Morphogenic Pathways Research Articles

Evolution and biomechanical variation in mammalian ungual phalanges with respect to locomotion

Past studies of the morphology of the claw-bearing phalanges among mammalian taxa has revealed a correspondence of ungual phalanx shape and the broad form of locomotor behavior carried out by those taxa. Further research has provided evidence for convergence in the morphology of ungual phalanges among distantly related taxa that engage in similar locomotion. This evolutionary parallelism is proposed to have been motivated by conserved, shared morphogenic pathways. Such evidence would argue that, despite phylogenetic divergence, we would expect to observe: 1) similar biomechanical properties in ungual cross-sectional geometry among unrelated taxa performing similar forms of locomotion; and 2) phylogenetic modeling of the evolution of ungual phalanx morphology should demonstrate evidence for selection and convergence. In this study, we test these hypotheses using linear dimensions and cross-sectional geometric properties (CGSP) obtained from 42 mammalian taxa. This sample represents a breadth of taxa that have previously been classified into five broad locomotor guilds (arboreal, cursorial, fossorial, generalist, scansorial). CSGP (Ix, Iy, J) were obtained using a Stratec pQCT scanner, and linear dimensions of the phalangeal length, mid-length mediolateral width and dorsopalmar height, and extensor and flexor lever arm lengths were taken using digital Mitutoyo sliding calipers. MANOVAs of linear and CSGP traits demonstrate significant differences in locomotor groups (Pillai's Trace = 0.7206; p=0.0007). Discriminant function analysis shows this is motivated by J and the proportion of mediolateral width to dorsopalmar height of the phalangeal mid-length. We then subjected these properties to phylogenetic models assessing evolutionary models of Brownian motion (using R packages ‘bayou’ and ‘OUwie’). Results present little evidence for shared selective regime changes among taxa. One notable model results indicates selective shifts that occurred among taxa that evolved from an ancestral state of mediolaterally narrow phalanges to broader phalanges (as would be associated with fossorial behaviors). Thus, while convergences in shape and CSGP that relate to locomotor behavior are observed among taxa, phylogenetic comparative methods support selection as occurring only in species that evolved from a narrow to broader ungual phalanx morphology. Further analyses using OU models were used to assess phylogenetic models of convergence.

14. Prenatal diagnosis of Acrofacial Dysostosis type 1 (Nager syndrome) by chromosomal microarray at the exon level

Nager syndrome or Acrofacial Dysostosis type 1 (OMIM #154400) is a rare, autosomal dominant syndrome characterized by craniofacial malformations. Fewer than 100 cases have been reported in the literature and of these, only 11 were suspected prenatally. Major phenotypic findings include micrognathia, ear anomalies and limb malformations; however, the severity is variable. Haploinsufficiency of the SF3B4 (1q21.2) gene is the most commonly recognized etiology of Nager syndrome and can be identified in more than 50% of clinically affected patients. SF3B4 encodes for the SAP49 protein which is a component of the spliceosome and has a role in the bone morphogenic protein pathway. The majority of Nager syndrome cases involve missense and nonsense mutations, or whole gene deletions. We report on a 20 year old primigravida who underwent amniocentesis at 23+1 weeks due to ultrasound findings of severe micrognathia, ear anomalies, mild hypotelorism, and absent left thumb. Chromosome microarray identified a 166kb partial deletion of the SF3B4 gene spanning exons 3-6. Copy number PCR was performed to confirm the finding and to determine whether the deletion was maternally inherited. Both independent DNA extractions of the amniotic fluid sample had a copy number of one and the maternal sample had a copy number of two, confirming the deletion. To the best of our knowledge, this prenatal diagnosis of Nager syndrome by chromosomal microarray is the first partial gene deletion of SF3B4 reported. Further, this demonstrates the importance of chromosomal microarray in correlating fetal ultrasound findings with genetic etiology. Knowledge of this result prior to birth allowed more robust family counseling about the findings and prognosis, along with adequate medical preparation for the anticipated complex interventional needs of the neonate at and after delivery.

