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.

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.

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.

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.

Genetic variations in ZmSAUR15 contribute to the formation of immature embryo-derived embryonic calluses in maize.

The ability of immature maize (Zea mays) embryos to form embryonic calluses (ECs) is highly genotype dependent, which limits transgenic breeding development in maize. Here, we report the association map-based cloning of ZmSAUR15 using an association panel (AP) consisting of 309 inbred lines with diverse formation abilities for ECs. We demonstrated that ZmSAUR15, which encodes a small auxin-upregulated RNA, acts as a negative effector in maize EC induction. Polymorphisms in the ZmSAUR15 promoter that influence the expression of ZmSAUR15 transcripts modulate the EC induction capacity in maize. ZmSAUR15 is involved in indole-3-acetic acid biosynthesis and cell division in immature embryo-derived callus. The ability of immature embryos to induce EC formation can be improved by the knockout of ZmSAUR15, which consequently increases the callus regeneration efficiency. Our study provides new insights into overcoming the genotypic limitations associated with EC formation and improving genetic transformation in maize.

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.

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.

The atypical histone variant H3.15 promotes callus formation in Arabidopsis thaliana.

Plants are capable of regenerating new organs after mechanical injury. The regeneration process involves genome-wide reprogramming of transcription, which usually requires dynamic changes in the chromatin landscape. We show that the histone 3 variant HISTONE THREE RELATED 15 (H3.15) plays an important role in cell fate reprogramming during plant regeneration in Arabidopsis H3.15 expression is rapidly induced upon wounding. Ectopic overexpression of H3.15 promotes cell proliferation to form a larger callus at the wound site, whereas htr15 mutation compromises callus formation. H3.15 is distinguished from other Arabidopsis histones by the absence of the lysine residue 27 that is trimethylated by the POLYCOMB REPRESSIVE COMPLEX 2 (PRC2) in constitutively expressed H3 variants. Overexpression of H3.15 promotes the removal of the transcriptional repressive mark H3K27me3 from chromatin, which results in transcriptional de-repression of downstream genes, such as WUSCHEL RELATED HOMEOBOX 11 (WOX11). Our results reveal a new mechanism for a release from PRC2-mediated gene repression through H3.15 deposition into chromatin, which is involved in reprogramming cell fate to produce pluripotent callus cells.

Specific Characteristic of Hyperplastic Callus in a Larger Cohort of Osteogenesis Imperfecta Type V.

Hyperplastic callus (HPC) is the most conspicuous features of osteogenesis imperfecta (OI) type V, of which accurate diagnosis and treatment are facing challenges. We investigate the clinical features, and impact factors of HPC in OI type V patients. In this retrospective single-center study, a total of 21 patients with type V OI confirmed by IFITM5 mutation were included. Radiological characteristics of bone were evaluated by X-rays, dual-energy X-ray absorptiometry, and computed tomography scan. Bone biopsy specimens were performed and stained by routine hematoxylin-eosin. The effects of bisphosphonates on HPC were investigated. Eleven patients (52.3%) had HPCs at 19 skeletal sites, 11 of which affected the femur. Three patients developed four (21.1%) HPCs after fractures, and 15 (78.9%) HPCs occurred in absence of bone fracture. The progress of HPCs was variable, of which most HPCs enlarged in the initial phase and remained stable, and only one HPC dwindled in size. One patient had a rapidly growing mass on the right humerus, and biopsy showed irregular trabeculae of woven bone and immature bone and cartilage in the loose and edematous collagenous network without signs of tumor. Bisphosphonates treatment had no significant effects on HPC of OI patients. HPC is the specific characteristic of OI type V patients, and its location, shape, size, and progression are variable, and the femur is the most frequently involved site. It is very important to make a diagnosis of HPC through detecting IFITM5 mutation and completing pathological diagnosis if necessary. The treatment of HPC is worth further exploration.

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.

Piezoelectric and Opto-Acoustic Material Properties of Bone.

In this chapter, recent piezoelectric and opto-acoustic studies on bone are introduced. The former are certainly related to ultrasound since piezoelectricity is one of the electro-mechanical properties. The latter are divided into two parts: Photo Acoustics (PA) and Brillouin Scattering (BS). PA is the energy conversion from light to ultrasound while Brillouin scattering is the interaction between phonons and photons. These studies seem very different; however, they are all studies on the ultrasonic material characterization of bone. Another common aspect of these studies is that they are generally targeting the material characterization of bone extracellular matrix. These studies have started later than the conventional ultrasonic bone studies and are expected to provide different characteristics of bone in the micrometer scale area.

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.

PUCHI regulates very long chain fatty acid biosynthesis during lateral root and callus formation.

