Differential gene expression analysis identified determinants of cell fate plasticity during radiation-induced regeneration in Drosophila.

Ionizing radiation (IR) is used to treat half of all cancer patients because of its ability to kill cells. IR, however, can induce stem cell-like properties in non-stem cancer cells, potentiating tumor regrowth and reduced therapeutic success. We identified previously a subpopulation of cells in Drosophila larval wing discs that exhibit IR-induced stem cell-like properties. These cells reside in the future wing hinge, are resistant to IR-induced apoptosis, and are capable of translocating, changing fate, and participating in regenerating the pouch that suffers more IR-induced apoptosis. We used here a combination of lineage tracing, FACS-sorting of cells that change fate, genome-wide RNAseq, and functional testing of 42 genes, to identify two key changes that are required cell-autonomously for IR-induced hinge-to-pouch fate change: (1) repression of hinge determinants Wg (Drosophila Wnt1) and conserved zinc-finger transcription factor Zfh2 and (2) upregulation of three ribosome biogenesis factors. Additional data indicate a role for Myc, a transcriptional activator of ribosome biogenesis genes, in the process. These results provide a molecular understanding of IR-induced cell fate plasticity that may be leveraged to improve radiation therapy.

Rapid and robust optogenetic control of gene expression in Drosophila.

Deciphering gene function requires the ability to control gene expression in space and time. Binary systems such as the Gal4/UAS provide a powerful means to modulate gene expression and to induce loss or gain of function. This is best exemplified in Drosophila, where the Gal4/UAS system has been critical to discover conserved mechanisms in development, physiology, neurobiology, and metabolism, to cite a few. Here we describe a transgenic light-inducible Gal4/UAS system (ShineGal4/UAS) based on Magnet photoswitches. We show that it allows efficient, rapid, and robust activation of UAS-driven transgenes in different tissues and at various developmental stages in Drosophila. Furthermore, we illustrate how ShineGal4 enables the generation of gain and loss-of-function phenotypes at animal, organ, and cellular levels. Thanks to the large repertoire of UAS-driven transgenes, ShineGal4 enriches the Drosophila genetic toolkit by allowing in vivo control of gene expression with high temporal and spatial resolutions.

Self-organization of human dorsal-ventral forebrain structures by light induced SHH.

Organizing centers secrete morphogens that specify the emergence of germ layers and the establishment of the body's axes during embryogenesis. While traditional experimental embryology tools have been instrumental in dissecting the molecular aspects of organizers in model systems, they are impractical in human in-vitro model systems to dissect the relationships between signaling and fate along embryonic coordinates. To systematically study human embryonic organizer centers, we devised a collection of optogenetic ePiggyBac vectors to express a photoactivatable Cre-loxP recombinase, that allows the systematic induction of organizer structures by shining blue-light on human embryonic stem cells (hESCs). We used a light stimulus to geometrically confine SHH expression in neuralizing hESCs. This led to the self-organization of mediolateral neural patterns. scRNA-seq analysis established that these structures represent the dorsal-ventral forebrain, at the end of the first month of development. Here, we show that morphogen light-stimulation is a scalable tool that induces self-organizing centers.

Exposure to Pulsed Electromagnetic Fields Improves the Developmental Competence and Quality of Somatic Cell Nuclear Transfer Buffalo (Bubalus bubalis) Embryos Produced Using Fibroblast Cells and Alters Their Epigenetic Status and Gene Expression.

We examined the effects of treatment with pulsed electromagnetic fields (PEMFs) on cumulus cells and buffalo somatic cell nuclear transfer (SCNT) embryos. PEMF treatment (30 µT for 3 hours) of cumulus cells increased (p < 0.05) the relative cell viability and cell proliferation and the expression level of OCT4, NANOG, SOX2, P53, CCNB1, and GPX, but decreased (p < 0.05) that of DNMT1, DNMT3a, GSK3b, and BAX, whereas the expression level of DNMT3b, GLUT1, BCL2, CASPASE3, SOD1, and CATALASE was not affected. PEMF treatment of SCNT embryos at the beginning of in vitro culture increased (p < 0.05) the blastocyst rate (51.4% ± 1.36% vs. 42.8% ± 1.29%) and decreased (p < 0.01) the apoptotic index to the level in in vitro fertilization blastocysts, but did not significantly alter the total cell number and the inner cell mass:trophectoderm cell number ratio of blastocysts compared to the controls. PEMF treatment increased the expression level of NANOG, SOX2, CDX2, GLUT1, P53, and BCL2 and decreased that of BAX, CASPASE3, GSK3b, and HSP70, but not OCT4, DNMT1, DNMT3a, DNMT3b, HDAC1, and CCNB1 in blastocysts. It increased (p < 0.001) the global level of H3K27me3 but not H3K18ac. These results suggest that PEMF treatment of SCNT embryos improves their developmental competence, reduces the level of apoptosis, and alters the expression level of several important genes related to pluripotency, apoptosis, metabolism, and stress.