STIs during the first and second wave of COVID-19 in Denmark

<h3>Abstract</h3> The increase in activity of the two-pore potassium-leak channel Kcnk5b maintains allometric juvenile growth of adult zebrafish appendages. However, it remains unknown how this channel maintains allometric growth and...

Medical Cannabis and Industrial Hemp Tissue Culture: Present Status and Future Potential.

In recent years high-THC (psychoactive) and low-THC (industrial hemp) type cannabis (Cannabis sativa L.) have gained immense attention in medical, food, and a plethora of other consumer product markets. Among the planting materials used for cultivation, tissue culture clones provide various advantages such as economies of scale, production of disease-free and true-to-type plants for reducing the risk of GMP-EuGMP level medical cannabis production, as well as the development and application of various technologies for genetic improvement. Various tissue culture methods have the potential application with cannabis for research, breeding, and novel trait development, as well as commercial mass propagation. Although tissue culture techniques for plant regeneration and micropropagation have been reported for different cannabis genotypes and explant sources, there are significant variations in the response of cultures and the morphogenic pathway. Methods for many high-yielding elite strains are still rudimentary, and protocols are not established. With a recent focus on sequencing and genomics in cannabis, genetic transformation systems are applied to medical cannabis and hemp for functional gene annotation via traditional and transient transformation methods to create novel phenotypes by gene expression modulation and to validate gene function. This review presents the current status of research focusing on different aspects of tissue culture, including micropropagation, transformation, and the regeneration of medicinal cannabis and industrial hemp transformants. Potential future tissue culture research strategies helping elite cannabis breeding and propagation are also presented.

Disruption of morphogenic and growth pathways in lysosomal storage diseases.

The lysosome achieved a new protagonism that highlights its multiple cellular functions, such as in the catabolism of complex substrates, nutrient sensing, and signaling pathways implicated in cell metabolism and growth. Lysosomal storage diseases (LSDs) cause lysosomal accumulation of substrates and deficiency in trafficking of macromolecules. The substrate accumulation can impact one or several pathways which contribute to cell damage. Autophagy impairment and immune response are widely studied, but less attention is paid to morphogenic and growth pathways and its impact on the pathophysiology of LSDs. Hedgehog pathway is affected with abnormal expression and changes in distribution of protein levels, and a reduced number and length of primary cilia. Moreover, growth pathways are identified with delay in reactivation of mTOR that deregulate termination of autophagy and reformation of lysosomes. Insulin resistance caused by changes in lipids rafts has been described in different LSDs. While the genetic and biochemical bases of deficient proteins in LSDs are well understood, the secondary molecular mechanisms that disrupt wider biological processes associated with LSDs are only now becoming clearer. Therefore, we explored how specific signaling pathways can be related to specific LSDs, showing that a system medicine approach could be a valuable tool for the better understanding of LSD pathogenesis. This article is categorized under: Metabolic Diseases > Molecular and Cellular Physiology.

Hedgehog Signaling in Colorectal Cancer: All in the Stroma?

Hedgehog (Hh) signaling regulates intestinal development and homeostasis. The role of Hh signaling in cancer has been studied for many years; however, its role in colorectal cancer (CRC) remains controversial. It has become increasingly clear that the “canonical” Hh pathway, in which ligand binding to the receptor PTCH1 initiates a signaling cascade that culminates in the activation of the GLI transcription factors, is mainly organized in a paracrine manner, both in the healthy colon and in CRC. Such canonical Hh signals largely act as tumor suppressors. In addition, stromal Hh signaling has complex immunomodulatory effects in the intestine with a potential impact on carcinogenesis. In contrast, non-canonical Hh activation may have tumor-promoting roles in a subset of CRC tumor cells. In this review, we attempt to summarize the current knowledge of the Hh pathway in CRC, with a focus on the tumor-suppressive role of canonical Hh signaling in the stroma. Despite discouraging results from clinical trials using Hh inhibitors in CRC and other solid cancers, we argue that a more granular understanding of Hh signaling might allow the exploitation of this key morphogenic pathway for cancer therapy in the future.

Reconstitution of Morphogen Signaling Gradients in Cultured Cells.

