Bone and periosteum protein analysis via tandem mass tag quantitative proteomics in pediatric patients with osteomyelitis.

Bone healing is crucial in managing osteomyelitis after fracture fixation. Understanding the mechanism of extensive callus formation in pediatric osteomyelitis is highly important. This study aims to analyze bone and periosteum samples from pediatric patients to elucidate the essential processes involved in callus formation during osteomyelitis. The study included eight patients from our hospital: four with positive microbial culture who underwent osteomyelitis debridement and four who had osteotomy surgery as contral. We used tandem mass tag quantitative proteomics to investigate proteomic changes in bone and periosteum tissues obtained from these patients. Differential expression proteins were analyzed for their pathways through Gene Ontology (GO) annotation, GO enrichment analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, and protein-protein interaction networks. A total of 4737 proteins were successfully identified. About 2224 differentially expressed proteins were detected in the bone tissues group and periosteum tissues group. Among the differentially expressed proteins, 10 protein genes in the bone group were associated with inflammation and osteogenesis, while in the periosteum group were nine. Cytochrome b-245, beta polypeptide (CYBB), nicotinamide phosphoribosyltransferase (NAMPT), tissue inhibitor of metalloproteinases 1 (TIMP-1), Raf-1 proto-oncogene, serine/threonine kinase (RAF-1), RELA proto-oncogene, NF-KB subunit (RELA), and sphingomyelin synthase 2 (SGMS2) may play an important role in callus formation in patients with osteomyelitis. This study provides novel clues for understanding callus formation in pediatric patients with osteomyelitis.

Novel mechanism of MicroRNA408 in callus formation from rice mature embryo.

Mature embryos are the main explants of tissue culture used in rice transgenic technology. However, the mechanism of mature embryo callus formation remains unclear. In this study, a microRNA-mediated gene regulatory network of rice calli was established using degradome sequencing. We identified a microRNA, OsmiR408, that regulates the formation of the callus derived from the mature rice embryo. OsUCLACYANIN 30 (OsUCL 30), a target gene of OsmiR408, was the most abundant cleavage mRNA in rice callus. OsUCL17 was verified as a target gene of OsmiR408 using RNA ligase-mediated 5'-RACE. In analysis of the OsmiR408 promoter reporter line and pri-miR408 transcript level, the promoter activity and transcript level of MIR408 were increased dramatically during callus formation. In phenotypic observations, OsmiR408 knockout caused severe defects in mature embryo callus formation, whereas OsmiR408 overexpression promoted callus formation. Transcriptome analysis demonstrated that OsUCLs and certain genes related to the plant hormone signal transduction and phenylpropanoid-flavonoid biosynthesis pathway had different differential expression patterns between OsmiR408 knockout and overexpression calli. Thus, OsmiR408 may regulate callus formation mainly by affecting plant hormone signal transduction and phenylpropanoid-flavonoid biosynthesis pathway. Our findings provide insight into OsmiR408/UCLs module function in callus formation.

Nicotinamide mononucleotide enhances fracture healing by promoting skeletal stem cell proliferation.

The process of skeletal regeneration initiated by stem cells following injury, especially in fractures, is significantly impaired by aging and adverse factors. Nicotinamide mononucleotide (NMN), a critical endogenous precursor of nicotinamide adenine dinucleotide (NAD), has garnered extensive attention for its multifaceted regulatory functions in living organisms and its wide-ranging therapeutic potential. However, whether NMN contributes to trauma-induced skeletal regeneration remains unclear. Methods: The transverse femoral shaft fracture model was employed to evaluate the potential advantages of NMN administration for overall repair during the initial fracture stages in male mice through micro-CT analysis, histochemistry, and biomechanical testing. The pro-proliferative function of NMN on skeletal stem cells (SSCs) was investigated through flow cytometry, qRT-PCR, NAD content measurement, and cell proliferation assay. Results: In this study, we observed that the administration of NMN during the initial phase of fracture in mice led to a larger callus and corresponding improvement in micro-CT parameters. NMN enhances the cartilaginous component of the callus by elevating the NAD content, consequently accelerating subsequent endochondral ossification and the fracture healing process. Subsequent analyses elucidated that NMN was beneficial in promoting the expansion of diverse stem cells in vivo and in vitro potentially via modulation of the Notch signaling pathway. Moreover, the depletion of macrophages profoundly obstructs the proliferation of SSCs. Conclusion: Our discoveries provide a potential strategy for enhancing fracture healing through stimulation of callus SSC proliferation at an early stage, shedding light on the translational value of NMN as an enhancer for skeletal regeneration and highlighting the pivotal role of macrophage-stem cell interactions in governing the regenerative influence of NMN on stem cells.

