Exploring distinct properties of endometrial stem cells through advanced single-cell analysis platforms.

The endometrium is a dynamic tissue that undergoes cyclic changes in response to ovarian hormones during the menstrual cycle. These changes are crucial for pregnancy establishment and maintenance. Endometrial stem cells play a pivotal role in endometrial regeneration and repair by differentiating into various cell types within the endometrium. However, their involvement in endometrial disorders such as endometriosis, infertility, and endometrial cancer is still not fully understood yet. Traditional bulk sequencing methods have limitations in capturing heterogeneity and complexity of endometrial stem cell populations. To overcome these limitations, recent single-cell analysis techniques, including single-cell RNA sequencing (scRNA-Seq), single-cell ATAC sequencing (scATAC-Seq), and spatial transcriptomics, have emerged as valuable tools for studying endometrial stem cells. In this review, although there are still many technical limitations that require improvement, we will summarize the current state-of-the-art single-cell analysis techniques for endometrial stem cells and explore their relevance to related diseases. We will discuss studies utilizing various single-cell analysis platforms to identify and characterize distinct endometrial stem cell populations and investigate their dynamic changes in gene expression and epigenetic patterns during menstrual cycle and differentiation processes. These techniques enable the identification of rare cell populations, capture heterogeneity of cell populations within the endometrium, and provide potential targets for more effective therapies.

Unveiling the potential effects of acetylsalicylic acid: insights into regeneration in endometrial stem cells.

BACKGROUND: Although acetylsalicylic acid has been widely used for decades to treat and prevent various diseases, its potential effects on endometrial receptivity and subsequent pregnancy rates are still controversial due to conflicting data: many reports have shown positive effects of acetylsalicylic acid, whereas others have found that it has no effect. Furthermore, the direct effects of acetylsalicylic acid on various functions of normal endometrial cells, especially endometrial stem cells, and their underlying molecular mechanisms have not yet been proven. Recently, studies have revealed that a reduced number of active stem/progenitor cells within endometrial tissue limits cyclic endometrial regeneration and subsequently decreases pregnancy success rates, suggesting that endometrial stem cells play a critical role in endometrial regeneration and subsequent endometrial receptivity. METHODS: We assessed whether aspirin treatment can inhibit various endometrial stem cell functions related to regenerative capacity, such as self-renewal, migration, pluripotency/stemness, and differentiation capacity, in vitro. Next, we evaluated whether SERPINB2 regulates the effects of aspirin on endometrial stem cell functions by depleting SERPINB2 expression with specific shRNA targeting SERPINB2. To further investigate whether aspirin also inhibits various endometrial stem cell functions in vivo, aspirin was administered daily to mice through intraperitoneal (i.p.) injection for 7 days. RESULTS: In addition to its previously identified roles, to the best of our knowledge, we found for the first time that acetylsalicylic acid directly inhibits various human endometrial stem cell functions related to regenerative capacity (i.e., self-renewal, migration, differentiation, and capacity) through its novel target gene SERPINB2 in vitro. Acetylsalicylic acid exerts its function by suppressing well-known prosurvival pathways, such as Akt and/or ERK1/2 signaling, through a SERPINB2 signaling cascade. Moreover, we also found that acetylsalicylic acid markedly inhibits regenerative capacity-related functions in endometrial stem cells within tissue. CONCLUSIONS: We have found that acetylsalicylic acid has diverse effects on various endometrial stem cell functions related to regenerative capacity. Our findings are a critical step toward the development of more effective therapeutic strategies to increase the chances of successful pregnancy. Video Abstract.

Seasonal Forest Changes of Color and Temperature: Effects on the Mood and Physiological State of University Students.