Effect of magnetic graphene oxide on cellular behaviors and osteogenesis under a moderate static magnetic field.

The biological behaviors of magnetic graphene oxide (MGO) in a static magnetic field (SMF) are unknown. The current study is to investigate the cellular behaviors, osteogenesis and the mechanism in BMSCs treated with MGO combined with an SMF. Results showed that the synthetic MGO particles were bio-compatible and could significantly improve the osteogenesis of BMSCs under SMFs, as verified by elevated alkaline phosphatase activity, mineralized nodule formation, and expressions of mRNA and protein levels. Under SMF at the same intensity, the addition of graphene oxide to Fe3O4 could increase the osteogenic ability of BMSCs. The Wnt/ß-catenin pathway was indicated to be related to the MGO-driven osteogenic behavior of the BMSCs under SMF. Taken together, our findings suggested that MGO under an SMF could promote osteogenesis in BMSCs through the Wnt/ß-catenin pathway and hence should attract more attention for practical applications in bone tissue regeneration.

Cryptic developmental events determine medulloblastoma radiosensitivity and cellular heterogeneity without altering transcriptomic profile.

It is unclear why medulloblastoma patients receiving similar treatments experience different outcomes. Transcriptomic profiling identified subgroups with different prognoses, but in each subgroup, individuals remain at risk of incurable recurrence. To investigate why similar-appearing tumors produce variable outcomes, we analyzed medulloblastomas triggered in transgenic mice by a common driver mutation expressed at different points in brain development. We genetically engineered mice to express oncogenic SmoM2, starting in multipotent glio-neuronal stem cells, or committed neural progenitors. Both groups developed medulloblastomas with similar transcriptomic profiles. We compared medulloblastoma progression, radiosensitivity, and cellular heterogeneity, determined by single-cell transcriptomic analysis (scRNA-seq). Stem cell-triggered medulloblastomas progressed faster, contained more OLIG2-expressing stem-like cells, and consistently showed radioresistance. In contrast, progenitor-triggered MBs progressed slower, down-regulated stem-like cells and were curable with radiation. Progenitor-triggered medulloblastomas also contained more diverse stromal populations, with more Ccr2+ macrophages and fewer Igf1+ microglia, indicating that developmental events affected the subsequent tumor microenvironment. Reduced mTORC1 activity in M-Smo tumors suggests that differential Igf1 contributed to differences in phenotype. Developmental events in tumorigenesis that were obscure in transcriptomic profiles thus remained cryptic determinants of tumor composition and outcome. Precise understanding of medulloblastoma pathogenesis and prognosis requires supplementing transcriptomic/methylomic studies with analyses that resolve cellular heterogeneity.

Crocetin Mitigates Irradiation Injury in an In Vitro Model of the Pubertal Testis: Focus on Biological Effects and Molecular Mechanisms.

Infertility is a potential side effect of radiotherapy and significantly affects the quality of life for adolescent cancer survivors. Very few studies have addressed in pubertal models the mechanistic events that could be targeted to provide protection from gonadotoxicity and data on potential radioprotective treatments in this peculiar period of life are elusive. In this study, we utilized an in vitro model of the mouse pubertal testis to investigate the efficacy of crocetin to counteract ionizing radiation (IR)-induced injury and potential underlying mechanisms. Present experiments provide evidence that exposure of testis fragments from pubertal mice to 2 Gy X-rays induced extensive structural and cellular damage associated with overexpression of PARP1, PCNA, SOD2 and HuR and decreased levels of SIRT1 and catalase. A twenty-four hr exposure to 50 µM crocetin pre- and post-IR significantly reduced testis injury and modulated the response to DNA damage and oxidative stress. Nevertheless, crocetin treatment did not counteract the radiation-induced changes in the expression of SIRT1, p62 and LC3II. These results increase the knowledge of mechanisms underlying radiation damage in pubertal testis and establish the use of crocetin as a fertoprotective agent against IR deleterious effects in pubertal period.