A point mutation in the IAA14 promoter enhances callus formation and regeneration.

Callus formation induced by auxin accumulation is considered the first step of in vitro plant regeneration. In Arabidopsis, degradation of the Aux/IAA protein, IAA14, in response to auxin signaling, which activates the AUXIN RESPONSE FACTOR 7 (ARF7) and ARF19 along with a series of downstream transcription factors, also plays a critical role in this process. However, the specific mechanism by which auxin regulates callus formation remains unclear. By screening mutant library in the solitary root 1 (iaa14/slr) Arabidopsis background we obtained the callus formation related 2 (cfr2) mutant. The cfr2 mutant exhibited a stronger capacity for callus formation, as well as lateral root and adventitious root regeneration from leaf explants than wild type (WT) seedlings, but did not recover gravitropism capability. The auxin signal in cfr2 was significantly enhanced, and the expression of some downstream transcription factors was increased. Map-based cloning, whole genome resequencing, and phenotypic complementation experiments showed that the phenotypes observed in the cfr2 mutant were caused by a point mutation in the IAA14 promoter region. This mutation, which is predicted to disrupt the binding of LBD16, LBD19, and LBD30 to the IAA14 promoter, changed the expression pattern of IAA14 in cfr2. Taken together, our results identified a new mutation in the IAA14 promoter region, which affects the expression pattern of IAA14 and in turn its ability to control plant regeneration. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-024-01493-y.

Promoting bone callus formation by taking advantage of the time-dependent fracture gap strain modulation.

Delayed union and non-union of fractures continue to be a major problem in trauma and orthopedic surgery. These cases are challenging for the surgeon. In addition, these patients suffer from multiple surgeries, pain and disability. Furthermore, these cases are a major burden on healthcare systems. The scientific community widely agrees that the stability of fixation plays a crucial role in determining the outcome of osteosynthesis. The extent of stabilization affects factors like fracture gap strain and fluid flow, which, in turn, influence the regenerative processes positively or negatively. Nonetheless, a growing body of literature suggests that during the fracture healing process, there exists a critical time frame where intervention can stimulate the bone's return to its original form and function. This article provides a summary of existing evidence in the literature regarding the impact of different levels of fixation stability on the strain experienced by newly forming tissues. We will also discuss the timing and nature of this "window of opportunity" and explore how current knowledge is driving the development of new technologies with design enhancements rooted in mechanobiological principles.

Glucosinolate O-methyltransferase mediated callus formation and affected ROS homeostasis in Arabidopsis thaliana.

Auxin-induced callus formation was largely dependent on the function of Lateral Organ Boundaries Domain (LBD) family transcription factors. We previously revealed that two IGMT (Indole glucosinolate oxy-methyl transferase) genes, IGMT2 and IGMT3, may be involved in the callus formation process as potential target genes of LBD29. Overexpression of the IGMT genes induces spontaneous callus formation. However, the details of the IGMT involvement in callus formation process were not well studied. IGMT1-4, but not IGMT5, are targeted and induced by LBD29 during the early stage of callus formation. Cell membrane and nucleus localized IGMT3 was mainly expressed in the elongation and maturation zones tissues of the primary root and lateral root, which could be further accumulated after CIM treatment. The igmts quadruple mutant, which obtained by CRISPR/Cas9 technology, exhibits a phenotype of attenuated callus formation. Enhanced indole glucosinolate anabolic pathway caused by IGMT1-4 overexpression promotes callus formation. In addition, the IGMT genes were involved in the reactive oxygen species homeostasis, which could be responsible for its role on callus formation. This study provides novel insights into the role of IGMTs gene-mediated callus formation. Activation of the Indole glucosinolate anabolic pathway is an inducing factor for plant callus initiation. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01409-2.

Management of Complex Open Tibial Plateau Fracture: A Case Report on the Application of Locked Plate External Fixation Technique during Bone Callus Formation stage to Replace transarticular External Fixation.

