Circadian key component CLOCK/BMAL1 interferes with segmentation clock in mouse embryonic organoids

In mammals, circadian clocks are strictly suppressed during early embryonic stages, as well as in pluripotent stem cells, by the lack of CLOCK/BMAL1-mediated circadian feedback loops. During ontogenesis, the innate circadian clocks emerge gradually at a late developmental stage, and with these, the circadian temporal order is invested in each cell level throughout a body. Meanwhile, in the early developmental stage, a segmented body plan is essential for an intact developmental process, and somitogenesis is controlled by another cell-autonomous oscillator, the segmentation clock, in the posterior presomitic mesoderm (PSM). In the present study, focusing upon the interaction between circadian key components and the segmentation clock, we investigated the effect of the CLOCK/BMAL1 on the segmentation clock Hes7 oscillation, revealing that the expression of functional CLOCK/BMAL1 severely interferes with the ultradian rhythm of segmentation clock in induced PSM and gastruloids. RNA sequencing analysis implied that the premature expression of CLOCK/BMAL1 affects the Hes7 transcription and its regulatory pathways. These results suggest that the suppression of CLOCK/BMAL1-mediated transcriptional regulation during the somitogenesis may be inevitable for intact mammalian development.

Comparative transcriptomic analysis reveals key components controlling spathe color in Anthurium andraeanum (Hort.)

Anthurium andraeanum (Hort.) is an important ornamental in the tropical cut-flower industry. However, there is currently insufficient information to establish a clear connection between the genetic model(s) proposed and the putative genes involved in the differentiation between colors. In this study, 18 cDNA libraries related to the spathe color and developmental stages of A. andraeanum were characterized by transcriptome sequencing (RNA-seq). For the de novo transcriptome, a total of 114,334,082 primary sequence reads were obtained from the Illumina sequencer and were assembled into 151,652 unigenes. Approximately 58,476 transcripts were generated and used for comparative transcriptome analysis between three cultivars that differ in spathe color (‘Sasha’ (white), ‘Honduras’ (red), and ‘Rapido’ (purple)). A large number of differentially expressed genes (8,324), potentially involved in multiple biological and metabolic pathways, were identified, including genes in the flavonoid and anthocyanin biosynthetic pathways. Our results showed that the chalcone isomerase (CHI) gene presented the strongest evidence for an association with differences in color and the highest correlation with other key genes (flavanone 3-hydroxylase (F3H), flavonoid 3’5’ hydroxylase (F3’5’H)/ flavonoid 3’-hydroxylase (F3’H), and leucoanthocyanidin dioxygenase (LDOX)) in the anthocyanin pathway. We also identified a differentially expressed cytochrome P450 gene in the late developmental stage of the purple spathe that appeared to determine the difference between the red- and purple-colored spathes. Furthermore, transcription factors related to putative MYB-domain protein that may control anthocyanin pathway were identified through a weighted gene co-expression network analysis (WGCNA). The results provided basic sequence information for future research on spathe color, which have important implications for this ornamental breeding strategies.

Cre-Activation in ErbB4-Positive Neurons of Floxed Grin1/NMDA Receptor Mice Is Not Associated With Major Behavioral Impairment

Extensive evidence suggests a dysfunction of the glutamate NMDA receptor (NMDAR) in schizophrenia, a severe psychiatric disorder with putative early neurodevelopmental origins, but clinical onset mainly during late adolescence. On the other hand, pharmacological models using NMDAR antagonists and the clinical manifestation of anti-NMDAR encephalitis indicate that NMDAR blockade/hypofunction can trigger psychosis also at adult stages, without any early developmental dysfunction. Previous genetic models of NMDAR hypofunction restricted to parvalbumin-positive interneurons indicate the necessity of an early postnatal impairment to trigger schizophrenia-like abnormalities, whereas the cellular substrates of NMDAR-mediated psychosis at adolescent/adult stages are unknown. Neuregulin 1 (NRG1) and its receptor ErbB4 represent schizophrenia-associated susceptibility factors that closely interact with NMDAR. To determine the neuronal populations implicated in “late” NMDAR-driven psychosis, we analyzed the effect of the inducible ablation of NMDARs in ErbB4-expressing cells in mice during late adolescence using a pharmacogenetic approach. Interestingly, the tamoxifen-inducible NMDAR deletion during this late developmental stage did not induce behavioral alterations resembling depression, schizophrenia or anxiety. Our data indicate that post-adolescent NMDAR deletion, even in a wider cell population than parvalbumin-positive interneurons, is also not sufficient to generate behavioral abnormalities resembling psychiatric disorders. Other neuronal substrates that have to be revealed by future studies, may underlie post-adolescent NMDAR-driven psychosis.