Development of multicellular organisms depends on the proper establishment of signaling information in space and time. Secreted molecules called morphogens form concentration gradients in space and provide positional information to differentiating cells within the organism. Although the key molecular components of morphogen pathways have been identified, how the architectures and key parameters of morphogen pathways control the properties of signaling gradients, such as their size, speed, and robustness to perturbations, remains challenging to study in developing embryos. Reconstituting morphogen gradients in cell culture provides an alternative approach to address this question. Here we describe the methodology for reconstituting Sonic Hedgehog (SHH) signaling gradients in mouse fibroblast cells. The protocol includes the design of morphogen sending and receiving cell lines, the setup of radial and linear gradients, the quantitative time-lapse imaging, and the data analysis. Similar approaches could potentially be applied to other cell-cell communication pathways.

Global 5-methylcytosine and physiological changes are triggers of indirect somatic embryogenesis in Coffea canephora.

Indirect somatic embryogenesis (ISE) establishment for Coffea species started in the 1970s. Since then, intraspecific variations in the morphogenic pathway have been reported, even in the common environmental condition in vitro. Several authors have suggested that these variations are the result of genetic, epigenetic, and/or physiological events, highlighting the need for investigations to know the causes. Along these lines, this study aimed to investigate and describe, for the first time, the global 5-methylcytosine and physiological changes that occur in the cells of the aggregate suspensions of Coffea canephora during proliferation and somatic embryo regeneration steps. The cell proliferation step was characterized by increase in cell mass in all subcultures; relatively low mean values of global 5-methylcytosine (5-mC%), abscisic acid (ABA), and indole-3-acetic acid (IAA); high mean value of 1-aminocyclopropane-1-carboxylic acid (ACC, an ethylene precursor); and increase followed by decrease in spermidine (Spd, a polyamine) level. Therefore, these epigenetic and physiologic aspects promoted the cell proliferation, which is fundamental for ISE. In turn, the somatic embryo regeneration was correlated with global 5-mC% and physiological changes. The competence acquisition, determination, and cell differentiation steps were marked by increases in mean values of 5-mC%, IAA and ABA, and decreases in ACC and Spd, evincing that these changes are the triggers for regeneration and maturation of somatic embryos. Therefore, dynamic and coordinated epigenetic and physiologic changes occur in the cells of the aggregate suspensions during the C. canephora ISE in liquid system.

Association analysis of polymorphisms rs12997 in ACVR1 and rs1043784 in BMP6 genes involved in bone morphogenic protein signaling pathway in primary angle-closure and pseudoexfoliation glaucoma patients of Saudi origin

BackgroundGlaucoma is a polygenic neurodegenerative disease and the second most common cause of blindness in Saudi Arabia. To test the hypothesis that genetic variants in the genes involved in the bone morphogenic protein (BMP) signaling pathway may be associated with glaucoma, we investigated the association between 3′ untranslated region variants, rs12997 in ACVR1 and rs1043784 in BMP6, and primary angle-closure glaucoma (PACG) and pseudoexfoliation glaucoma (PXG).MethodsIn a case-control study, TaqMan® real-time PCR-based genotyping was done in 444 subjects consisting of 250 controls, 101 PACG and 95 PXG cases, and tested for genetic association with glaucoma-types and other clinical phenotypes.ResultsRs12997[G] allele in ACVR1 exhibited significant 2-fold increased risk of PACG (p = 0.005) in women but not in men. Similarly, genotype analysis also showed that subjects carrying rs12997[G/G] genotype were at > 2-fold risk of PACG that remained significant after adjustment for age, sex, and Bonferroni correction in the recessive model. Furthermore, this effect was also significant in women only. In PXG, the rs12997[G/G] genotype showed a significant trend towards increased risk of the disease (OR = 2.04, 95% CI = 0.99–4.18, p = 0.049) but did not survive the Bonferroni correction. Regression analysis showed that rs12997[G/G] genotype was a significant predictor of PACG independent of age, sex, and rs1043784 genotypes. Likewise, age and rs12997[G/G] genotype showed significant effect on PXG outcome. The rs12997[A/G] genotype showed significant association with cup/disc ratio as compared to wild-type (p = 0.005) in PXG. Genotype and allele frequencies of rs1043784 in BMP6 did not show any significant association either with PACG or PXG.ConclusionsOur results suggest that the polymorphism rs12997 in the ACVR1 gene involved in the BMP signaling pathway is significantly associated with PACG and PXG in a Saudi cohort. This is the first study to associate this variant/gene with PACG and PXG. However, further studies would be needed to replicate these findings in a large population-based cohort.