In this study, we attempted to analyze the effect of color and temperature changes in the forest environment over time on the mood and physiological state of university students. The survey was conducted four times considering forest changes such as new leaf appearance and growth, autumn leaf changes, and fallen leaves. The participants' moods and physiological states were first evaluated in an indoor environment; a second evaluation was conducted after contact with the forest. The color visual information of the forest environment was analyzed through color extraction from photographs taken each survey day. The participants' moods and physiological states were measured using the Korean Profile of Mood States-Brief and a heart rate variability measuring device, respectively. Changes in the forest experience according to the season had an effect on university students' mood states. In particular, the effects of the spring forest experience included the relaxation of tension and the activation of vigor. This result is considered to be influenced by factors such as the season's temperature and the green color, which is predominant in the spring forest. However, no physiological changes were found in the participants according to each season. The results of this study can lead to greater consideration of the role of color in urban forest planning for universities and other public spaces.

Current Status and Future Outlook of Additive Manufacturing Technologies for the Reconstruction of the Trachea.

Tracheal stenosis and defects occur congenitally and in patients who have undergone tracheal intubation and tracheostomy due to long-term intensive care. Such issues may also be observed during tracheal removal during malignant head and neck tumor resection. However, to date, no treatment method has been identified that can simultaneously restore the appearance of the tracheal skeleton while maintaining respiratory function in patients with tracheal defects. Therefore, there is an urgent need to develop a method that can maintain tracheal function while simultaneously reconstructing the skeletal structure of the trachea. Under such circumstances, the advent of additive manufacturing technology that can create customized structures using patient medical image data provides new possibilities for tracheal reconstruction surgery. In this study, the three-dimensional (3D) printing and bioprinting technologies used in tracheal reconstruction are summarized, and various research results related to the reconstruction of mucous membranes, cartilage, blood vessels, and muscle tissue, which are tissues required for tracheal reconstruction, are classified. The prospects for 3D-printed tracheas in clinical studies are also described. This review serves as a guide for the development of artificial tracheas and clinical trials using 3D printing and bioprinting.

Comparative structural analysis on the mitochondrial DNAs from various strains of Lentinula edodes.

The evolution of mitochondria through variations in mitochondrial DNA (mtDNA) is one of the intriguing questions in eukaryotic cells. In order to assess the causes of the variations in mitochondria, the mtDNAs of the 21 strains of Lentinula edodes were assembled for this study, and analyzed together with four published mtDNA sequences. The mtDNAs were within the sizes of 117 kb ~ 122 kb. The gene number was observed consistent except for two mtDNAs, which carry a duplicated trnG1-trnG2 unit or a putative gene deletion. The size variation was largely attributed to the number of introns, repeated sequences, transposable elements (TEs), and plasmid-related sequences. Intron loss and gain were found from cox1, rnl, and rns of three mtDNAs. Loss of two introns in cox1 of KY217797.1 reduced its size by 2.7 kb, making it the smallest cox1 gene (8.4 kb) among the cox1s of the 25 mtDNAs, whereas gain of a Group II intron (2.65 kb) and loss of a Group I intron (1.7 kb) in cox1 of MF774813.1 resulted in the longest cox1 (12 kb). In rnl of L. edodes, we discovered four intron insertion consensus sequences which were unique to basidiomycetes but not ascomycetes. Differential incorporation of introns was the primary cause of the rnl size polymorphism. Homing endonucleases (HEGs) were suggestively involved in the mobilization of the introns because all of the introns have HEG genes of the LAGRIDADG or GIY-YIG families with the conserved HEG cleavage sites. TEs contributed to 11.04% of the mtDNA size in average, of which 7.08% was LTR-retrotransposon and 3.96% was DNA transposon, whereas the repeated sequences covered 4.6% of the mtDNA. The repeat numbers were variable in a strain-dependent manner. Both the TEs and repeated sequences were mostly found in the intronic and intergenic regions. Lastly, two major deletions were found in the plasmid-related sequence regions (pol2-pol3 and pol1-atp8) in the five mtDNAs. Particularly, the 6.8 kb-long deletion at pol2-pol3 region made MF774813.1 the shortest mtDNA of all. Our results demonstrate that mtDNA is a dynamic molecule that persistently evolves over a short period of time by insertion/deletion and repetition of DNA segments at the strain level.

The complete mitochondrial genome of the poisonous mushroom Trichoderma cornu-damae (Hypocreaceae).

