted in the early embryo by two main mechanisms: de novo transcription and degradation. The most prominent change can be seen in the emerging blastodisc in the animal pole, where there is a significant increase in localized transcripts. In contrast with research suggesting active relocalization of RNAs by ooplasmic streaming in zebrafish, we hypothesized that the change in RNA localization is caused by regionalized zygotic transcription in trout. Regardless of these differing mechanisms, the cross-species comparison revealed a conservation of many transcripts involved in germ cell development and cell proliferation. Moreover, using hybrid trout embryos, we were able to reveal the early onset of de novo transcription. Altogether, these findings indicate how species with large oocytes and prolonged development utilize unique RNA localization strategies. This knowledge expands our understanding of early development across teleost species.

The understanding of how drugs interact with carrier proteins is crucial in the field of pharmacology and the life sciences, especially in the field of drug invention. In the present work described the molecular interaction of pharmaceutically important phyto-molecule khellin on bovine serum albumin. Khellin is recognized for its ability to widen blood vessels, making it useful for heart health. It's a major component of the plant Ammi visnaga and Dioscorea species helps to protect the heart. Bovine serum albumin (BSA) is a model protein of Human serum albumin hence, BSA has been used for drug-binding properties studies. Various biophysical techniques to examine the interactions between khellin and BSA. The biophysical techniques such as fluorescence quenching by fluorescence spectroscopy studies, micro-environmental changes by synchronous fluorescence, protein structural changes by circular dichroism spectroscopy, molecular docking, ADMET properties studies, SWISS Target Prediction for target analysis and pharmacokinetic analysis by insilico. The Khellin-BSA interaction was examined using fluorescence analysis, which showed that the binding constant was 1.29 ± 0.2 × 10 M and binding free energy was -7.99 kcal/mol by invitro. The binding energy was compared with computation molecular docking studies of ligand and protein interaction showed the binding energy of -5.1 kcal/mol. It is nearer to the in vitro binding energy values of khellin on BSA. The micro-environmental changes of the ligand-protein complex were observed with peak shifts at Δλ15 for tyrosine, Δλ60 for tryptophane, and Δλ90 for phenylalanine. Also, the secondary structural changes of BSA after titrating the khellin were observed and found that there were secondary structural changes in the free BSA after adding the khellin. With possible targets found through SWISS Target Prediction, khellin is a promising druggable candidate, according to ADMET analysis, which revealed zero violations. Finally, we concluded that the Phyto-active constituent khellin possesses good binding affinity on BSA. Further, it can be taken for drug discovery experiments on clinical trials.

The zebrafish otic vesicle initially develops with only two sensory maculae, each with distinct functions. The anterior utricular macula is indispensable for vestibular function, while the posterior saccular macula is the primary auditory endorgan in zebrafish. The unique identities of these maculae are specified in the early otic vesicle by differing levels of Fgf vs. Shh signaling, but few downstream effectors have been identified. pou3f3b is the only saccule-specific marker known, but its function has not been established. We generated a knockout allele of pou3f3b and found that it causes a persistent delay in accumulation of saccular hair cells due to a failure to activate saccular expression of fgf3. In addition, saccular hair cells exhibit reduced expression of Otoferlin caused by ectopic expression of neurog1. Defects in saccular hair cell development are fully rescued by misexpressing fgf3 or knocking down neurog1. Misexpression of pou3f3b causes loss of utricular pax5 expression and further truncates neurog1 in the posterior otic vesicle but does not otherwise alter macular development. In addition to regulating saccular development, pou3f3b is also expressed in a previously undescribed population of non-neuronal cells that delaminate from the otic vesicle and migrate together with developing neuroblasts to promote their maturation. Mutant neuroblasts show a marked delay in activation of expression of neurod1, causing a transient delay in accumulation of mature SAG neurons. Thus pou3f3b is required for timely development of SAG neurons and saccular/auditory hair cells.