Transcriptome-Wide Regulation of Key Developmental Pathways in the Mouse Neural Tube by Prenatal Alcohol Exposure.

Early gestational alcohol exposure is associated with severe craniofacial and CNS dysmorphologies and behavioral abnormalities during adolescence and adulthood. Alcohol exposure during the formation of the neural tube (gestational day [GD] 8 to 10 in mice; equivalent to4th week of human pregnancy) disrupts development of ventral midline brain structures such as the pituitary, septum, and ventricles. This study identifies transcriptomic changes in the rostroventral neural tube (RVNT), the region of the neural tube that gives rise to the midline structures sensitive to alcohol exposure during neurulation. Female C57BL/6J mice were administered 2 doses of alcohol (2.9g/kg) or vehicle 4hours apart on GD 9.0. The RVNTs of embryos were collected 6 or 24hours after the first dose and processed for RNA-seq. Six hours following GD 9.0 alcohol exposure (GD 9.25), over 2,300 genes in the RVNT were determined to be differentially regulated by alcohol. Enrichment analysis determined that PAE affected pathways related to cell proliferation, p53 signaling, ribosome biogenesis, and immune activation. In addition, over 100 genes involved in primary cilia formation and function and regulation of morphogenic pathways were altered 6hours after alcohol exposure. The changes to gene expression were largely transient, as only 91 genes identified as differentially regulated by prenatal alcohol at GD 10 (24hours postexposure). Functionally, the differentially regulated genes at GD 10 were related to organogenesis and cell migration. These data give a comprehensive view of the changing landscape of the embryonic transcriptome networks in regions of the neural tube that give rise to brain structures impacted by a neurulation-stage alcohol exposure. Identification of gene networks dysregulated by alcohol will help elucidate the pathogenic mechanisms of alcohol's actions.

Mutual Zonated Interactions of Wnt and Hh Signaling Are Orchestrating the Metabolism of the Adult Liver in Mice and Human

The Hedgehog (Hh) and Wnt/β-Catenin (Wnt) cascades are morphogen pathways whose pronounced influence on adult liver metabolism has been identified in recent years. How both pathways communicate and control liver metabolic functions are largely unknown. Detecting core components of Wnt and Hh signaling and mathematical modeling showed that both pathways in healthy liver act largely complementary to each other in the pericentral (Wnt) and the periportal zone (Hh) and communicate mainly by mutual repression. The Wnt/Hh module inversely controls the spatiotemporal operation of various liver metabolic pathways, as revealed by transcriptome, proteome, and metabolome analyses. Shifting the balance to Wnt (activation) or Hh (inhibition) causes pericentralization and periportalization of liver functions, respectively. Thus, homeostasis of the Wnt/Hh module is essential for maintaining proper liver metabolism and to avoid the development of certain metabolic diseases. With caution due to minor species-specific differences, these conclusions may hold for human liver as well.

Is fibrodysplasia ossificans progressiva an interleukin-1 driven auto-inflammatory syndrome?

BackgroundFibrodysplasia ossificans progressiva (FOP) is the most catastrophic form of heterotopic ossification, due to ongoing intracellular signaling through the bone morphogenic protein pathway. The paroxysmal appearance of inflammatory lumps and elevated inflammatory markers during flares, suggest that FOP is an auto-inflammatory disease. Based on evidence, demonstrating a role for interleukin-1β (IL-1β) in other forms of heterotopic ossification, we hypothesized that treating FOP patients with anti-IL-1 agents could help lower the rate of FOP paroxysms and/or limit the symptoms and residual lesions.Case presentationA 13.5-year-old Arab boy was diagnosed with FOP. Treatment with anti-inflammatory drugs did not change the disease course. New lumps appeared in a rate of approximately one every 8 days. Treatment with the anti-IL-1 agents anakinra and canakinumab resulted in significantly lower rate of paroxysms (every 22–25 days, of which almost all involved only 2 existing lumps), as well as shorter duration. High levels of IL-1β were found in the patient’s plasma samples, collected during a paroxysm that appeared 8 weeks after the last canakinumab dose. In contrast, IL-1β plasma levels were undetectable in the previous three plasma samples, obtained while he was treated with anti-IL-1 agents.ConclusionsOur data demonstrate the efficacy of anti-IL-1 agents in the treatment of a patient with FOP. Results showing the marked increase in IL-1β plasma levels during a paroxysm support a role for IL-1β in the pathogenesis of FOP and further provide the rationale for the use of anti-IL-1 agents in FOP treatment.