Lateral root organogenesis plays an essential role in elaborating plant root system architecture. In Arabidopsis, the AP2 family transcription factor PUCHI controls cell proliferation in lateral root primordia. To identify potential targets of PUCHI, we analyzed a time course transcriptomic dataset of lateral root formation. We report that multiple genes coding for very long chain fatty acid (VLCFA) biosynthesis enzymes are induced during lateral root development in a PUCHI-dependent manner. Significantly, several mutants perturbed in VLCFA biosynthesis show similar lateral root developmental defects as puchi-1 Moreover, puchi-1 roots display the same disorganized callus formation phenotype as VLCFA biosynthesis-deficient mutants when grown on auxin-rich callus-inducing medium. Lipidomic profiling of puchi-1 roots revealed reduced VLCFA content compared with WT. We conclude that PUCHI-regulated VLCFA biosynthesis is part of a pathway controlling cell proliferation during lateral root and callus formation.

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.

OsHDA710-Mediated Histone Deacetylation Regulates Callus Formation of Rice Mature Embryo.

Histone deacetylases (HDACs) play important roles in the regulation of eukaryotic gene expression. The role of HDACs in specialized transcriptional regulation and biological processes is poorly understood. In this study, we evaluated the global expression patterns of genes related to epigenetic modifications during callus initiation in rice. We found that the repression of HDAC activity by trichostatin A (TSA) or by OsHDA710 mutation (hda710) results in impaired callus formation of rice mature embryo and increased global histone H3 acetylation levels. The HDAC inhibition decreased auxin response and cell proliferation in callus formation. Meanwhile, the transcriptional repressors OsARF18 and OsARF22 were upregulated in the callus of hda710. The chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) analysis demonstrated that the callus of hda710 exhibited enhanced histone H3 acetylation levels at the chromatin regions of OsARF18 and OsARF22. Furthermore, we found that OsARF18 and OsARF22 were regulated through OsHDA710 recruitment to their target loci. In addition, overexpression of OsARF18 decreased the transcription of downstream genes PLT1 and PLT2 and inhibited callus formation of the mature embryo. These results demonstrate that OsHDA710 regulates callus formation by suppressing repressive OsARFs via histone deacetylation during callus formation of rice mature embryo. This indicates that OsHDA710-mediated histone deacetylation is an epigenetic regulation pathway for maintaining auxin response during cell dedifferentiation.

Analysis of low-dose estrogen on callus BMD as measured by pQCT in postmenopausal women.

BACKGROUND: Osteoporosis affects elderly patients of both sexes. It is characterized by an increased fracture risk due to defective remodeling of the bone microarchitecture. It affects in particular postmenopausal women due to their decreased levels of estrogen. Preclinical studies with animals demonstrated that loss of estrogen had a negative effect on bone healing and that increasing the estrogen level led to a better bone healing. We asked whether increasing the estrogen level in menopausal patients has a beneficial effect on bone mineral density (BMD) during callus formation after a bone fracture. METHODS: To investigate whether estrogen has a beneficial effect on callus BMD of postmenopausal patients, we performed a prospective double-blinded randomized study with 76 patients suffering from distal radius fractures. A total of 31 patients (71.13 years ±11.99) were treated with estrogen and 45 patients (75.62 years ±10.47) served as untreated controls. Calculated bone density as well as cortical bone density were determined by peripheral quantitative computed tomography (pQCT) prior to and 6 weeks after the surgery. Comparative measurements were performed at the fractured site and at the corresponding position of the non-fractured arm. RESULTS: We found that unlike with preclinical models, bone fracture healing of human patients was not improved in response to estrogen treatment. Furthermore, we observed no dependence between age-dependent bone tissue loss and constant callus formation in the patients. CONCLUSIONS: Transdermally applied estrogen to postmenopausal women, which results in estrogen levels similar to the systemic level of premenopausal women, has no significant beneficial effect on callus BMD as measured by pQCT, as recently shown in preclinical animal models. TRIAL REGISTRATION: Low dose estrogen has no significant effect on bone fracture healing measured by pQCT in postmenopausal women, DRKS00019858 . Registered 25th November 2019 - Retrospectively registered. Trial registration number DRKS00019858 .

Variable fixation promotes callus formation: an experimental study on transverse tibial osteotomies stabilized with locking plates.

