scholarly journals Transcriptomic profiling of mTOR and ryanodine receptor signaling molecules in developing zebrafish in the absence and presence of PCB 95

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e4106 ◽  
Author(s):  
Daniel F. Frank ◽  
Galen W. Miller ◽  
Richard E. Connon ◽  
Juergen Geist ◽  
Pamela J. Lein
Keyword(s):  
Signaling Pathways ◽  
Ryanodine Receptor ◽  
Expression Patterns ◽  
Signaling Molecules ◽  
Transcriptomic Profiling ◽  
Downstream Target ◽  
Spatiotemporal Expression ◽  
Waterborne Exposure

The mechanistic target of rapamycin (mTOR) and ryanodine receptor (RyR) signaling pathways regulate fundamental processes of neurodevelopment, and genetic mutations within these pathways have been linked to neurodevelopmental disorders. While previous studies have established that these signaling molecules are expressed in developing zebrafish, a detailed characterization of the ontogenetic profile of these signaling molecules is lacking. Thus, we evaluated the spatiotemporal expression of key transcripts in mTOR and RyR signaling pathways in wildtype zebrafish at 24, 72 and 120 hours post fertilization (hpf). We further determined whether transcriptional profiles of a subset of genes in both pathways were altered by exposure to PCB 95 (2,2′,3,5′,6-pentachlorobiphenyl), a pervasive environmental contaminant known to cause developmental neurotoxicity in mammalian systems via RyR-dependent mechanisms. Quantitative PCR revealed that transcription generally increased across development. Genes in the signaling pathway upstream of the mTORC1 complex, and the RyR-paralogs, ryr2a and ryr3, were robustly upregulated, and in situ hybridization of ryr3 coincided with a transcriptional shift from muscle to neuronal tissue after 24 hpf. Static waterborne exposure to PCB 95 beginning at 6 hpf significantly altered transcription of genes in both pathways. These changes were concentration- and time-dependent, and included downregulation of rptor, a member of the mTORC1 complex, at both 72 and 120 hpf, and increased transcript levels of the RyR paralog ryr2b and downstream target of RyR signaling, Wingless-type 2ba (wnt2ba) at 72 hpf. The detailed transcriptomic profiling of key genes within these two signaling pathways provides a baseline for identifying other environmental factors that modify normal spatiotemporal expression patterns of mTOR and RyR signaling pathways in the developing zebrafish, as illustrated here for PCB 95.

scholarly journals Spatiotemporal Expression of Anticoagulation Factor Antistasin in Freshwater Leeches

2019 ◽  
Vol 20 (16) ◽  
pp. 3994 ◽  
Author(s):  
Hee-Jin Kwak ◽  
Jeong-Su Park ◽  
Brenda Irene Medina Jiménez ◽  
Soon Cheol Park ◽  
Sung-Jin Cho
Keyword(s):  
Hedgehog Signaling ◽  
Signal Transduction Pathway ◽  
The Body ◽  
Spatial Expression ◽  
Downstream Target ◽  
Spatiotemporal Expression ◽  
Stage 4 ◽  
Segment Polarity ◽  
Late Stages

Antistasin, which was originally discovered in the salivary glands of the Mexican leech Haementeria officinalis, was newly isolated from Helobdella austinensis. To confirm the temporal expression of antistasin during embryogenesis, we carried out semi-quantitative RT-PCR. Hau-antistasin1 was uniquely expressed at stage 4 of the cleavage and was strongly expressed in the late stages of organogenesis, as were other antistasin members. In order to confirm the spatial expression of antistasin, we performed fluorescence in situ hybridization in the late stages of organogenesis. The expression of each antistasin in the proboscis showed a similar pattern and varied in expression in the body. In addition, the spatial expression of antistasin orthologs in different leeches showed the possibility of different function across leech species. Hau-antistasin1 was expressed in the same region as hedgehog, which is a known mediator of signal transduction pathway. Hau-antistasin1 is probably a downstream target of Hedgehog signaling, involved in segment polarity signal pathway.

scholarly journals Expression and Function Studies of CYC/TB1-Like Genes in the Asymmetric Flower Canna (Cannaceae, Zingiberales)

2020 ◽  
Vol 11 ◽  
Author(s):  
Qianxia Yu ◽  
Xueyi Tian ◽  
Canjia Lin ◽  
Chelsea D. Specht ◽  
Jingping Liao
Keyword(s):  
Flower Development ◽  
Expression Patterns ◽  
Asymmetric Distribution ◽  
Inflorescence Architecture ◽  
Flower Primordia ◽  
Fertile Stamen ◽  
Spatiotemporal Expression ◽  
And Function

