The Circadian Clock of the Unicellular Eukaryotic Model Organism Chlamydomonas reinhardtii

2003 ◽  
Vol 384 (5) ◽  
pp. 689-695 ◽  
Author(s):  
M. Mittag ◽  
V. Wagner
Keyword(s):  
Circadian Clock ◽  
Chlamydomonas Reinhardtii ◽  
Rna Binding ◽  
Outer Space ◽  
Expression Patterns ◽  
Model Organism ◽  
Nuclear Genome ◽  
Circadian System ◽  
Transcriptional Level ◽  
Circadian Expression

Abstract The green unicellular alga Chlamydomonas reinhardtii, also called 'green yeast', emerged in the past years as a model organism for specific scientific questions such as chloroplast biogenesis and function, the composition of the flagella including its basal apparatus, or the mechanism of the circadian clock. Sequencing of its chloroplast and mitochondrial genomes have already been completed and a first draft of its nuclear genome has also been released recently. In C. reinhardtii several circadian rhythms are physiologically well characterized, and one of them has even been shown to operate in outer space. Circadian expression patterns of nuclear and plastid genes have been studied. The mode of regulation of these genes occurs at the transcriptional level, although there is also evidence for posttranscriptional control. A clock-controlled, phylogenetically conserved RNA-binding protein was characterized in this alga, which interacts with several mRNAs that all contain a common cis-acting motif. Its function within the circadian system is currently under investigation. This review summarizes the current state of the knowledge about the circadian system in C. reinhardtii and points out its potential for future studies.

scholarly journals Functional Development of the Circadian Clock in the Zebrafish Pineal Gland

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Zohar Ben-Moshe ◽  
Nicholas S. Foulkes ◽  
Yoav Gothilf
Keyword(s):  
Pineal Gland ◽  
Circadian Clock ◽  
Rapid Development ◽  
Model Organism ◽  
Circadian System ◽  
Zebrafish Model ◽  
Peripheral Oscillators ◽  
Clock Gene Expression ◽  
Functional Development ◽  
Central Clock

The zebrafish constitutes a powerful model organism with unique advantages for investigating the vertebrate circadian timing system and its regulation by light. In particular, the remarkably early and rapid development of the zebrafish circadian system has facilitated exploring the factors that control the onset of circadian clock function during embryogenesis. Here, we review our understanding of the molecular basis underlying functional development of the central clock in the zebrafish pineal gland. Furthermore, we examine how the directly light-entrainable clocks in zebrafish cell lines have facilitated unravelling the general mechanisms underlying light-induced clock gene expression. Finally, we summarize how analysis of the light-induced transcriptome and miRNome of the zebrafish pineal gland has provided insight into the regulation of the circadian system by light, including the involvement of microRNAs in shaping the kinetics of light- and clock-regulated mRNA expression. The relative contributions of the pineal gland central clock and the distributed peripheral oscillators to the synchronization of circadian rhythms at the whole animal level are a crucial question that still remains to be elucidated in the zebrafish model.

scholarly journals Efficient Editing of the Nuclear APT Reporter Gene in Chlamydomonas reinhardtii via Expression of a CRISPR-Cas9 Module

2019 ◽  
Vol 20 (5) ◽  
pp. 1247 ◽  
Author(s):  
Daniel Guzmán-Zapata ◽  
José Sandoval-Vargas ◽  
Karla Macedo-Osorio ◽  
Edgar Salgado-Manjarrez ◽  
José Castrejón-Flores ◽  
...  
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Transient Expression ◽  
Mammalian Cells ◽  
High Efficiency ◽  
Model Organism ◽  
Nuclear Genome ◽  
Single Copy ◽  
Point Of View ◽  
Selection Marker ◽  
Gene Encoding

The clustered regularly interspaced short palindromic repeat/CRISPR-associated protein 9 (CRISPR/Cas9) technology is a versatile and useful tool to perform genome editing in different organisms ranging from bacteria and yeast to plants and mammalian cells. For a couple of years, it was believed that the system was inefficient and toxic in the alga Chlamydomonas reinhardtii. However, recently the system has been successfully implemented in this model organism, albeit relying mostly on the electroporation of ribonucleoproteins (RNPs) into cell wall deficient strains. This requires a constant source of RNPs and limits the application of the technology to strains that are not necessarily the most relevant from a biotechnological point of view. Here, we show that transient expression of the Streptococcus pyogenes Cas9 gene and sgRNAs, targeted to the single-copy nuclear apt9 gene, encoding an adenine phosphoribosyl transferase (APT), results in efficient disruption at the expected locus. Introduction of indels to the apt9 locus results in cell insensitivity to the otherwise toxic compound 2-fluoroadenine (2-FA). We have used agitation with glass beads and particle bombardment to introduce the plasmids carrying the coding sequences for Cas9 and the sgRNAs in a cell-walled strain of C. reinhardtii (CC-125). Using sgRNAs targeting exons 1 and 3 of apt9, we obtained disruption efficiencies of 3 and 30% on preselected 2-FA resistant colonies, respectively. Our results show that transient expression of Cas9 and a sgRNA can be used for editing of the nuclear genome inexpensively and at high efficiency. Targeting of the APT gene could potentially be used as a pre-selection marker for multiplexed editing or disruption of genes of interest.