Increasing plant density increases Bt toxin concentration of boll wall in cotton by decreasing boll setting speed

Background In order to uncover the mechanism of significantly reduced insect resistance at the late developmental stage in cotton (Gossypium hirsutum L.), the relationship between boll setting rate under different planting densities and Bacillus thuringiensis (Bt) insecticidal concentrations in the boll wall were investigated in the present study. Two studies were arranged at Yangzhou, China during the 2017–2018 cotton growth seasons. Five planting densities (15 000, 25 000, 45 000, 60 000 and 75 000 plants per hectare) and the flower-removal treatment were imposed separately on Bt cotton cultivar Sikang3 to arrange different boll setting rates, and the boll setting rates and Bt toxin content were compared. Results Higher boll setting rate together with lower Bt toxin contents in boll wall was observed under low planting density, whereas lower boll setting rate and higher Bt toxin contents were found under high planting density. Also, higher Bt protein concentration was associated with higher soluble protein content, glutamic-pyruvic transaminase (GPT), and glutamic oxaloacetate transaminase (GOT) activities, but lower amino acid content, and protease and peptidase activities. It was further confirmed that a higher boll setting rate with lower Bt protein content under flower-removal. Conclusions This study demonstrated that the insecticidal efficacy of boll walls was significantly impacted by boll formation. Reduced protein synthesis and enhanced protein degradation were related to the reduced Bt toxin concentration.

Ediacaran algal cysts from the Doushantuo Formation, South China

Early-middle Ediacaran organic-walled microfossils from the Doushantuo Formation studied in several sections in the Yangtze Gorges area, South China, show ornamented cyst-like vesicles of very high diversity. These microfossils are diagenetically permineralized and observed in petrographic thin-sections of chert nodules. Exquisitely preserved specimens belonging to seven species of Appendisphaera, Mengeosphaera, Tanarium, Urasphaera and Tianzhushania contain either single or multiple spheroidal internal bodies inside the vesicles. These structures indicate reproductive stages, endocyst and dividing cells, respectively, and are preserved at early to late ontogenetic stages in the same taxa. This new evidence supports the algal affiliations for the studied taxa and refutes previous suggestions of Tianzhushania being animal embryo or holozoan. The first record of a late developmental stage of a completely preserved specimen of T. spinosa observed in thin-section demonstrates the interior of vesicles with clusters of identical cells but without any cavity that is diagnostic for recognizing algal cysts vs animal diapause cysts. Various lines of evidence to infer biological affinities of these microfossils – morphology, reproductive characters, spatial arrangement of cells, and biochemical properties of the vesicle wall – are collectively characteristic of algal clades. Recognizing the biological affinities of these microfossils is key to understanding whether animals capable of producing such morphologically complex diapause cysts had an early Ediacaran fossil record (633–610 Ma), or the microfossils were non-animal holozoans or algae as argued herein for Tianzhushania spinosa and other studied microfossils.

Differentiation and localization of interneurons in the developing spinal cord depends on DOT1L expression

Genetic and epigenetic factors contribute to the development of the spinal cord. Failure in correct exertion of the developmental programs, including neurulation, neural tube closure and neurogenesis of the diverse spinal cord neuronal subtypes results in clinical phenotypes with variable severity. The histone methyltransferase Disruptor of Telomeric 1 Like (DOT1L), which mediates histone H3 lysine 79 (H3K79) methylation, is fundamental for proper development of the cerebral cortex and cerebellum, and here we report on its essential role for development of the spinal cord. Conditional inactivation of DOT1L using Wnt1-cre as driver in the developing murine spinal cord did not result in neural tube closure defect (NTCD). Transcriptome analysis revealed that DOT1L deficiency favored differentiation over progenitor proliferation. Dot1l -cKO mainly decreased the numbers of dI1 interneurons expressing Lhx2 . Loss of DOT1L affected localization but not generation of dI2, dI3, and dI5 interneurons. The resulting derailed interneuron patterns might be responsible for increased cell death that occurred at the late developmental stage E18.5. Together our data indicate that DOT1L is essential for subtype- specific neurogenesis, migration and localization of interneurons in the developing spinal cord, in part by regulating transcriptional activation of Lhx2 .

Differentiation and localization of interneurons in the developing spinal cord depends on DOT1L expression

Genetic and epigenetic factors contribute to the development of the spinal cord. Failure in correct exertion of the developmental programs, including neurulation, neural tube closure and neurogenesis of the diverse spinal cord neuronal subtypes results in clinical phenotypes with variable severity. The histone methyltransferase Disruptor of Telomeric 1 Like (DOT1L), which mediates histone H3 lysine 79 (H3K79) methylation, is fundamental for proper development of the cerebral cortex and cerebellum, and here we report on its essential role for development of the spinal cord. Conditional inactivation of DOT1L using Wnt1-cre as driver in the developing murine spinal cord did not result in neural tube closure defect (NTCD). Transcriptome analysis revealed that DOT1L deficiency favored differentiation over progenitor proliferation. Dot1l -cKO mainly decreased the numbers of dI1 interneurons expressing Lhx2 . Loss of DOT1L affected localization but not generation of dI2, dI3, and dI5 interneurons. The resulting derailed interneuron patterns might be responsible for increased cell death that occurred at the late developmental stage E18.5. Together our data indicate that DOT1L is essential for subtype- specific neurogenesis, migration and localization of interneurons in the developing spinal cord, in part by regulating transcriptional activation of Lhx2 .