PUMA facilitates EMI1-promoted cytoplasmic Rad51 ubiquitination and inhibits DNA repair in stem and progenitor cells.

Maintenance of genetic stability via proper DNA repair in stem and progenitor cells is essential for the tissue repair and regeneration, while preventing cell transformation after damage. Loss of PUMA dramatically increases the survival of mice after exposure to a lethal dose of ionizing radiation (IR), while without promoting tumorigenesis in the long-term survivors. This finding suggests that PUMA (p53 upregulated modulator of apoptosis) may have a function other than regulates apoptosis. Here, we identify a novel role of PUMA in regulation of DNA repair in embryonic or induced pluripotent stem cells (PSCs) and immortalized hematopoietic progenitor cells (HPCs) after IR. We found that PUMA-deficient PSCs and HPCs exhibited a significant higher double-strand break (DSB) DNA repair activity via Rad51-mediated homologous recombination (HR). This is because PUMA can be associated with early mitotic inhibitor 1 (EMI1) and Rad51 in the cytoplasm to facilitate EMI1-mediated cytoplasmic Rad51 ubiquitination and degradation, thereby inhibiting Rad51 nuclear translocation and HR DNA repair. Our data demonstrate that PUMA acts as a repressor for DSB DNA repair and thus offers a new rationale for therapeutic targeting of PUMA in regenerative cells in the context of DNA damage.

Light-induced and circadian expressions of melanopsin genes opn4xa and opn4xb in the eyes of juvenile grass puffer Takifugu alboplumbeus.

Animals regulate a variety of aspects of physiology according to environmental light conditions via nonvisual opsins such as melanopsin. In order to study photic regulation of fish physiology, expression changes of the genes for melanopsin (opn4xa and opn4xb) and effects of light on them were examined in juvenile grass puffer Takifugu alboplumbeus using quantitative real-time PCR. In the brain of juvenile fish, no significant diurnal nor circadian changes were observed in opn4x mRNA levels. On the other hand, in the eyes, the mRNA level of opn4xa showed a significant diurnal rhythm with a peak at Zeitgeber time (ZT) 4, while no apparent circadian changes were observed. The mRNA level of opn4xb in the eyes showed a diurnal change similar to that of opn4xa, while it showed a significant circadian change. Furthermore, continuous exposure to light during a subjective night significantly increased the mRNA levels of opn4xa in the eyes at ZT24, suggesting that light induces gene expression of opn4xa in the eyes and that the induction occurs only during the night-day transition period. These results suggest that Opn4xa and Opn4xb play differential roles in the eyes of juvenile grass puffer to mediate the physiological effects of environmental light information.

IAA3-mediated repression of PIF proteins coordinates light and auxin signaling in Arabidopsis.

The exogenous light signal and endogenous auxin are two critical factors that antagonistically regulate hypocotyl growth. However, the regulatory mechanisms integrating light and auxin signaling pathways need further investigation. In this study, we identified a direct link between the light and auxin signaling pathways mediated by the auxin transcriptional repressor IAA3 and light-controlled PIF transcription factors in Arabidopsis. The gain-of-function mutation in IAA3 caused hyposensitivity to light, whereas disruption of IAA3 led to an elongated hypocotyl under different light intensity conditions, indicating that IAA3 is required in light regulated hypocotyl growth. Genetic studies showed that the function of IAA3 in hypocotyl elongation is dependent on PIFs. Our data further demonstrated that IAA3 interacts with PIFs in vitro and in vivo, and it attenuates the DNA binding activities of PIFs to the target genes. Moreover, IAA3 negatively regulates the expression of PIFs-dependent genes. Collectively, our study reveals an interplay mechanism of light and auxin on the regulation of hypocotyl growth, coordinated by the IAA3 and PIFs transcriptional regulatory module.

TAEL 2.0: An Improved Optogenetic Expression System for Zebrafish.

Inducible gene expression systems are valuable tools for studying biological processes. We previously developed an optogenetic gene expression system called TAEL that is optimized for use in zebrafish. When illuminated with blue light, TAEL transcription factors dimerize and activate gene expression downstream of the TAEL-responsive C120 promoter. By using light as the inducing agent, the TAEL/C120 system overcomes limitations of traditional inducible expression systems by enabling fine spatial and temporal regulation of gene expression. In this study, we describe ongoing efforts to improve the TAEL/C120 system. We made modifications to both the TAEL transcriptional activator and the C120 regulatory element, collectively referred to as TAEL 2.0. We demonstrate that TAEL 2.0 consistently induces higher levels of reporter gene expression and at a faster rate, but with comparable background and toxicity as the original TAEL system. With these improvements, we were able to create functional stable transgenic lines to express the TAEL 2.0 transcription factor either ubiquitously or with a tissue-specific promoter. We demonstrate that the ubiquitous line in particular can be used to induce expression at late embryonic and larval stages, addressing a major deficiency of the original TAEL system. This improved optogenetic expression system will be a broadly useful resource for the zebrafish community.