Trichoderma cornu-damae is a poisonous mushroom that contains trichothecene mycotoxins. The complete mitochondrial genome of this mushroom was determined using next-generation sequencing. This mitogenome is a circular molecule 94,608 bp in length with a GC content of 27.94% and contains 15 protein-coding genes, two rRNA genes (rnl and rns), and 25 tRNA genes. Phylogenetic analysis placed T. cornu-damae in the family Hypocreaceae group, which includes the genus Trichoderma. The mitogenome of T. cornu-damae will contribute to our understanding of the phylogeny, taxonomy, and population genetics of this mushroom.

Pseudo-Chromosomal Genome Assembly in Combination with Comprehensive Transcriptome Analysis in Agaricus bisporus Strain KMCC00540 Reveals Mechanical Stimulus Responsive Genes Associated with Browning Effect.

Agaricus bisporus is one of the world's most popular edible mushrooms, including in South Korea. We performed de novo genome assembly with a South Korean white-colored cultivar of A. bisporus, KMCC00540. After generating a scaffold-level genomic sequence, we inferred chromosome-level assembly by genomic synteny analysis with the representative A. bisporus strains H97 and H39. The KMCC00540 genome had 13 pseudochromosomes comprising 33,030,236 bp mostly covering both strains. A comparative genomic analysis with cultivar H97 indicated that most genomic regions and annotated proteins were shared (over 90%), ensuring that our cultivar could be used as a representative genome. However, A. bisporus suffers from browning even from only a slight mechanical stimulus during transportation, which significantly lowers its commercial value. To identify which genes respond to a mechanical stimulus that induces browning, we performed a time-course transcriptome analysis based on the de novo assembled genome. Mechanical stimulus induces up-regulation in long fatty acid ligase activity-related genes, as well as melanin biosynthesis genes, especially at early time points. In summary, we assembled the chromosome-level genomic information on a Korean strain of A. bisporus and identified which genes respond to a mechanical stimulus, which provided key hints for improving the post-harvest biological control of A. bisporus.

Effects of smoking on the tissue regeneration-associated functions of human endometrial stem cells via a novel target gene SERPINB2.

BACKGROUND: Smokers directly inhale mainstream cigarette smoke, which contains numerous known and potential toxic substances, and thus, smoking is expected to have broad harmful effects that cause tissue injury and dysfunction. Interestingly, many studies have suggested that the recent decline in female fertility and increased rate of spontaneous abortion could be associated with increased smoking rates. Indeed, women that smoked for 10 years or more were reported to have a ~ 20% higher infertility rate than women that had never smoked. However, the reasons for the underlying harmful aspects of smoking on female fertility remain a matter of debate. Importantly, a previous study revealed that resident endometrial stem cell deficiency significantly limits the cyclic regeneration potential of endometrium, which, in turn, decreases successful pregnancy outcomes. In this context, we postulated that exposure to mainstream cigarette smoke extracts might decrease female fertility by inhibiting the functions of resident endometrial stem cells. METHODS: We investigated whether cigarette mainstream smoke exposure directly inhibits various tissue regeneration-associated functions of endometrial stem cells, such as self-renewal, migration, pluripotency, and differentiation capacity in vitro. Next, we determined whether SERPINB2 mediates cigarette smoke-induced suppressive effects on various tissue regeneration-associated functions by depleting SERPINB2 expression with specific shRNA targeting SERPINB2. Mice were injected intraperitoneally with low (0.5 mg/kg) or high (1 mg/kg) doses of cigarette smoke extract (10 times for two weeks), and endometrial stem cells were then isolated from mice uterine tissues. RESULTS: We found that exposure to cigarette smoke extracts remarkably suppressed various tissue regeneration-associated functions of endometrial stem cells, such as self-renewal, migration, multilineage differentiation ability, and pluripotency in vitro and in vivo by activating the SERPINB2 gene. Indeed, cigarette smoke-induced inhibitory effects on various endometrial stem cell functions were significantly abolished by SERPINB2 knockdown. CONCLUSIONS: These findings provide valuable information on the harmful effects of cigarette smoking on resident endometrial stem cells and hopefully will facilitate the developments of promising therapeutic strategies for subfertile or infertile women that smoke cigarettes.