Dissecting the dynamics of signaling events in the BMP, WNT, and NODAL cascade during self-organized fate patterning in human gastruloids.

During gastrulation, the pluripotent epiblast self-organizes into the 3 germ layers—endoderm, mesoderm and ectoderm, which eventually form the entire embryo. Decades of research in the mouse embryo have revealed that a signaling cascade involving the Bone Morphogenic Protein (BMP), WNT, and NODAL pathways is necessary for gastrulation. In vivo, WNT and NODAL ligands are expressed near the site of gastrulation in the posterior of the embryo, and knockout of these ligands leads to a failure to gastrulate. These data have led to the prevailing view that a signaling gradient in WNT and NODAL underlies patterning during gastrulation; however, the activities of these pathways in space and time have never been directly observed. In this study, we quantify BMP, WNT, and NODAL signaling dynamics in an in vitro model of human gastrulation. Our data suggest that BMP signaling initiates waves of WNT and NODAL signaling activity that move toward the colony center at a constant rate. Using a simple mathematical model, we show that this wave-like behavior is inconsistent with a reaction-diffusion–based Turing system, indicating that there is no stable signaling gradient of WNT/NODAL. Instead, the final signaling state is homogeneous, and spatial differences arise only from boundary effects. We further show that the durations of WNT and NODAL signaling control mesoderm differentiation, while the duration of BMP signaling controls differentiation of CDX2-positive extra-embryonic cells. The identity of these extra-embryonic cells has been controversial, and we use RNA sequencing (RNA-seq) to obtain their transcriptomes and show that they closely resemble human trophoblast cells in vivo. The domain of BMP signaling is identical to the domain of differentiation of these trophoblast-like cells; however, neither WNT nor NODAL forms a spatial pattern that maps directly to the mesodermal region, suggesting that mesoderm differentiation is controlled dynamically by the combinatorial effect of multiple signals. We synthesize our data into a mathematical model that accurately recapitulates signaling dynamics and predicts cell fate patterning upon chemical and physical perturbations. Taken together, our study shows that the dynamics of signaling events in the BMP, WNT, and NODAL cascade in the absence of a stable signaling gradient control fate patterning of human gastruloids.

Rejuvenation of brain, liver and muscle by simultaneous pharmacological modulation of two signaling determinants, that change in opposite directions with age

We hypothesize that altered intensities of a few morphogenic pathways account for most/all the phenotypes of aging. Investigating this has revealed a novel approach to rejuvenate multiple mammalian tissues by defined pharmacology. Specifically, we pursued the simultaneous youthful in vivo calibration of two determinants: TGF-beta which activates ALK5/pSmad 2,3 and goes up with age, and oxytocin (OT) which activates MAPK and diminishes with age. The dose of Alk5 inhibitor (Alk5i) was reduced by 10-fold and the duration of treatment was shortened (to minimize overt skewing of cell-signaling pathways), yet the positive outcomes were broadened, as compared with our previous studies. Alk5i plus OT quickly and robustly enhanced neurogenesis, reduced neuro-inflammation, improved cognitive performance, and rejuvenated livers and muscle in old mice. Interestingly, the combination also diminished the numbers of cells that express the CDK inhibitor and marker of senescence p16 in vivo. Summarily, simultaneously re-normalizing two pathways that change with age in opposite ways (up vs. down) synergistically reverses multiple symptoms of aging.