Intestinal long non-coding RNAs in response to simulated microgravity stress in Caenorhabditis elegans.

Long non-coding RNAs (lncRNAs) are important in regulating the response to environmental stresses in organisms. In this study, we used Caenorhabditis elegans as an animal model to determine the functions of intestinal lncRNAs in regulating response to simulated microgravity stress. Among the intestinal lncRNAs, linc-2, linc-46, linc-61, and linc-78 were increased by simulated microgravity treatment, and linc-13, linc-14, linc-50, and linc-125 were decreased by simulated microgravity treatment. Among these 8 intestinal lncRNAs, RNAi knockdown of linc-2 or linc-61 induced a susceptibility to toxicity of simulated microgravity, whereas RNAi knockdown of linc-13, linc-14, or linc-50 induced a resistance to toxicity of simulated microgravity. In simulated microgravity treated nematodes, linc-50 potentially binds to three transcriptional factors (DAF-16, SKN-1, and HLH-30). RNAi knockdown of daf-16, skn-1, or hlh-30 could suppress resistance of linc-50(RNAi) nematodes to the toxicity of simulated microgravity. Therefore, our results provide an important basis for intestinal lncRNAs, such as the linc-50, in regulating the response to simulated microgravity in nematodes.

Ionising Radiation Induces Promoter DNA Hypomethylation and Perturbs Transcriptional Activity of Genes Involved in Morphogenesis during Gastrulation in Zebrafish.

Embryonic development is particularly vulnerable to stress and DNA damage, as mutations can accumulate through cell proliferation in a wide number of cells and organs. However, the biological effects of chronic exposure to ionising radiation (IR) at low and moderate dose rates (< 6 mGy/h) remain largely controversial, raising concerns for environmental protection. The present study focuses on the molecular effects of IR (0.005 to 50 mGy/h) on zebrafish embryos at the gastrula stage (6 hpf), at both the transcriptomics and epigenetics levels. Our results show that exposure to IR modifies the expression of genes involved in mitochondrial activity from 0.5 to 50 mGy/h. In addition, important developmental pathways, namely, the Notch, retinoic acid, BMP and Wnt signalling pathways, were altered at 5 and 50 mGy/h. Transcriptional changes of genes involved in the morphogenesis of the ectoderm and mesoderm were detected at all dose rates, but were prominent from 0.5 to 50 mGy/h. At the epigenetic level, exposure to IR induced a hypomethylation of DNA in the promoter of genes that colocalised with both H3K27me3 and H3Kme4 histone marks and correlated with changes in transcriptional activity. Finally, pathway enrichment analysis demonstrated that the DNA methylation changes occurred in the promoter of important developmental genes, including morphogenesis of the ectoderm and mesoderm. Together, these results show that the transcriptional program regulating morphogenesis in gastrulating embryos was modified at dose rates greater than or equal to 0.5 mGy/h, which might predict potential neurogenesis and somitogenesis defects observed at similar dose rates later in development.

stim2b Knockout Induces Hyperactivity and Susceptibility to Seizures in Zebrafish Larvae.

In neurons, stromal interaction molecule (STIM) proteins regulate store-operated Ca2+ entry (SOCE) and are involved in calcium signaling pathways. However, STIM activity in neurological diseases is unclear and should be clarified by studies that are performed in vivo rather than in cultured cells in vitro. The present study investigated the role of neuronal Stim2b protein in zebrafish. We generated stim2b knockout zebrafish, which were fertile and had a regular lifespan. Using various behavioral tests, we found that stim2b-/- zebrafish larvae were hyperactive compared with wild-type fish. The mutants exhibited increases in mobility and thigmotaxis and disruptions of phototaxis. They were also more sensitive to pentylenetetrazol and glutamate treatments. Using lightsheet microscopy, a higher average oscillation frequency and higher average amplitude of neuronal Ca2+ oscillations were observed in stim2b-/- larvae. RNA sequencing detected upregulation of the annexin 3a and gpr39 genes and downregulation of the rrm2, neuroguidin, and homer2 genes. The latter gene encodes a protein that is involved in several processes that are involved in Ca2+ homeostasis in neurons, including metabotropic glutamate receptors. We propose that Stim2b deficiency in neurons dysregulates SOCE and triggers changes in gene expression, thereby causing abnormal behavior, such as hyperactivity and susceptibility to seizures.