Finite element analysis of femoral component sagittal alignment in mobile-bearing total knee arthroplasty.

BACKGROUND: Recently, there has been an increasing interest in mobile-bearing total knee arthroplasty (TKA). However, changes in biomechanics for femoral component alignment in mobile-bearing TKA have not been explored in depth. OBJECTIVE: This study aims to evaluate the biomechanical effect of sagittal alignment of the femoral component in mobile-bearing TKA. METHODS: We developed femoral sagittal alignment models with -3°, 0°, 3°, 5°, and 7° flexion. We also examine the kinematics of the tibiofemoral (TF) joint, contact point on the TF joint, contact stress on the patellofemoral (PF) joint, collateral ligament force, and quadriceps force using a validated computational model under a deep-knee-bend condition. RESULTS: Posterior kinematics of the TF joint increases as the femoral component flexes. The contact stress on the PF joint, collateral ligament force, and the quadriceps force decreases as the femoral component flexes. CONCLUSIONS: Our results show that a slight, approximately 0°âˆ¼3°, flexion of the implantation could be an effective substitute technique. However, excessive flexion should be avoided because of the potential loosening of the TF joint.

Patient-specific instrumentation improves the reproducibility of preoperative planning for the positioning of baseplate components with reverse total shoulder arthroplasty: a comparative clinical study in 39 patients.

BACKGROUND: The application of patient-specific instrumentation (PSI) for reverse total shoulder replacement has been rapidly increasing, which could reduce errors in implant positioning. Although PSI theoretically holds promise, evidence of the accuracy and reliability of PSI in shoulder replacement surgery is limited. METHODS: Thirty-nine shoulders that underwent reverse total shoulder arthroplasty were included in this study and categorized into two groups: the conventional (n = 20) and PSI (n = 19) groups. Screw (length and angle) and baseplate (version, inclination, translation, and rotation) positioning were calculated based on postoperative computed tomography images using a three-dimensional measurement tool. The difference between the values of the preoperative target and postoperative measurement was calculated to evaluate the reproducibility of preoperative planning. Screw involvement in the suprascapular and spinoglenoid notches was assessed. Thus, the correlation between the position of the baseplate and the screws was assessed. RESULTS: The mean differences between the planned length and angle (anteroposterior and superoinferior angles) and postoperative measurement in the PSI group were significantly smaller than those in the conventional group. Similarly, the mean difference in baseplate rotation between the planned and postoperative measurements in the PSI group was significantly lower than that in the conventional group (4.5° vs 10.6°; P < .001). The spinoglenoid notch was involved in 10 cases in the conventional group and 2 cases in the PSI group, and this difference was significant (P = .014). Overall, the mean difference between the preoperative and postoperative version, inferior inclination, and rotation values for the baseplate position was significantly correlated with the values for screw position (length and angle). CONCLUSIONS: PSI improves the reproducibility of preoperative planning for baseplate and screw positioning and reduces the risk of neurovascular injury in reverse total shoulder arthroplasty.

Targeting Cancer Stem Cell Markers or Pathways: A Potential Therapeutic Strategy for Oral Cancer Treatment.

Cancer stem cells (CSCs) are a small subset of cancer cells with stem cell-like properties, self-renewal potential, and differentiation capacity into multiple cell types. Critical genetic alterations or aberrantly activated signaling pathways associated with drug resistance and recurrence have been observed in multiple types of CSCs. In this context, CSCs are considered to be responsible for tumor initiation, growth, progression, therapeutic resistance, and metastasis. Therefore, to effectively eradicate CSCs, tremendous efforts have been devoted to identify specific target molecules that play a critical role in regulating their distinct functions and to develop novel therapeutics, such as proteins, monoclonal antibodies, selective small molecule inhibitors, and small antisense RNA (asRNA) drugs. Similar to other CSC types, oral CSCs can be characterized by certain pluripotency-associated markers, and oral CSCs can also survive and form 3D tumor spheres in suspension culture conditions. These oral CSC-targeting therapeutics selectively suppress specific surface markers or key signaling components and subsequently inhibit the stem-like properties of oral CSCs. A large number of new therapeutic candidates have been tested, and some products are currently in the pre-clinical or clinical development phase. In the present study, we review new oral CSC-targeted therapeutic strategies and discuss the various specific CSC surface markers and key signaling components involved in the stem-like properties, growth, drug resistance, and tumorigenicity of oral CSCs.