The asymmetric flower, lacking any plane of symmetry, is rare among angiosperms. Canna indica L. has conspicuously asymmetric flowers resulting from the presence of a half-fertile stamen, while the other androecial members develop as petaloid staminodes or abort early during development. The molecular basis of the asymmetric distribution of fertility and petaloidy in the androecial whorls remains unknown. Ontogenetic studies have shown that Canna flowers are borne on monochasial (cincinnus) partial florescences within a racemose inflorescence, with floral asymmetry likely corresponding to the inflorescence architecture. Given the hypothesized role of CYC/TB1 genes in establishing floral symmetry in response to the influence of the underlying inflorescence architecture, the spatiotemporal expression patterns of three Canna CYC/TB1 homologs (CiTBL1a, CiTBL1b-1, and CiTBL1b-2) were analyzed during inflorescence and floral development using RNA in situ hybridization and qRT-PCR. In the young inflorescence, both CiTBL1a and CiTBL1b-1 were found to be expressed in the bracts and at the base of the lateral florescence branches, whereas transcripts of CiTBL1b-2 were mainly detected in flower primordia and inflorescence primordia. During early flower development, expression of CiTBL1a and CiTBL1b-1 were both restricted to the developing sepals and petals. In later flower development, expression of CiTBL1a was reduced to a very low level while CiTBL1b-1 was detected with extremely high expression levels in the petaloid androecial structures including the petaloid staminodes, the labellum, and the petaloid appendage of the fertile stamen. In contrast, expression of CiTBL1b-2 was strongest in the fertile stamen throughout flower development, from early initiation of the stamen primordium to maturity of the ½ anther. Heterologous overexpression of CiTBL genes in Arabidopsis led to dwarf plants with smaller petals and fewer stamens, and altered the symmetry of mature flowers. These data provide evidence for the involvement of CYC/TB1 homologs in the development of the asymmetric Cannaceae flower.

scholarly journals Patterns of gene expression in schistosomes: localization by whole mountin situhybridization

Parasitology ◽  
2007 ◽  
Vol 134 (11) ◽  
pp. 1589-1597 ◽  
Author(s):  
G. P. DILLON ◽  
J. C. ILLES ◽  
H. V. ISAACS ◽  
R. A. WILSON
Keyword(s):  
Gene Expression ◽  
Cathepsin L ◽  
Expression Patterns ◽  
Proteinase K ◽  
Fluorescent Substrate ◽  
Rna Probes ◽  
Fast Red ◽  
Proteinase K Treatment

SUMMARYAs a consequence of comprehensive transcriptome analysis followed by sequencing and draft assembly of the genome, the emphasis of schistosome research is shifting from the identification of genes to the characterization of their functions and interactions. Developmental biologists have long used whole mountin situhybridization (WISH) to determine gene expression patterns, as a vital tool for formulating and testing hypotheses about function. This paper describes the application of WISH to the study of gene expression in larval and adult schistosomes. Fixed worms were permeablized by proteinase K treatment for hybridization with digoxygenin-labelled RNA probes, with binding being detected by alkaline phosphatase-coupled anti-digoxygenin antibodies, and BM Purple substrate. Discrete staining patterns for the transcripts of the molecules Sm29, cathepsin L, antigen 10.3 and chorion were observed in the tegument cell bodies, gut epithelium, oesophageal gland and vitelline lobules, respectively, of adult worms. Transcripts of the molecules SGTP4, GP18-22 and cathepsin L were localized to tegument cell bodies and embryonic gut, respectively, of lung schistosomula. We also showed that Fast Red TR fluorescent substrate can refine the pattern of localization permitting use of confocal microscopy. We believe that method of WISH will find broad application, in synergy with other emerging post-genomic techniques, such as RNA interference, to studies focused at increasing our molecular understanding of schistosomes.

scholarly journals Spatiotemporal characterization of periocular mesenchyme heterogeneity during anterior segment development

2019 ◽  
Author(s):  
Kristyn L. Van Der Meulen ◽  
Oliver Vöcking ◽  
Megan L. Weaver ◽  
Jakub K. Famulski
Keyword(s):  
Expression Patterns ◽  
Anterior Segment ◽  
Critical Event ◽  
Transgenic Zebrafish ◽  
Anterior Segment Dysgenesis ◽  
Heterogenous Population ◽  
First Time