Effects of Low-Dose Gamma-Ray Radiation on Apoptosis and Development of Zebrafish Embryo Brain.

To investigate the effects of low-dose γ irradiation on apoptosis and development of the brain in zebrafish embryos, cumulative 15 mGy doses of γ rays from a 137Cs source were used to irradiate zebrafish embryos at 2 h post-fertilization (hpf) for 120 h. Apoptosis of the brain, brain morphological development, cell submicroscopic structure and mRNA expression were analyzed, respectively. Results indicate that after 15 mGy exposure, the apoptosis of zebrafish brain increased, vacuoles appeared in brain tissue, some organelles were damaged and vacuoles appeared locally in brain cells. The mRNA expression level of axin2 was significantly upregulated, and those of frizzled, ß-catenin, camk2, TCF/ LEF and bcl9 were significantly downregulated in brain tissue. These genes are involved in the Wnt signaling pathway. The findings of this work suggest that low-dose radiation may influence the apoptosis and development of the brain in the zebrafish embryo by inhibiting the Wnt signaling pathway.

Novel in vitro assay to investigate radiation induced changes in the functionality of human embryonic stem cell-derived neurospheres.

Ionizing radiation (IR) is increasingly used for diagnostics and therapy of severe brain diseases. However, IR also has adverse effects on the healthy brain tissue, particularly on the neuronal network. This is true for adults but even more pronounced in the developing brain of unborn and pediatric patients. Epidemiological studies on children receiving radiotherapy showed an increased risk for cognitive decline ranging from mild deficits in academic functioning to severe late effects in intellectual ability and language as a consequence of altered neuronal development and connectivity. To provide a comprehensive approach for the analysis of radiation-induced alterations in human neuronal functionality, we developed an in vitro assay by combining microelectrode array (MEA) analyses and human embryonic stem cell (hESC) derived three-dimensional neurospheres (NS). In our proof of principle study, we irradiated hESC with 1 Gy X-rays and let them spontaneously differentiate into neurons within NS. After the onset of neuronal activity, we recorded and analyzed the activity pattern of the developing neuronal networks. The network activity in NS derived from irradiated hESC was significantly reduced, whereas no differences in molecular endpoints such as cell proliferation and transcript or protein expression analyses were found. Thus, the combination of MEA analysis with a 3D model for neuronal functionality revealed radiation sequela that otherwise would not have been detected. We therefore strongly suggest combining traditional biomolecular methods with the new functional assay presented in this work to improve the risk assessment for IR-induced effects on the developing brain.

Irradiation-induced senescence of bone marrow mesenchymal stem cells aggravates osteogenic differentiation dysfunction via paracrine signaling.

The role of cellular senescence induced by radiation in bone loss has attracted much attention. As one of the common complications of anticancer radiotherapy, irradiation-induced bone deterioration is common and clinically significant, but the pathological mechanism has not been elucidated. This study was performed to explore the cellular senescence and senescence-associated secretory phenotype (SASP) induction of bone marrow-derived mesenchymal stem cells (BMSCs) by irradiation and its role in osteogenic differentiation dysfunction. It was observed that irradiated BMSCs lost typical fibroblast-like morphology, exhibited suppressed viability and differentiation potential accompanied with senescence phenotypes, including an increase in senescence-associated ß-galactosidase (SA-ß-gal) staining-positive cells, and upregulated senescence-related genes p53/p21, whereas no changes happened to p16. Additionally, DNA damage γ-H2AX foci, G0/G1 phase of cell cycle arrest, and cellular and mitochondrial reactive oxygen species (ROS) increased in an irradiation dose-dependent manner. Meanwhile, the JAK1/STAT3 pathway was activated and accompanied by an increase in SASP secretion, such as IL-6, IL-8, and matrix metalloproteinase-9 (MMP9), whereas 0.8 µM JAK1 inhibitor (JAKi) treatment effectively inhibited the JAK pathway and SASP production. Furthermore, conditioned medium (CM) from irradiation-induced senescent (IRIS) BMSCs exhibited a markedly reduced ability in osteogenic differentiation and marker gene expression of osteoblasts, whereas CM with JAKi intervention may effectively improve these deterioration effects. In conclusion, irradiation could provoke BMSC senescence and SASP secretion and further aggravate osteogenic differentiation dysfunction via paracrine signaling, whereas SASP targeting may be a possible intervention strategy for alleviating irradiation-induced bone loss.