Transarticular external fixation is primarily used for open fractures involving the joint. However, its biggest drawback is the potential forjoint dysfunction. The article reports a successful case with complex open tibial plateau fracture treated using locked plate external fixation technique during bone callus formation stage to replace transarticular external fixation. We present a case of a 55-year-old male who sustained a complex open fracture of the tibial plateau. In addition, he also suffered from multiple rib fractures, a fibula fracture, a clavicle fracture, hemorrhagic shock, and lung contusion. The patient has occurred tibial bone infection after undergoing open reduction and transarticular external fixation for fracture management. Our team skillfully applied locked plate external fixation technique during bone callus formation stage to replace transarticular external fixation. Ultimately, the approach not only successfully controls infection and achieves fracture healing but also preserves knee joint function after five years of follow-up. In conclusion,the application of locked plate external fixation technique during bone callus formation stage to replace transarticular external fixation is a valuable approach that orthopedic clinicians should consider and learn from when managing complex intra-articular fractures.

Unraveling Epigenetic Changes in A. thaliana Calli: Impact of HDAC Inhibitors.

The ability for plant regeneration from dedifferentiated cells opens up the possibility for molecular bioengineering to produce crops with desirable traits. Developmental and environmental signals that control cell totipotency are regulated by gene expression via dynamic chromatin remodeling. Using a mass spectrometry-based approach, we investigated epigenetic changes to the histone proteins during callus formation from roots and shoots of Arabidopsis thaliana seedlings. Increased levels of the histone H3.3 variant were found to be the major and most prominent feature of 20-day calli, associated with chromatin relaxation. The methylation status in root- and shoot-derived calli reached the same level during long-term propagation, whereas differences in acetylation levels provided a long-lasting imprint of root and shoot origin. On the other hand, epigenetic signs of origin completely disappeared during 20 days of calli propagation in the presence of histone deacetylase inhibitors (HDACi), sodium butyrate, and trichostatin A. Each HDACi affected the state of post-translational histone modifications in a specific manner; NaB-treated calli were epigenetically more similar to root-derived calli, and TSA-treated calli resembled shoot-derived calli.

The Healing Callus-Promoting Effect of Fenugreek in a Humerus Shaft Fracture: A Case Report.

A 54-year-old male presented with a fractured shaft in the right humerus and refused surgery. The patient was treated with a cast, and a follow-up plain radiography revealed good callus formation after 32 days. The patient had a history of receiving fenugreek seed extract from the first week after the fracture. We did our best to exclude any other factors that helped rapid fracture healing with good callus formation in our patient. The current case supports the hypothesis that fenugreek seed extract promotes bone healing. This hypothesis is supported by a literature review. Previous studies have suggested several mechanisms by which fenugreek promotes bone healing.

The HOS1-PIF4/5 module controls callus formation in Arabidopsis leaf explants.

A two-step plant regeneration has been widely exploited to genetic manipulation and genome engineering in plants. Despite technical importance, understanding of molecular mechanism underlying in vitro plant regeneration remains to be fully elucidated. Here, we found that the HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES 1 (HOS1)-PHYTOCHROME INTERACTING FACTOR 4/5 (PIF4/5) module participates in callus formation. Consistent with the repressive role of HOS1 in PIF transcriptional activation activity, hos1-3 mutant leaf explants exhibited enhanced callus formation, whereas pif4-101 pif5-3 mutant leaf explants showed reduced callus size. The HOS1-PIF4/5 function would be largely dependent on auxin biosynthesis and signaling, which are essential for callus initiation and proliferation. Our findings suggest that the HOS1-PIF4/5 module plays a pivotal role in auxin-dependent callus formation in Arabidopsis.

Three-Dimensional Computational Model Simulating the Initial Callus Growth during Fracture Healing in Long Bones: Application to Different Fracture Types.

Bone fractures are among the most common and potentially serious injuries to the skeleton, femoral shaft fractures being especially severe. Thanks to recent advances in the area of in silico analysis, several approximations of the bone healing process have been achieved. In this context, the objective of this work was to simulate the initial phase of callus formation in long bones, without a pre-meshed domain in the 3D space. A finite element approach was computationally implemented to obtain the values of the cell concentrations along the whole domain and evaluate the areas where the biological quantities reached the thresholds necessary to trigger callus growth. A voxel model was used to obtain the 3D domain of the bone fragments and callus. A mesh growth algorithm controlled the addition of new elements to the domain at each step of the iterative procedure until complete callus formation. The implemented approach is able to reproduce the generation of the primary callus, which corresponds to the initial phase of fracture healing, independently of the fracture type and complexity, even in the case of several bone fragments. The proposed approach can be applied to the most complex bone fractures such as oblique, severely comminuted or spiral-type fractures, whose simulation remains hardly possible by means of the different existing approaches available to date.

