Cell fate determination is a necessary and tightly regulated process for producing different cell types and structures during development. Cranial neural crest cells (CNCCs) are unique to vertebrate embryos and emerge from the neural plate borders into multiple cell lineages that differentiate into bone, cartilage, neurons, and glial cells. We previously reported that Irf6 genetically interacts with Twist1 during CNCC-derived tissue formation. Here, we investigated the mechanistic role of Twist1 and Irf6 at early stages of craniofacial development. Our data indicates that TWIST1 is expressed in endocytic vesicles at the apical surface and interacts with β/δ-CATENINS during neural tube closure, and Irf6 is involved in defining neural fold borders by restricting AP2α expression. Twist1 suppresses Irf6 and other epithelial genes in CNCCs during epithelial-to-mesenchymal transition (EMT) process and cell migration. Conversely, a loss of Twist1 leads to a sustained expression of epithelial and cell adhesion markers in migratory CNCCs. Disruption of TWIST1 phosphorylation in vivo leads to epidermal blebbing, edema, neural tube defects, and CNCC-derived structural abnormalities. Altogether, this study describes an uncharacterized function of mammalian Twist1 and Irf6 in the neural tube and CNCCs and provides new target genes of Twist1 involved in cytoskeletal remodeling. DNA variations within TWIST1 putative enhancers are significantly associated with human facial morphology in a large European cohort.

Auxin response factors (ARFs) are a family of transcription factors within plants that serve to regulate the plant’s response to auxin, a growth initiating hormone. ARFs can either serve as transcription activators or repressors to auxin regulation in the presence of auxin. The majority ARFs have 3 domains: a PB1 protein interaction domain, the low‐complexity middle region, and a DNA binding domain. Major differences between ARF activators and repressors lay in large sequence differences in the middle region, but sequence analysis of the PB1s domains show key differences with implications for their role in transcription regulation. PB1 domains are responsible the interactions necessary for auxin regulation. All ARF PB1 domains are type I/II meaning they contain both a positive and negative residue on opposite sides for electrostatic interactions with copies of themselves or other PB1 domains. The potential for bidentate interaction interfaces complicates the model of auxin response by requiring us to consider oligomerization through the PB1 domain including the potential for hetero‐oligomerizatin between ARF activators and repressors. The work examines the degree of oligomerization and quantifies dimerization affinity in both the wild‐type ARF5 PB1, a transcription activator, and the ARF1 PB1, a transcription repressor. More specifically, to answer these questions crosslinking, gel filtration, and fluorescence anisotropy were used to compare a broad range of concentrations for analysis as well as to improve quantification. Both wild‐type ARF5 and ARF1 self‐associated as expected under varying conditions, forming small oligomers, but ARF5 remained primarily in the dimer state. This work suggests a more nuanced model of transcriptional regulation depending on the specific population of regulatory PB1s present.

A wide variety of common craniofacial birth defects and pediatric cancers arise from errors in the development of cranial neural crest cells (CNCC) that give rise to craniofacial skeletal and peripheral nervous systems. Identification of the genetic and environmental factors that regulate cell fate of CNCCs is crucial for uncovering the causes behind craniofacial disorders. Our lab has identified a dual‐transcription factor pathway that regulates CNCCs during EMT and migration. Initially, we identified a causative mutation in a TWIST1 binding site within an Interferon Regulatory Factor 6 (IRF6) enhancer in a multi‐generational family affected with syndromic cleft lip and palate called Van der Woude syndrome. Our subsequent animal work has shown that Twist1 and Irf6 genetically interact during early craniofacial development, and that the compound heterozygotes exhibit craniofacial abnormalities. We recently demonstrated that Irf6 is required for neural tube and CNCC development and that it interacts with Twist1 during EMT and migration of CNCCs. We also showed that TWIST1 is highly phosphorylated in CNCCs and its phosphorylation is crucial for regulating Irf6 and potentially miR10 family members in CNCC‐derived craniofacial tissues. Our recent in‐vivo and neural tube explants using dual fluorescent cell tracing system showed that Twist1 conditional knockout (CKO) in CNCCs significantly reduced cell delamination and disrupted EMT. The detached Twist1 CKO in CNCCs retained their epithelial signatures and migrated as clusters of epithelial‐like cells over significantly shorter distances. Finally, Twist1 phospho‐incompetent mice for two serine residues were recently generated to determine their impact on TWIST1 activity. Our preliminary results showed that Twist1S68A/S68A phospho‐mutant mice have craniofacial bone defects and forebrain hemorrhage. Support or Funding Information UTHealth start‐up funds and a small grant from theRolanette and Berdon Lawrence Bone Disease Program of Texas

Background Ganoderma lucidum, a valuable medicinal fungus, is widely distributed in China. It grows alongside with a complex microbial ecosystem in the substrate. As sequencing technology advances, it is possible to reveal the composition and functions of substrate-associated bacterial communities. Methods We analyzed the bacterial community dynamics in the substrate during the four typical growth stages of G. lucidum using next-generation sequencing. Results The physicochemical properties of the substrate (e.g. acidity, moisture, total nitrogen, total phosphorus and total potassium) changed between different growth stages. A total of 598,771 sequences from 12 samples were obtained and assigned to 22 bacterial phyla. Proteobacteria and Firmicutes were the dominant phyla. Bacterial community composition and diversity significantly differed between the elongation stage and the other three growth stages. LEfSe analysis revealed a large number of bacterial taxa (e.g. Bacteroidetes, Acidobacteria and Nitrospirae) with significantly higher abundance at the elongation stage. Functional pathway prediction uncovered significant abundance changes of a number of bacterial functional pathways between the elongation stage and other growth stages. At the elongation stage, the abundance of the environmental information processing pathway (mainly membrane transport) decreased, whereas that of the metabolism-related pathways increased. Discussion The changes in bacterial community composition, diversity and predicted functions were most likely related to the changes in the moisture and nutrient conditions in the substrate with the growth of G. lucidum, particularly at the elongation stage. Our findings shed light on the G. lucidum-bacteria-substrate relationships, which should facilitate the industrial cultivation of G. lucidum.

Background Ganoderma lucidum, a valuable medicinal fungus, is widely distributed in China. It grows alongside with a complex microbial ecosystem in the substrate. As sequencing technology advances, it is possible to reveal the composition and functions of substrate-associated bacterial communities. Methods We analyzed the bacterial community dynamics in the substrate during the four typical growth stages of G. lucidum using next-generation sequencing. Results The physicochemical properties of the substrate (e.g. acidity, moisture, total nitrogen, total phosphorus and total potassium) changed between different growth stages. A total of 598,771 sequences from 12 samples were obtained and assigned to 22 bacterial phyla. Proteobacteria and Firmicutes were the dominant phyla. Bacterial community composition and diversity significantly differed between the elongation stage and the other three growth stages. LEfSe analysis revealed a large number of bacterial taxa (e.g. Bacteroidetes, Acidobacteria and Nitrospirae) with significantly higher abundance at the elongation stage. Functional pathway prediction uncovered significant abundance changes of a number of bacterial functional pathways between the elongation stage and other growth stages. At the elongation stage, the abundance of the environmental information processing pathway (mainly membrane transport) decreased, whereas that of the metabolism-related pathways increased. Discussion The changes in bacterial community composition, diversity and predicted functions were most likely related to the changes in the moisture and nutrient conditions in the substrate with the growth of G. lucidum, particularly at the elongation stage. Our findings shed light on the G. lucidum-bacteria-substrate relationships, which should facilitate the industrial cultivation of G. lucidum.