Genome-wide identification of Arabidopsis long noncoding RNAs in response to the blue light.

Long non-coding RNAs (lncRNAs) have been shown in animals to play roles in a wide range of biological processes. In plant, light modulates the growth and development as a key external signal. However, little is known about the role of plant lncRNA in response to light. In this study, we sequenced the messenger RNAs (mRNAs), lncRNAs and microRNAs (miRNAs) in Arabidopsis seedlings under blue light for 2 h and 8 h. Compared to dark, we identified 4197 mRNAs, 375 miRNAs and 481 lncRNAs, or 5207 mRNAs, 286 miRNAs and 545 lncRNAs of differential expressions under blue light treatments for 2 h or 8 h respectively. Subsequently, a total of 407 competing endogenous RNA (ceRNA) pairs (lncRNA-mRNA-miRNA) were constructed. We identified a blue light-induced lncRNA which plays roles in blue light-directed plant photomorphogenesis and response to mannitol stress by serving as a ceRNA to sequester miR167 in a type of target mimicry. These results revealed previously unknown roles of the lncRNA in blue light signaling and mannitol stress, and provided useful resources of lncRNAs associated with miRNAs in response to blue light.

Quantitative Proteomic Analyses Identify STO/BBX24 -Related Proteins Induced by UV-B.

Plants use solar radiation for photosynthesis and are inevitably exposed to UV-B. To adapt to UV-B radiation, plants have evolved a sophisticated strategy, but the mechanism is not well understood. We have previously reported that STO (salt tolerance)/BBX24 is a negative regulator of UV-B-induced photomorphogenesis. However, there is limited knowledge of the regulatory network of STO in UV-B signaling. Here, we report the identification of proteins differentially expressed in the wild type (WT) and sto mutant after UV-B radiation by iTRAQ (isobaric tags for relative and absolute quantitation)-based proteomic analysis to explore differential proteins that depend on STO and UV-B signaling. A total of 8212 proteins were successfully identified, 221 of them were STO-dependent proteins in UV-B irradiated plants. The abundances of STO-dependent PSB and LHC (light-harvesting complex) proteins in sto mutants decreased under UV-B radiation, suggesting that STO is necessary to maintain the normal accumulation of photosynthetic system complex under UV-B radiation to facilitate photosynthesis photon capture. The abundance of phenylalanine lyase-1 (PAL1), chalcone synthetase (CHS), and flavonoid synthetase (FLS) increased significantly after UV-B irradiation, suggesting that the accumulation of flavonoids do not require STO, but UV-B is needed. Under UV-B radiation, STO stabilizes the structure of antenna protein complex by maintaining the accumulation of PSBs and LHCs, thereby enhancing the non-photochemical quenching (NPQ) ability, releasing extra energy, protecting photosynthesis, and ultimately promoting the elongation of hypocotyl. The accumulation of flavonoid synthesis key proteins is independent of STO under UV-B radiation. Overall, our results provide a comprehensive regulatory network of STO in UV-B signaling.

Proteomic Analysis of Irradiation with Millimeter Waves on Soybean Growth under Flooding Conditions.

Improving soybean growth and tolerance under environmental stress is crucial for sustainable development. Millimeter waves are a radio-frequency band with a wavelength range of 1-10 mm that has dynamic effects on organisms. To investigate the potential effects of millimeter-waves irradiation on soybean seedlings, morphological and proteomic analyses were performed. Millimeter-waves irradiation improved the growth of roots/hypocotyl and the tolerance of soybean to flooding stress. Proteomic analysis indicated that the irradiated soybean seedlings recovered under oxidative stress during growth, whereas proteins related to glycolysis and ascorbate/glutathione metabolism were not affected. Immunoblot analysis confirmed the promotive effect of millimeter waves to glycolysis- and redox-related pathways under flooding conditions. Sugar metabolism was suppressed under flooding in unirradiated soybean seedlings, whereas it was activated in the irradiated ones, especially trehalose synthesis. These results suggest that millimeter-waves irradiation on soybean seeds promotes the recovery of soybean seedlings under oxidative stress, which positively regulates soybean growth through the regulation of glycolysis and redox related pathways.