Kihara, S. | bioRxiv Science

bioRxiv2025

Expression dynamics and functional characterization of the pigmented and non-diapausing egg gene (pnd) and pnd-2, responsible for the initiation of embryonic diapause in the silkworm, Bombyx mori

Diapause in insects is a developmental arrest that serves as an adaptation to severe environmental conditions. In the silkworm Bombyx mori, embryonic diapause in the next generation is maternally induced by the diapause hormone (DH). However, the downstream factors responsible for initiating diapause have remained unidentified. The pnd and pnd-2 mutants produce pigmented and non-diapausing eggs despite intact DH signaling, indicating that the pnd and pnd-2 genes act downstream as potential initiators of diapause. Here, we analyzed their expression dynamics and functions. Expression analyses showed that pnd was strongly upregulated in diapause-destined eggs in a DH-signal-dependent manner and was suppressed by HCl treatment that prevents diapause initiation. In contrast, pnd-2 exhibited a transient expression peak independent of DH-signal and was unaffected by HCl treatment. RNAi-mediated knockdown of either gene inhibited the diapause initiation, and injection of pnd mRNA into non-diapause-destined eggs induced a diapause-like state in embryos. These findings demonstrate that pnd and pnd-2 are required downstream of the DH-signal to initiate diapause, providing key insights into the molecular mechanism underlying embryonic diapause induction in B. mori. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=128 SRC="FIGDIR/small/667111v1_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@7e9516org.highwire.dtl.DTLVardef@6dddc2org.highwire.dtl.DTLVardef@fa11baorg.highwire.dtl.DTLVardef@1a2054e_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2019

Modeling the Human Segmentation Clock with Pluripotent Stem Cells

Pluripotent stem cells (PSCs) have increasingly been used to model different aspects of embryogenesis and organ formation1. Despite recent advances in the in vitro induction of major mesodermal lineages and mesoderm-derived cell types2,3, experimental model systems that can recapitulate more complex biological features of human mesoderm development and patterning are largely missing. Here, we utilized induced pluripotent stem cells (iPSCs) for the stepwise in vitro induction of presomitic mesoderm (PSM) and its derivatives to model distinct aspects of human somitogenesis. We focused initially on modeling the human segmentation clock, a major biological concept believed to underlie the rhythmic and controlled emergence of somites, which give rise to the segmental pattern of the vertebrate axial skeleton. We succeeded to observe oscillatory expression of core segmentation clock genes, including HES7 and DKK1, and identified novel oscillatory genes in human iPSC-derived PSM. We furthermore determined the period of the human segmentation clock to be around five hours and showed the presence of dynamic traveling wave-like gene expression within in vitro induced human PSM. Utilizing CRISPR/Cas9-based genome editing technology, we then targeted genes, for which mutations in patients with abnormal axial skeletal development such as spondylocostal dysostosis (SCD) (HES7, LFNG and DLL3) or spondylothoracic dysostosis (STD) (MESP2) have been reported. Subsequent analysis of patient-like iPSC knock-out lines as well as patient-derived iPSCs together with their genetically corrected isogenic controls revealed gene-specific alterations in oscillation, synchronization or differentiation properties, validating the overall utility of our model system, to recapitulate not only key features of human somitogenesis but also to provide novel insights into diseases associated with the formation and patterning of the human axial skeleton.

developmental biology↗