Closed reduction with percutaneous Kirschner wire drill-and-pry for pediatric supracondylar humeral fractures with bony callus formation and delayed presentation.

BACKGROUND: Supracondylar humeral fractures are the most common type of pediatric elbow fractures, and are primarily treated using closed reduction and percutaneous pinning. For patients who are treated ≥14 days after the injury, after callus formation has occurred, closed reduction is usually not possible. The purpose of this study is to report the clinical outcomes of closed reduction with percutaneous Kirschner wire (K-wire) drill-and-pry for the delayed treatment of pediatric supracondylar humeral fractures with bony callus formation. METHODS: We retrospectively reviewed the data of 16 patients who underwent percutaneous K-wire drill-and-pry between November 2019 and August 2021 for the treatment of supracondylar humeral fractures with bony callus formation ≥14 days after the injury. Clinical outcomes were assessed using the Flynn criteria. The postoperative Baumann angle and pin configuration were evaluated using x-ray examinations. RESULTS: All patients were followed up for 8-28 months (average, 16.63 months). The fractures healed in 4-6 weeks (average, 4.38 weeks). The operative time ranged from 10 to 124 min (average, 35.12 min). No iatrogenic vascular or nerve injury occurred. No patient developed cubitus varus. According to the Flynn criteria, 12 patients had excellent outcomes, 2 patients had good outcomes, 1 patient had a fair outcome and 1 patient had a poor outcome. CONCLUSION: Closed reduction with percutaneous K-wire drill-and-pry is a mini invasive technique for supracondylar humeral fractures with bony callus formation in children. Most patients had a good clinical and cosmetic outcomes without scarring.

Molecular Mechanisms of Plant Regeneration from Differentiated Cells: Approaches from Historical Tissue Culture Systems.

Plants can exert remarkable capacity for cell reprogramming even from differentiated cells. This ability allows plants to regenerate tissues/organs and even individuals in nature and in vitro. In recent decades, Arabidopsis research has uncovered molecular mechanisms of plant regeneration; however, our understanding of how plant cells retain both differentiated status and developmental plasticity is still obscure. In this review, we first provide a brief outlook of the representative modes of plant regeneration and key factors revealed by Arabidopsis research. We then re-examine historical tissue culture systems that enable us to investigate the molecular details of cell reprogramming in differentiated cells and discuss the different approaches, specifically highlighting our recent progress in shoot regeneration from the epidermal cell of Torenia fournieri.

Unveiling the Time Course Mechanism of Bone Fracture Healing by Transcriptional Profiles.

BACKGROUND: Bone fracture healing is a time-consuming and high-priority orthopedic problem worldwide. OBJECTIVE: Discovering the potential mechanism of bone healing at a time course and transcriptional level may better help manage bone fracture. METHODS: In this study, we analyze a time-course bone fracture healing transcriptional dataset in a rat model (GSE592, GSE594, and GSE1371) of Gene Expression Omnibus (GEO). RNA was obtained from female Sprague-Dawley rats with a femoral fracture at the initial time (day 3) as well as early (week 1), middle (week 2), and late (week 4) time periods, with nonfracture rats used as control. Gene Ontology (GO) functional analysis and pathway examinations were performed for further measurements of GSEA and hub genes. RESULTS: Results indicated that the four stages of bone fracture healing at the initial, early, middle, and late time periods represent the phases of hematoma formation, callus formation, callus molding, and mature lamellar bone formation, respectively. Extracellular organization was positively employed throughout the four stages. At the hematoma formation phase, the muscle contraction process was downregulated. Antibacterial peptide pathway was downregulated at all phases. The upregulation of Fn1 (initial, early, middle, and late time periods), Col3a1 (initial, early, and middle time periods), Col11a1 (initial and early time periods), Mmp9 (middle and late time periods), Mmp13 (early, middle, and late time periods) and the downregulation of RatNP-3b (initial, early, middle, and late time periods) were possible symbols for bone fracture healing and may be used as therapeutic targets. CONCLUSION: These findings suggest some new potential pathways and genes in the process of bone fracture healing and further provide insights that can be used in targeted molecular therapy for bone fracture healing.

Auxin-Induced WUSCHEL-RELATED HOMEOBOX13 Mediates Asymmetric Activity of Callus Formation upon Cutting.