Long-term in vitro culture affects phenotypic plasticity of Neoregelia johannis plants

Factors such as stress and prolonged in vitro culture may lead microplants to senescence, reducing and/or altering their morphogenic potential. Considering that plants phenotypic plasticity is improved under stress, this research investigated this physiological event in Neoregelia johannis microplants in long-term in vitro culture. Histological and histochemical analyses were done to monitor the morphogenic potential during the propagules induction by microplants in vitro for 1 and 5 years, and from young leaf used as explants from five subcultures of microplants in both origins. During the monitor of morphogenic pathways, have not been seen alteration for the axillary meristems establishment and development besides that, the microplants multiplication have shown parenchyma cells in the proximal and in shoot bases acting as a pluripotent cell niche to direct adventitious organogenesis (adventitious buds) for young and older cultures. These events were evidenced by amyloplasts and polysaccharides in shoot bases and its reduction along the subcultures to induce adventitious and axillary buds. The culture medium renewal induced stress in microplants as well as the probable aging in older microplants, and it conducted the epidermal and subepidermal cells of older leaves to act as target cells niches for pluripotency and totipotency from the second subculture of leaf explants. All the adventitious roots were originated directly from the meristematic activity of procambial cells and older microplants roots acquired competence to indirect somatic embryogenesis after the second foliar subculture. Therefore, our results indicates that long-term in vitro culture supports new morphogenic competence in microplants, resulting from cell and phenotypic plasticity, that could help in the species survival. Long-term in vitro culture acquired in microplants new morphogenic competences due to the stress caused by the culture medium renewal and its probable aging, resulting to cell and phenotypic plasticity.

Embryogenic competence of microspores is associated with their ability to form a callosic, osmoprotective subintinal layer

Microspore embryogenesis is an experimental morphogenic pathway with important applications in basic research and applied plant breeding, but its genetic, cellular, and molecular bases are poorly understood. We applied a multidisciplinary approach using confocal and electron microscopy, detection of Ca2+, callose, and cellulose, treatments with caffeine, digitonin, and endosidin7, morphometry, qPCR, osmometry, and viability assays in order to study the dynamics of cell wall formation during embryogenesis induction in a high-response rapeseed (Brassica napus) line and two recalcitrant rapeseed and eggplant (Solanum melongena) lines. Formation of a callose-rich subintinal layer (SL) was common to microspore embryogenesis in the different genotypes. However, this process was directly related to embryogenic response, being greater in high-response genotypes. A link could be established between Ca2+ influx, abnormal callose/cellulose deposition, and the genotype-specific embryogenic competence. Callose deposition in inner walls and SLs are independent processes, regulated by different callose synthases. Viability and control of internal osmolality are also related to SL formation. In summary, we identified one of the causes of recalcitrance to embryogenesis induction: a reduced or absent protective SL. In responding genotypes, SLs are markers for changes in cell fate and serve as osmoprotective barriers to increase viability in imbalanced in vitro environments. Genotype-specific differences relate to different responses against abiotic (heat/osmotic) stresses.

Self-Assembly of Embryonic and Two Extraembryonic Stem Cell Types Into Gastrulating Embryo-like Structures

(Abstracted from Nature Cell Biology 2018;20:979–989) Three stem cell types found in the natural embryo differentiate to become 3 distinct tissues: the embryonic tissue or epiblast, the trophectoderm, and the primitive endoderm. Signaling interactions between each of these tissue types are important for morphogenic pathways causing the cell rearrangements of gastrulation and establishing anterior and posterior identity.

Interplay between microRNAs and Wnt, transforming growth factor-β, and bone morphogenic protein signaling pathways promote osteoblastic differentiation of mesenchymal stem cells.