Evaluation of Genetic Diversity and Population Structure Analysis among Germplasm of Agaricus bisporus by SSR Markers.

Agaricus bisporus is a popular edible mushroom that is cultivated worldwide. Due to its secondary homothallic nature, cultivated A. bisporus strains have low genetic diversity, and breeding novel strains is challenging. The aim of this study was to investigate the genetic diversity and population structure of globally collected A. bisporus strains using simple sequence repeat (SSR) markers. Agaricus bisporus strains were divided based on genetic distance-based groups and model-based subpopulations. The major allele frequency (MAF), number of genotypes (NG), number of alleles (NA), observed heterozygosity (HO), expected heterozygosity (HE), and polymorphic information content (PIC) were calculated, and genetic distance, population structure, genetic differentiation, and Hardy-Weinberg equilibrium (HWE) were assessed. Strains were divided into two groups by distance-based analysis and into three subpopulations by model-based analysis. Strains in subpopulations POP A and POP B were included in Group I, and strains in subpopulation POP C were included in Group II. Genetic differentiation between strains was 99%. Marker AB-gSSR-1057 in Group II and subpopulation POP C was confirmed to be in HWE. These results will enhance A. bisporus breeding programs and support the protection of genetic resources.

3D stem cell-laden artificial endometrium: successful endometrial regeneration and pregnancy.

Thin endometrium lining or severe endometrial injury which may occur during artificial abortion can cause defective endometrial receptivity and subsequent infertility. Therefore, much effort has been devoted toward regenerating thin or damaged endometrial lining by applying multiple types of stem cells. Even though there are some positive preliminary outcomes, repairing the injured endometrium with stem cells is considerably challenging, due to the lack of an adequate microenvironment for the administrated stem cells within the tissues and subsequent poor therapeutic efficiency. In this context, as an alternative, we fabricated a 3D stem cell-laden artificial endometrium by incorporating several biodegradable biomaterials (collagen and hyaluronic acid) and multiple cellular components of endometrium (endometrial stem cells, stromal cells, and vessel cells) to properly recapitulate the multicellular microenvironment and multilayered structure. Agarose was used as an inert filler substrate to enhance the mechanical integrity of the three-layered artificial endometrium. Various mechanical characteristics, such as morphology, compression properties, swelling, and viscosity, have been evaluated. Various biological features, such as steroid hormone responsiveness, specific endometrial cell-surface marker expressions, and the secretion of multiple growth factors and steroid hormones, as well as the viability of encapsulated endometrial cells are relatively well maintained within the artificial endometrium. More importantly, severe tissue injuries were significantly relieved by transplanting our 3D artificial endometrium into endometrial ablation mice. Remarkably, artificial endometrium transplantation resulted in a successful pregnancy with subsequent live birth without any morphological or chromosomal abnormalities.

Development of CAPS Markers for Evaluation of Genetic Diversity and Population Structure in the Germplasm of Button Mushroom (Agaricus bisporus).

Agaricus bisporus is a globally cultivated mushroom with high economic value. Despite its widespread cultivation, commercial button mushroom strains have little genetic diversity and discrimination of strains for identification and breeding purposes is challenging. Molecular markers suitable for diversity analyses of germplasms with similar genotypes and discrimination between accessions are needed to support the development of new varieties. To develop cleaved amplified polymorphic sequences (CAPs) markers, single nucleotide polymorphism (SNP) mining was performed based on the A. bisporus genome and resequencing data. A total of 70 sets of CAPs markers were developed and applied to 41 A. bisporus accessions for diversity, multivariate, and population structure analyses. Of the 70 SNPs, 62.85% (44/70) were transitions (G/A or C/T) and 37.15% (26/70) were transversions (A/C, A/T, C/G, or G/T). The number of alleles per locus was 1 or 2 (average = 1.9), and expected heterozygosity and gene diversity were 0.0-0.499 (mean = 0.265) and 0.0-0.9367 (mean = 0.3599), respectively. Multivariate and cluster analyses of accessions produced similar groups, with F-statistic values of 0.134 and 0.153 for distance-based and model-based groups, respectively. A minimum set of 10 markers optimized for accession identification were selected based on high index of genetic diversity (GD, range 0.299-0.499) and major allele frequency (MAF, range 0.524-0.817). The CAPS markers can be used to evaluate genetic diversity and population structure and will facilitate the management of emerging genetic resources.