Plants have the regenerative ability to reconnect cut organs, which is physiologically important to survive severe tissue damage. The ability to reconnect organs is utilized as grafting to combine two different individuals. Callus formation at the graft junction facilitates organ attachment and vascular reconnection. While it is well documented that local wounding signals provoke callus formation, how callus formation is differentially regulated at each cut end remains elusive. Here, we report that callus formation activity is asymmetrical between the top and bottom cut ends and is regulated by differential auxin accumulation. Gene expression analyses revealed that cellular auxin response is preferentially upregulated in the top part of the graft. Disruption of polar auxin transport inhibited callus formation from the top, while external application of auxin was sufficient to induce callus formation from the bottom, suggesting that asymmetric auxin accumulation is responsible for active callus formation from the top end. We further found that the expression of a key regulator of callus formation, WUSCHEL-RELATED HOMEOBOX 13 (WOX13), is induced by auxin. The ectopic callus formation from the bottom end, which is triggered by locally supplemented auxin, requires WOX13 function, demonstrating that WOX13 plays a pivotal role in auxin-dependent callus formation. The asymmetric WOX13 expression is observed both in grafted petioles and incised inflorescence stems, underscoring the generality of our findings. We propose that efficient organ reconnection is achieved by a combination of local wounding stimuli and disrupted long-distance signaling.

Accelerated tibia fracture healing in traumatic brain injury in accordance with increased hematoma formation.

BACKGROUND: Traumatic brain injury (TBI) has been known to accelerate bone healing. Many cells and molecules have been investigated but the exact mechanism is still unknown. The neuroinflammatory state of TBI has been reported recently. We aimed to investigate the effect of TBI on fracture healing in patients with tibia fractures and assess whether the factors associated with hematoma formation changed more significantly in the laboratory tests in the fractures accompanied with TBI. METHODS: We retrospectively investigated patients who were surgically treated for tibia fractures and who showed secondary bone healing. Patients with and without TBI were divided for comparative analyses. Radiological parameters were time to callus formation and the largest callus ratio during follow-up. Preoperative levels of complete blood count and chemical battery on admission were measured in all patients. Subgroup division regarding age, gender, open fracture, concomitant fracture and severity of TBI were compared. RESULTS: We included 48 patients with a mean age of 44.9 (range, 17-78), of whom 35 patients (72.9%) were male. There were 12 patients with TBI (Group 1) and 36 patients without TBI (Group 2). Group 1 showed shorter time to callus formation (P <  0.001), thicker callus ratio (P = 0.015), leukocytosis and lymphocytosis (P ≤ 0.028), and lower red blood cell counts (RBCs), hemoglobin, and hematocrit (P <  0.001). Aging and severity of TBI were correlated with time to callus formation and callus ratio (P ≤ 0.003) while gender, open fracture, and concomitant fracture were unremarkable. CONCLUSION: Tibia fractures with TBI showed accelerated bone healing and superior measurements associated with hematoma formation (lymphocytes, RBCs, hemoglobin, hematocrit). Promoted fracture healing in TBI was correlated with the enhanced proinflammatory state. LEVEL OF EVIDENCE: III, case control study.

Submergence promotes auxin-induced callus formation through ethylene-mediated post-transcriptional control of auxin receptors.

Plant cells in damaged tissue can be reprogrammed to acquire pluripotency and induce callus formation. However, in the aboveground organs of many species, somatic cells that are distal to the wound site become less sensitive to auxin-induced callus formation, suggesting the existence of repressive regulatory mechanisms that are largely unknown. Here we reveal that submergence-induced ethylene signals promote callus formation by releasing post-transcriptional silencing of auxin receptor transcripts in non-wounded regions. We determined that short-term submergence of intact seedlings induces auxin-mediated cell dedifferentiation across the entirety of Arabidopsis thaliana explants. The constitutive triple response 1-1 (ctr1-1) mutation induced callus formation in explants without submergence, suggesting that ethylene facilitates cell dedifferentiation. We show that ETHYLENE-INSENSITIVE 2 (EIN2) post-transcriptionally regulates the abundance of transcripts for auxin receptor genes by facilitating microRNA393 degradation. Submergence-induced calli in non-wounded regions were suitable for shoot regeneration, similar to those near the wound site. We also observed submergence-promoted callus formation in Chinese cabbage (Brassica rapa), indicating that this may be a conserved mechanism in other species. Our study identifies previously unknown regulatory mechanisms by which ethylene promotes cell dedifferentiation and provides a new approach for boosting callus induction efficiency in shoot explants.

The plant stem-cell niche and pluripotency: 15 years of an epigenetic perspective.

