Development Growth & Differentiation最新文献

Zebrafish pou5f3 encodes a Class V POU transcription factor, Pou5f3, which regulates various developmental processes, including neurogenesis and brain formation. In the current study, we attempted to comprehensively identify the Pou5f3 downstream genes around the end of epiboly, when the competence of the mid-hindbrain region to Pou5f3 suppression changes drastically, by the microarray method and a heat-inducible dominant-interference pou5f3 gene (en-pou5f3) that functionally suppresses pou5f3. At late epiboly and early somitogenesis stages, we identified genes whose expression was altered in en-pou5f3-induced embryos, revealing numerous genes regulated differently by Pou5f3 at the two stages. The validity of the microarray data was confirmed by whole mount in situ hybridization and quantitative RT-PCR. Many of the downstream genes were implicated by the Gene ontology (GO) analyses in transcriptional regulation and neural development and were enriched with sox genes and bHLH genes such as her genes. Interestingly, we noticed a tendency that Notch-dependent her genes were activated, whereas Notch-independent her genes were downregulated by Pou5f3 suppression. Among the Notch-independent her genes, her3, which is orthologous to mammalian Hes3, was suggested to be strongly activated endogenously by Pou5f3. In the upstream DNA of this gene, we found two noncoding conserved sequences (NCRs), which harbored consensus binding sites for Pou5f3, Sox, and Nanog. We further showed in reporter assays that the transcriptional regulatory activity of the her3 upstream DNA was strongly enhanced by SoxB1, and this SoxB1-mediated activation was weakened by Pou5f3. Deletion experiments showed that both upstream NCRs were involved in transcriptional repression.

In living organisms, including humans, the developmental processes that construct their morphology from a single fertilized egg are influenced not only by genetic regulation but also by various external factors. One such factor is mechanical stimulation. Although mechanical forces are suggested to contribute to brain formation during development, quantitative information on intraventricular pressure during neurogenesis remains limited. We developed a high time-resolution system efficiently using a piezoresistive sensor to measure brain intraventricular pressure in mouse embryos from E12.5 to E16.5 (embryonic stages in days). Ex utero measurements revealed intraventricular pressure increasing from 53.76 ± 4.16 Pa at E12.5 to 158.10 ± 19.94 Pa by E16.5. In utero analyses uncovered striking periodicity in sync with uterine contractions, reaching up to 1430 ± 195.2 Pa at E12.5, indicating dynamic mechanical stimuli beyond ex utero observations. Additionally, perforation experiments at E9.0–E15.5 showed rapid neuroepithelial thickening and apical surface contraction upon pressure release, indicative of a tensile effect by the positive intraventricular pressure. This effect diminished after E15.5, implying that tension wanes or the neuroepithelium becomes more robust. These results highlight the dynamic nature of embryonic intraventricular pressure, governed by internal fluid production and uterine forces, and emphasize the importance of mechanical cues in neuroepithelial architecture. Our findings provide a steppingstone to clarify how mechanical forces integrate with genetic and molecular processes to shape normal brain development and may render new perspectives on brain evolution.

Enteroendocrine cells (EECs) are sensory epithelial cells that sense the gut luminal environment and convey sensory information to the brain via the visceral afferent pathway. Although EECs are a part of gut epithelial cells, which generally undergo rapid turnover, some EECs have been reported to be long-lived. EECs consist of multiple subtypes, each of which displays distinct hormone production and distribution patterns. It remains unknown whether a long lifespan is a characteristic shared by all EEC subtypes. To address this issue, we conducted genetic pulse labeling of three EEC subtypes expressing serotonin (5-HT), peptide YY (PYY), and gastric inhibitory polypeptide (GIP) in mice and tracked their survival. In the proximal small intestine, all labeled GIP⁺ EECs disappeared completely within 5 days, whereas some PYY⁺ EECs survived for more than 7 days. In the proximal colon, some labeled 5-HT⁺ EECs lived for more than 28 days, whereas no PYY⁺ cells survived beyond 14 days. These long-lived 5-HT⁺ EECs were almost exclusively found in the upper half of the crypt in the mucosal fold, where visceral sensory fibers were enriched. This study reveals subtype- and region-dependent survival of EECs and suggests that EEC–nerve communication may underlie the long lifespan of certain EECs.

The 18th International Zebrafish Conference “IZFC2024” was held in Kyoto from August 17 to 21, 2024.

The genetic modification of rats is a key technology for advancing biomedical research on human diseases. CRISPR/Cas9-mediated genome editing enables the generation of knockout rats in a single step, without the need for embryonic stem cells, by directly injecting genome editing components into zygotes. This simplifies the process, reduces costs, and accelerates gene function analysis in rats. However, the insertion of a gene cassette into a target site has remained inefficient, limiting the generation of knockin (KI) rats. To overcome this issue, we developed an optimized method that covers the entire process from zygote harvesting with superovulation to timed microinjection, ensuring the consistent generation of KI rats. We successfully generated four different fluorescent reporter lines at the ROSA26 locus in rats. Our study provides detailed, step-by-step protocols for donor vector design, zygote collection, microinjection, founder screening, and cryopreservation in rats.

The first Asian community conference on “Scientific research using Xenopus as an animal model system” was held in Osaka, Japan, from November 24 to 26, 2024. This event was organized by the Korean Society for Xenopus Development of KSMCB (Korean Society for the Molecular and Cellular Biology), Xenopus Community in Japan (XCIJ), KNU G-LAMP Project, National BioResource Project—Clawed frogs/Newts (NBRP), and Osaka University. The conference covered various research topics in biological sciences using Xenopus. The ultimate goal of this conference was to promote collaboration and encourage a new and sustainable relationship among researchers in the Asian region who utilize Xenopus in their studies by sharing scientific findings. The conference not only shared research findings from various biological fields in the Asian region but also provided opportunities to invite global experts in the Xenopus research for “Keynote Lectures” and rising researchers who use other animal models for the “Friends of Xenopus” session to foster interdisciplinary exchange. In addition, organizers planned a luncheon seminar to help trainee students learn how to write a scientific paper and how not to.