Noncanonical functions of glucocorticoids: A novel role for glucocorticoids in performing multiple beneficial functions in endometrial stem cells.

Chronic stress has a negative impact on many fertility-related functions; thus, the recent decline in female fertility seems to be at least partially associated with increased stress. The secretion of glucocorticoids is a typical endocrine response to chronic stress and indirectly reduces uterine receptivity through the hypothalamus-pituitary-gonadal (HPG) axis. However, in addition to its well-known canonical role, the direct effects of chronic stress-induced glucocorticoids on various uterine functions and their underlying molecular mechanisms are complex and have not yet been revealed. Recent studies have found that resident stem cell deficiency is responsible for the limited regenerative potential of the endometrium (the innermost lining of the uterine cavity) during each menstrual cycle, which subsequently increases infertility rates. In this context, we hypothesized that stress-induced glucocorticoids directly damage endometrial stem cells and consequently negatively affect endometrial reconstruction, which is important for uterine receptivity. In addition to its well-known canonical roles, we identified for the first time that cortisol, the most abundant and potent glucocorticoid in humans, directly suppresses the multiple beneficial functions (self-renewal, transdifferentiation, and migratory potential) of human endometrial stem cells through its functional receptor, glucocorticoid receptor (GR). Glucocorticoids inhibit well-known survival signals, such as the PI3K/Akt and FAK/ERK1/2 pathways. More importantly, we also found that immobilization of stress-induced glucocorticoids suppresses the various beneficial functions of tissue resident stem cells in vivo. To the best of our knowledge, this is the first study to investigate the direct effects of glucocorticoids on the regenerative capacity of endometrial stem cells, and the findings will facilitate the development of more promising therapeutic approaches to increase female fertility.

Brassinosteroid-BZR1/2-WAT1 module determines the high level of auxin signalling in vascular cambium during wood formation.

The tight regulation of local auxin homeostasis and signalling maxima in xylem precursor cells specifies the organising activity of the vascular cambium and consequently promotes xylem differentiation and wood formation. However, the molecular mechanisms underlying the local auxin signalling maxima in the vascular cambium are largely unknown. Here, we reveal that brassinosteroid (BR)-activated WALLS ARE THIN1 (WAT1) facilitates wood formation by enhancing local auxin signalling in the vascular cambium in Solanum lycopersicum. Growth defects and low auxin signalling readouts in the BR-deficient tomato cultivar, Micro-Tom, were associated with a novel recessive allele, Slwat1-copi, created by the insertion of a retrotransposon in the last exon of the SlWAT1 locus. Molecular and genetic studies by generating the gain-of-function and loss-of-function tomato mutants revealed that SlWAT1 is a critical regulator for fine tuning local auxin homeostasis and signalling outputs in vascular cambium to facilitate secondary growth. Finally, we discovered that BR-regulated SlBZR1/2 directly activated downstream auxin responses by SlWAT1 upregulation in xylem precursor cells to facilitate xylem differentiation and subsequent wood formation. Our data suggest that the BR-SlBZR1/2-WAT1 signalling network contributes to the high level of auxin signalling in the vascular cambium for secondary growth.

Influence of Variation in Sagittal Placement of the Femoral Component after Cruciate-Retaining Total Knee Arthroplasty.