Osteoblasts are terminally differentiated cells with mesenchymal origins, known to possess pivotal roles in sustaining bone microstructure and homeostasis. These cells are implicated in the pathophysiology of various bone disorders, especially osteoporosis. Over the last few decades, strategies to impede bone resorption, principally by bisphosphonates, have been mainstay of treatment of osteoporosis; however, in recent years more attention has been drawn on bone-forming approaches for managing osteoporosis. MicroRNAs (miRNAs) are a broad category of noncoding short sequence RNA fragments that posttranscriptionally regulate the expression of diverse functional and structural genes in a negative manner. An accumulating body of evidence signifies that miRNAs direct mesenchymal stem cells toward osteoblast differentiation and bone formation through bone morphogenic protein, transforming growth factor-β, and Wnt signaling pathways. MiRNAs are regarded as excellent future therapeutic candidates because of their small size and ease of delivery into the cells. Considering their novel therapeutic significance, this review discusses the main miRNAs contributing to the anabolic aspects of bone formation and illustrates their interactions with corresponding signaling pathways involved in osteoblastic differentiation.

In vitro responses in Passiflora species with different chromosome numbers, ploidy levels and nuclear 2C values: revisiting and providing new insights

Tissue culture in Passiflora has emerged as a strategy to propagate species with agronomic relevance, which is the main focus of most in vitro studies. Different morphogenic responses have been obtained under the same environmental in vitro conditions, mainly for species of the subgenus Passiflora with distinct 2n chromosome numbers. The aims of this study were to verify and compare the in vitro responses in Passiflora species with distinct 2n chromosome numbers, ploidy levels and nuclear 2C values. Under the same in vitro conditions, only friable calli occurred from mature zygotic embryo explants of Passiflora coriacea (2n = 2x = 12 chromosomes, 2C = 1.00 pg), Passiflora lindeniana (2n = 4x = 24, 2C = 2.42 pg) and Passiflora contracta (2n = 8x = 48, 2C = 4.78 pg). In contrast, plantlets were regenerated from Passiflora foetida (2n = 20, 2C = 1.04 pg) and Passiflora miniata (2n = 18, 2C = 3.40 pg) via indirect organogenesis and indirect somatic embryogenesis, respectively. By now, from mature zygotic embryo explants, de novo shoot organogenesis and somatic embryos have been recovered for Passiflora species with 2n = 18 chromosomes and relative high nuclear 2C value (more than 2C = 2.93 pg—Passiflora cincinata), and only de novo shoot organogenesis for P. foetida with 2n = 20 chromosomes and relative low 2C value (2C = 1.04 pg). Therefore, in a taxonomic and evolutive context, this study showed that the in vitro morphogenic pathways pretty varied between the Passiflora species with distinct karyotype features.

The NFκB subunit RELA is a master transcriptional regulator of the committed epithelial-mesenchymal transition in airway epithelial cells

The epithelial-mesenchymal transition (EMT) is a multistep dedifferentiation program important in tissue repair. Here, we examined the role of the transcriptional regulator NF-κB in EMT of primary human small airway epithelial cells (hSAECs). Surprisingly, transforming growth factor β (TGFβ) activated NF-κB/RELA proto-oncogene, NF-κB subunit (RELA) translocation within 1 day of stimulation, yet induction of its downstream gene regulatory network occurred only after 3 days. A time course of TGFβ-induced EMT transition was analyzed by RNA-Seq in the absence or presence of inducible shRNA-mediated silencing of RELA. In WT cells, TGFβ stimulation significantly affected the expression of 2,441 genes. Gene set enrichment analysis identified WNT, cadherin, and NF-κB signaling as the most prominent TGFβ-inducible pathways. By comparison, RELA controlled expression of 3,138 overlapping genes mapping to WNT, cadherin, and chemokine signaling pathways. Conducting upstream regulator analysis, we found that RELA controls six clusters of upstream transcription factors, many of which overlapped with a transcription factor topology map of EMT developed earlier. RELA triggered expression of three key EMT pathways: 1) the WNT/β-catenin morphogen pathway, 2) the JUN transcription factor, and 3) the Snail family transcriptional repressor 1 (SNAI1). RELA binding to target genes was confirmed by ChIP. Experiments independently validating WNT dependence on RELA were performed by silencing RELA via genome editing and indicated that TGFβ-induced WNT5B expression and downstream activation of the WNT target AXIN2 are RELA-dependent. We conclude that RELA is a master transcriptional regulator of EMT upstream of WNT morphogen, JUN, SNAI1-ZEB1, and interleukin-6 autocrine loops.