Pluripotent stem-cells are slowly dividing cells giving rise to daughter cells that can either differentiate to new tissues and organs, or remain stem-cells. In plants, stem-cells are located in specific niches of the shoot and root apical meristems (SAMs and RAMs). After ablation of stem-cell niches, pluripotent meristematic cells can establish new stem-cells, whereas the removal of the whole meristem destructs the regeneration process. In tissue cultures, after detached plant organs are transferred to rooting or callus induction medium (G5 or CIM), vasculature-associated pluripotent cells (VPCs) immediately start proliferation to form adventitious roots or callus, respectively, while other cell types of the organ explants basically play no part in the process. Hence, in contrast to the widely-held assumption that all plant cells have the ability to reproduce a complete organism, only few cell types are pluripotent in practice, raising the question how pluripotent stem-cells differ from differentiated cells. It is now clear that, in addition to gene regulatory networks of pluripotency factors and phytohormone signaling, epigenetics play a crucial role in initiation, maintenance and determination of plant stem-cells. Although, more and more epigenetic regulators have been shown to control plant stem-cell fate, only a few studies demonstrate how they are recruited and how they change the chromatin structure and transcriptional regulation of pluripotency factors. Here, we highlight recent breakthroughs but also revisited classical studies of epigenetic regulation and chromatin dynamics of plant stem-cells and their pluripotent precursor-cells, and point out open questions and future directions.

Ectopic expression of WOX5 promotes cytokinin signaling and de novo shoot regeneration.

KEY MESSAGE: WOX5 has a potential in activating cytokinin signaling and shoot regeneration, in addition to its role in pluripotency acquisition. Thus, overexpression of WOX5 maximizes plant regeneration capacity during tissue culture. In vitro plant regeneration involves two steps: callus formation and de novo shoot organogenesis. The WUSCHEL-RELATED HOMEOBOX 5 (WOX5) homeodomain transcription factor is known to be mainly expressed during incubation on callus-inducing medium (CIM) and involved in pluripotency acquisition in callus, but whether WOX5 also affects de novo shoot regeneration on cytokinin-rich shoot-inducing medium (SIM) remains unknown. Based on the recent finding that WOX5 promotes cytokinin signaling, we hypothesized that ectopic expression of WOX5 beyond CIM would further enhance overall plant regeneration capacity, because intense cytokinin signaling is particularly required for shoot regeneration on SIM. Here, we found that overexpression of the WOX5 gene on SIM drastically promoted de novo shoot regeneration from callus with the repression of type-A ARABIDOPSIS RESPONSE REGULATOR (ARR) genes, negative regulators of cytokinin signaling. The enhanced shoot regeneration phenotypes were indeed dependent on cytokinin signaling, which were partially suppressed in the progeny derived from crossing WOX5-overexpressing plants with cytokinin-insensitive 35S:ARR7 plants. The function of WOX5 in enhancing cytokinin-dependent shoot regeneration is evolutionarily conserved, as conditional overexpression of OsWOX5 on SIM profoundly enhanced shoot regeneration in rice callus. Overall, our results provide the technical advance that maximizes in vitro plant regeneration by constitutively expressing WOX5, which unequivocally promotes both callus pluripotency and de novo shoot regeneration.

High auxin stimulates callus through SDG8-mediated histone H3K36 methylation in Arabidopsis.

Callus induction, which results in fate transition in plant cells, is considered as the first and key step for plant regeneration. This process can be stimulated in different tissues by a callus-inducing medium (CIM), which contains a high concentration of phytohormone auxin. Although a few key regulators for callus induction have been identified, the multiple aspects of the regulatory mechanism driven by high levels of auxin still need further investigation. Here, we find that high auxin induces callus through a H3K36 histone methylation-dependent mechanism, which requires the methyltransferase SET DOMAIN GROUP 8 (SDG8). During callus induction, the increased auxin accumulates SDG8 expression through a TIR1/AFBs-based transcriptional regulation. SDG8 then deposits H3K36me3 modifications on the loci of callus-related genes, including a master regulator WOX5 and the cell proliferation-related genes, such as CYCB1.1. This epigenetic regulation in turn is required for the transcriptional activation of these genes during callus formation. These findings suggest that the massive transcriptional reprogramming for cell fate transition by auxin during callus formation requires epigenetic modifications including SDG8-mediated histone H3K36 methylation. Our results provide insight into the coordination between auxin signaling and epigenetic regulation during fundamental processes in plant development.