Prosthetic alignment is an important factor for long-term survival in cruciate-retaining (CR) total knee arthroplasty (TKA). The purpose of this study is to investigate the influence of sagittal placement of the femoral component on tibiofemoral (TF) kinematics and kinetics in CR-TKA. Five sagittal placements of femoral component models with -3, 0, 3, 5, and 7 degrees of flexion are developed. The TF joint kinematics, quadriceps force, patellofemoral contact force, and posterior cruciate ligament force are evaluated using the models under deep knee-bend loading. The kinematics of posterior TF translation is found to occur with the increase in femoral-component flexion. The quadriceps force and patellofemoral contact force decrease with the femoral-component flexion increase. In addition, extension of the femoral component increases with the increase in posterior cruciate ligament force. The flexed femoral component in CR-TKA provides a positive biomechanical effect compared with a neutral position. Slight flexion could be an effective alternative technique to enable positive biomechanical effects with TKA prostheses.

Functional Characterization of BRASSINAZOLE-RESISTANT 1 in Panax Ginseng (PgBZR1) and Brassinosteroid Response during Storage Root Formation.

Brassinosteroids (BRs) play crucial roles in the physiology and development of plants. In the model plant Arabidopsis, BR signaling is initiated at the level of membrane receptors, BRASSINOSTEROIDS INSENSITIVE 1 (BRI1) and BRI1-ASSOCIATED RECEPTOR KINASE 1 (BAK1) complex, thus activating the transcription factors (TFs) BRASSINAZOLE RESISTANT 1/BRI1-EMS-SUPPRESSOR 1 (BZR1/BES1) to coordinate BR responsive genes. BRASSINOSTEROIDS INSENSITIVE 2 (BIN2), glycogen synthase kinase 3 (GSK3) like-kinase, negatively regulates BZR1/BES1 transcriptional activity through phosphorylation-dependent cytosolic retention and shuttling. However, it is still unknown whether this mechanism is conserved in Panax ginseng C. A. Mayer, a member of the Araliaceae family, which is a shade-tolerant perennial root crop. Despite its pharmacological and agricultural importance, the role of BR signaling in the development of P. ginseng and characterization of BR signaling components are still elusive. In this study, by utilizing the Arabidopsisbri1 mutant, we found that ectopic expression of the gain of function form of PgBZR1 (Pgbzr1-1D) restores BR deficiency. In detail, ectopic expression of Pgbzr1-1D rescues dwarfism, defects of floral organ development, and hypocotyl elongation of bri1-5, implying the functional conservation of PgBZR1 in P. ginseng. Interestingly, brassinolide (BL) and BRs biosynthesis inhibitor treatment in two-year-old P. ginseng storage root interferes with and promotes, respectively, secondary growth in terms of xylem formation. Altogether, our results provide new insight into the functional conservation and potential diversification of BR signaling and response in P. ginseng.

Functional Analysis of a Novel ABL (Abnormal Browning Related to Light) Gene in Mycelial Brown Film Formation of Lentinula edodes.

Lentinula edodes is a globally important edible mushroom species that is appreciated for its medicinal properties as well as its nutritional value. During commercial cultivation, a mycelial brown film forms on the surface of the sawdust growth medium at the late vegetative stage. Mycelial film formation is a critical developmental process that contributes to the quantity and quality of the mushroom yield. However, little is known regarding the genetic underpinnings of brown film formation on the surface of mycelial tissue. A novel causal gene associated with the formation of the mycelial brown film, named ABL (Abnormal browning related to light), was identified in this study. The comparative genetic analysis by dihybrid crosses between normal and abnormal browning film cultivars demonstrated that a single dominant allele was responsible for the abnormal mycelium browning phenotype. Whole-genome sequencing analysis of hybrid isolates revealed five missense single-nucleotide polymorphisms (SNPs) in the ABL locus of individuals forming abnormal partial brown films. Additional whole-genome resequencing of a further 16 cultivars showed that three of the five missense SNPs were strongly associated with the abnormal browning phenotype. Overexpression of the dominant abl-D allele in a wild-type background conferred the abnormal mycelial browning phenotype upon transformants, with slender hyphae observed as a general defective mycelial growth phenotype. Our methodology will aid the future discovery of candidate genes associated with favorable traits in edible mushrooms. The discovery of a novel gene, ABL, associated with mycelial film formation will facilitate marker-associated breeding in L. edodes.