BACKGROUND: A new locking screw technology, named variable fixation, has been developed aiming at promoting bone callus formation providing initial rigid fixation followed by progressive fracture gap dynamisation. In this study, we compared bone callus formation in osteotomies stabilized with standard locking fixation against that of osteotomies stabilized with variable fixation in an established tibia ovine model. METHODS: A 3 mm tibial transverse osteotomy gap was stabilized in three groups of six female sheep each with a locking plate and either 1) standard fixation in both segments (group LS) or 2) variable fixation in the proximal and standard fixation in the distal bone segment (group VFLS3) or 3) variable fixation in both segments (group VFLS6). The implantation site and fracture healing were compared between groups by means of radiologic, micro tomographic, biomechanical, and histological investigations. RESULTS: Compared to LS callus, VFLS3 callus was 40% larger and about 3% denser, while VFLS6 callus was 93% larger and its density about 7.2% lower. VFLS3 showed 65% and VFLS6 163% larger amount of callus at the cis-cortex. There wasn't a significant difference in the amount of callus at the cis and trans-cortex in groups featuring variable fixation only. Investigated biomechanical variables were not significantly different among groups and histology showed comparable good healing in all groups. Tissues adjacent to the implants did not show any alteration of the normal structure in all groups. CONCLUSIONS: Variable fixation promoted the formation of a larger amount of bone callus, equally distributed at the cis and trans cortices. The histological and biomechanical properties of the variable fixation callus were equivalent to those of the standard fixation callus. The magnitude of variable fixation had a biological effect on the formation of bone callus. At the implantation site, the usage of variable fixation did not raise additional concerns with respect to standard fixation. The formation of a larger amount of mature callus suggests that fractures treated with variable fixation might have a higher probability to bridge the fracture gap. The conditions where its usage can be most beneficial for patients needs to be clinically defined.

Factors Associated with Callus Formation in the Plantar Region through Gait Measurement in Patients with Diabetic Neuropathy: An Observational Case-Control Study.

Callus has been identified as a risk factor leading to severe diabetic foot ulcer; thus, it is necessary to prevent its formation. Callus formation under the first, second, and fifth metatarsal heads (MTHs) is associated with external forces (pressure and shear stress) during walking. However, the gait factors increasing the external forces remain undetermined. Thus, this study aims to identify the factors increasing the external forces to prevent callus formation. In 59 patients with diabetic neuropathy wearing their usual shoes, the external forces, and the lower extremity joint angles were measured using MEMS force sensors and motion sensors. The external forces and their relationship with the lower extremity joint angles and footwear size were determined. Risk factors causing high external forces on the first MTH included small flexion of the knee joint (p = 0.015) and large ankle pronation motion (p = 0.034) to obtain propulsion. For the second MTH, wearing excessively long footwear was identified (p = 0.026). For the fifth MTH, high external force was related to tight width footwear (p = 0.005). An effective intervention for preventing callus formation for the first MTH would involve assisting the push-off foot motion using rocker-sole footwear or gait training. For the second and fifth MTHs, wearing appropriate size footwear would be effective.

WIND1 induces dynamic metabolomic reprogramming during regeneration in Brassica napus.

Plants often display a high competence for regeneration under stress conditions. Signals produced in response to various types of stress serve as critical triggers for de novo organogenesis, but the identity of these signaling molecules underlying cellular reprogramming are largely unknown. We previously identified an AP2/ERF transcription factor, WOUND INDUCED DEDIFFERENTIATION1 (WIND1), as a key regulator involved in wound-induced cellular reprogramming in Arabidopsis. In this study, we found that activation of Arabidopsis WIND1 (AtWIND1) in hypocotyl explants of Brassica napus (B. napus) enhances callus formation and subsequent organ regeneration. Gene expression analyses revealed that AtWIND1 enhances expression of B. napus homologs of ENHANCER OF SHOOT REGENERATION1/DORNRÖSCHEN (ESR1/DRN), which is a direct target of WIND1 in Arabidopsis. Further, time-course hormonal analyses showed that an altered balance of endogenous auxin/cytokinin exists in AtWIND1-activated B. napus explants. Our mass spectrometry analyses, in addition, uncovered dynamic metabolomic reprogramming in AtWIND1-activated explants, including accumulation of several compounds, e.g. proline, gamma aminobutyric acid (GABA), and putrescine, that have historically been utilized as additives to enhance plant cell reprogramming in tissue culture. Our findings thus provide new insights into how WIND1 functions to promote cell reprogramming.

Stem cells and plant regeneration.

Multicellular organisms show the ability to replace damage cells, tissues and even whole organs through regeneration mechanisms. Plants show a remarkable regenerative potential. While the basic principles of plant regeneration have been known for a number of decades, the molecular and cellular mechanisms underlying such principles are currently starting to emerge. Some of these mechanisms point to the existence of highly reprogrammable cells. Developmental plasticity is a hallmark for stem cells, and stem cells are responsible for the generation of distinctive cell types forming plants. In the last years, a number of players and molecular mechanism regulating stem cell maintenance have been described, and some of them have also been involved in regenerative processes. These discoveries in plant stem cell regulation and regeneration invite us to rethink several of the classical concepts in plant biology such as cell fate specification and even the actual meaning of what we consider stem cells in plants. In this review we will cover some of these discoveries, focusing on the role of the plant stem cell function and regulation during cell and organ regeneration.