Identification of Drought Tolerant Mutant Lines of TEF [ERAGROSTIS TEF (ZUCC.) TROTTER]

Introduction: Tef [Eragrostis tef (Zucc.) Trotter] is the major food crop in Ethiopia where it is annually cultivated on about 30% of the total area allocated to cereals. The productivity of tef is, however, significantly reduced by drought which can occur at either the early or the late developmental stage of the crop. Methodology: A total of 10,000 EMS (ethyl methane sulfonate) mutagenized M2 tef populations were screened first for early and second for terminal drought tolerance. The performance of candidate lines was investigated using diverse agronomical and physiological parameters. Results and discussion: Several promising lines were obtained for both early and terminal drought tolerance. Among these, genotypes with outstanding performance were found for two drought tolerant tef lines targeting early drought tolerance (dtt2 and dtt13) and three terminal drought tolerant lines targeting terminal drought tolerance (tdt9, tdt15 and tdt19). Conclusions: Candidate lines for both early and terminal drought tolerance will play vital roles in deve- loping drought tolerance in tef improvement. KEY WORDS: DROUGHT TOLERANCE, ERAGROSTIS TEF, ORPHAN CROP, MOISTURE SCARCITY, TEF

Genetic diversity, functional properties and expression analysis of NnSBE genes involved in starch synthesis of lotus (Nelumbo nucifera Gaertn.)

Background Starch branching enzyme (SBE) is one of the key enzymes in starch biosynthetic metabolism, determining amylopectin structure. Methods Full length coding sequences (CDS) of SBE genes were cloned using reverse transcription PCR (RT-PCR) technology, and neighbor-joining (NJ) tree was used for phylogenetic analysis. Single nucleotide polymorphisms (SNPs) were determined to assess the genetic polymorphisms and variation indexes between individuals and clusters. Quantitative real time PCR (qRT-PCR) was performed to analyze the spatial and temporal expression of NnSBE genes. The effect of NnSBE genes on amylopectin’s fine structures was explored using affinity and the enzyme activity analysis of two isoforms in amylopectin and amylose. Results In this study, two SBE family genes, NnSBEI and NnSBEIII, were identified in lotus (Nelumbo nucifera Gaertn.). Phylogenetic analysis sorted NnSBEI into SBE family B and NnSBEIII into SBE family A. UPGMA phylogenetic tree divided 45 individuals of lotus into three classes. The homozygous haplotype (A G G A G) of NnSBEIII was observed in seed lotus. During the seed embryo development stage, NnSBEIII reached the peak in the middle of the development stage, while NnSBEI increased in the mid-late developmental stage. The different affinity activity of the two isozymes binding amylopectin and amylose assay indicated NnSBEI has higher activity and wider affinity. Discussion Genetic diversity showed that NnSBE genes received artificial selection during the process of cultivation and domestication in lotus seeds. Furthermore, the expression pattern and affinity activity analysis indicated that NnSBE genes were related to the chain length of amylopectin.

Genetic diversity, functional properties and expression analysis of NnSBE genes involved in starch synthesis of lotus (Nelumbo nucifera Gaertn.)

Background. Starch branching enzyme (SBE) is one of the key enzymes in starch biosynthetic metabolism and determines the structure of amylopectin. Methods. Full length of SBE genes were cloned by reverse transcription PCR (RT-PCR) technology, and used neighbor-joining (NJ) tree for phylogenetic analysis. The single nucleotide polymorphisms (SNPs) were determined to assess the genetic polymorphisms and variation indexes between individuals and clusters. Quantitative real time PCR (qRT-PCR) was performed to analyse spatial and temporal expression of SBE genes. Effect of NnSBE genes on fine structure of amylopectin was explore by affinity and enzyme activity analysis of two isoforms in amylopectin and amylose. Results. In this study, two SBE family genes, NnSBEI and NnSBEIII were identified from lotus (Nelumbo nucifera Gaertn.). Phylogenetic analysis divided the NnSBEI into SBE family B and NnSBEIII was belong to SBE family A. The homozygous haplotype (AA GG GG AA GG) of NnSBEIII was observed in seed lotus. During the seed embryo development stage, NnSBEIII reached the peak in middle of development stage and while NnSBEI increased in mid-late developmental stage. The different affinity activity of two isozymes bind amylopectin and amylose assay indicated NnSBEI has higher activity and wider affinity. Discussion. This study showed that NnSBE genes received artificial selection during the process of cultivation and domestication in seed lotus especially. Furthermore, the expression pattern and affinity activity analysis indicated that NnSBE genes was related to chain length of amylopectin.