ABSTRACTEstablishment of the ocular anterior segment (AS) is a critical event during development of the vertebrate visual system. Failure in this process leads to Anterior Segment Dysgenesis (ASD), which is characterized by congenital blindness and predisposition to glaucoma. The anterior segment is largely formed via a neural crest-derived population, the Periocular Mesenchyme (POM). In this study, we aimed to characterize POM behaviors and identities during zebrafish AS development. POM distributions and migratory dynamics were analyzed using transgenic zebrafish embryos (Tg[foxC1b:GFP], Tg[foxD3:GFP], Tg[pitx2:GFP], Tg[lmx1b.1:GFP], and Tg[sox10:GFP] throughout the course of early AS development (24-72hpf). In vivo imaging analysis revealed unique AS distribution and migratory behavior among the reporter lines, suggesting AS mesenchyme (ASM) is a heterogenous population. This was confirmed using double in situ hybridization. Furthermore, we generated ASM transcriptomic profiles from our reporter lines and using a four-way comparison analysis uncovered unique ASM subpopulation expression patterns. Taken together, our data reveal for the first time that AS-associated POM is not homogeneous but rather comprised of several unique subpopulations identifiable by their distributions, behaviors, and transcriptomic profiles.

scholarly journals Functional conservation of erythropoietin signaling in zebrafish

Blood ◽  
2007 ◽  
Vol 110 (7) ◽  
pp. 2718-2726 ◽  
Author(s):  
Noëlle Paffett-Lugassy ◽  
Nelson Hsia ◽  
Paula G. Fraenkel ◽  
Barry Paw ◽  
Irene Leshinsky ◽  
...  
Keyword(s):  
Expression Patterns ◽  
Functional Characterization ◽  
Erythroid Progenitor ◽  
Mature Erythrocyte ◽  
Functional Conservation ◽  
Hypoxic Conditions ◽  
Definitive Erythropoiesis ◽  
Mammalian Gene Expression

Erythropoietin (Epo) and its cognate receptor (EpoR) are required for maintaining adequate levels of circulating erythrocytes during embryogenesis and adulthood. Here, we report the functional characterization of the zebrafish epo and epor genes. The expression of epo and epor was evaluated by quantitative reverse transcriptase–polymerase chain reaction (RT-PCR) and whole-mount in situ hybridization, revealing marked parallels between zebrafish and mammalian gene expression patterns. Examination of the hypochromic mutant, weissherbst, and adult hypoxia-treated hearts indicate that zebrafish epo expression is induced by anemia and hypoxia. Overexpression of epo mRNA resulted in severe polycythemia, characterized by a striking increase in the number of cells expressing scl, c-myb, gata1, ikaros, epor, and βe1-globin, suggesting that both the erythroid progenitor and mature erythrocyte compartments respond to epo. Morpholino-mediated knockdown of the epor caused a slight decrease in primitive and complete block of definitive erythropoiesis. Abrogation of STAT5 blocked the erythropoietic expansion by epo mRNA, consistent with a requirement for STAT5 in epo signaling. Together, the characterization of zebrafish epo and epor demonstrates the conservation of an ancient program that ensures proper red blood cell numbers during normal homeostasis and under hypoxic conditions.

scholarly journals Sox Genes Show Spatiotemporal Expression during Murine Tongue and Eyelid Development

2018 ◽  
Vol 2018 ◽  
pp. 1-13 ◽  
Author(s):  
Ryuichi Ishikawa ◽  
Maiko Kawasaki ◽  
Katsushige Kawasaki ◽  
Akane Yamada ◽  
Supaluk Trakanant ◽  
...  
Keyword(s):  
Expression Pattern ◽  
Expression Patterns ◽  
Family Members ◽  
Biological Processes ◽  
Tongue Muscle ◽  
Spatiotemporal Expression ◽  
Critical Organ ◽  
Eyelid Development ◽  
Sox Genes

The tongue is a critical organ, involved in functions such as speaking, swallowing, mastication, and degustation. Although Sox genes are known to play critical roles in many biological processes, including organogenesis, the expression of the Sox family members during tongue development remains unclear. We therefore performed a comparative in situ hybridization analysis of 17 Sox genes (Sox1–14, 17, 18, and 21) during murine tongue development. Sox2, 4, 6, 8, 9, 10, 11, 12, and 21 were found to be expressed in the tongue epithelium, whereas Sox2, 4–6, 8–11, 13, and 21 showed expression in the mesenchyme of the developing tongue. Expression of Sox1, 4, 6, 8–12, and 21 were observed in the developing tongue muscle. Sox5 and 13 showed expression only at E12, while Sox1 expression was observed only on E18. Sox6, 8, 9, and 12 showed expression at several stages. Although the expression of Sox2, 4, 10, 11, and 21 was detected during all the four stages of tongue development, their expression patterns differed among the stages. We thus identified a dynamic spatiotemporal expression pattern of the Sox genes during murine tongue development. To understand whether Sox genes are involved in the development of other craniofacial organs through similar roles to those in tongue development, we also examined the expression of Sox genes in eyelid primordia, which also contain epithelium, mesenchyme, and muscle. However, expression patterns and timing of Sox genes differed between tongue and eyelid development. Sox genes are thus related to organogenesis through different functions in each craniofacial organ.

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