scholarly journals Real-Time Monitoring of Chloroplast Gene Expression by a Luciferase Reporter: Evidence for Nuclear Regulation of Chloroplast Circadian Period

2006 ◽  
Vol 26 (3) ◽  
pp. 863-870 ◽  
Author(s):  
Takuya Matsuo ◽  
Kiyoshi Onai ◽  
Kazuhisa Okamoto ◽  
Jun Minagawa ◽  
Masahiro Ishiura
Keyword(s):  
Gene Expression ◽  
Circadian Clock ◽  
Real Time ◽  
Nuclear Genome ◽  
Firefly Luciferase ◽  
Luciferase Reporter ◽  
Chloroplast Gene ◽  
Chloroplast Gene Expression ◽  
Period Gene ◽  
Circadian Expression

ABSTRACT Chloroplast-encoded genes, like nucleus-encoded genes, exhibit circadian expression. How the circadian clock exerts its control over chloroplast gene expression, however, is poorly understood. To facilitate the study of chloroplast circadian gene expression, we developed a codon-optimized firefly luciferase gene for the chloroplast of Chlamydomonas reinhardtii as a real-time bioluminescence reporter and introduced it into the chloroplast genome. The bioluminescence of the reporter strain correlated well with the circadian expression pattern of the introduced gene and satisfied all three criteria for circadian rhythms. Moreover, the period of the rhythm was lengthened in per mutants, which are phototactic rhythm mutants carrying a long-period gene in their nuclear genome. These results demonstrate that chloroplast gene expression rhythm is a bona fide circadian rhythm and that the nucleus-encoded circadian oscillator determines the period length of the chloroplast rhythm. Our reporter strains can serve as a powerful tool not only for analysis of the circadian regulation mechanisms of chloroplast gene expression but also for a genetic approach to the molecular oscillator of the algal circadian clock.

scholarly journals Circadian RNA expression elicited by 3’-UTR IRAlu-paraspeckle associated elements

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Manon Torres ◽  
Denis Becquet ◽  
Marie-Pierre Blanchard ◽  
Séverine Guillen ◽  
Bénédicte Boyer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Nuclear Bodies ◽  
Transcriptional Level ◽  
Pituitary Cells ◽  
Nuclear Retention ◽  
Circadian Expression ◽  
Expression Control ◽  
Non Coding Rna

Paraspeckles are nuclear bodies form around the long non-coding RNA, Neat1, and RNA-binding proteins. While their role is not fully understood, they are believed to control gene expression at a post-transcriptional level by means of the nuclear retention of mRNA containing in their 3’-UTR inverted repeats of Alu sequences (IRAlu). In this study, we found that, in pituitary cells, all components of paraspeckles including four major proteins and Neat1 displayed a circadian expression pattern. Furthermore the insertion of IRAlu at the 3’-UTR of the EGFP cDNA led to a rhythmic circadian nuclear retention of the egfp mRNA that was lost when paraspeckles were disrupted whereas insertion of a single antisense Alu had only a weak effect. Using real-time video-microscopy, these IRAlu were further shown to drive a circadian expression of EGFP protein. This study shows that paraspeckles, thanks to their circadian expression, control circadian gene expression at a post-transcriptional level.

scholarly journals Global Analysis of Circadian Expression in the Cyanobacterium Synechocystis sp. Strain PCC 6803

2005 ◽  
Vol 187 (6) ◽  
pp. 2190-2199 ◽  
Author(s):  
Ken-ichi Kucho ◽  
Kazuhisa Okamoto ◽  
Yuka Tsuchiya ◽  
Satoshi Nomura ◽  
Mamoru Nango ◽  
...  
Keyword(s):  
Circadian Clock ◽  
Dna Microarrays ◽  
Global Analysis ◽  
Bacterial Species ◽  
Physiological State ◽  
Expression Patterns ◽  
Main Role ◽  
Circadian Expression ◽  
Pcc 6803

ABSTRACT Cyanobacteria are the only bacterial species found to have a circadian clock. We used DNA microarrays to examine circadian expression patterns in the cyanobacterium Synechocystis sp. strain PCC 6803. Our analysis identified 54 (2%) and 237 (9%) genes that exhibited circadian rhythms under stringent and relaxed filtering conditions, respectively. The expression of most cycling genes peaked around the time of transition from subjective day to night, suggesting that the main role of the circadian clock in Synechocystis is to adjust the physiological state of the cell to the upcoming night environment. There were several chromosomal regions where neighboring genes were expressed with similar circadian patterns. The physiological functions of the cycling genes were diverse and included a wide variety of metabolic pathways, membrane transport, and signal transduction. Genes involved in respiration and poly(3-hydroxyalkanoate) synthesis showed coordinated circadian expression, suggesting that the regulation is important for the supply of energy and carbon source in the night. Genes involved in transcription and translation also followed circadian cycling patterns. These genes may be important for output of the rhythmic information generated by the circadian clock. Our findings provided critical insights into the importance of the circadian clock on cellular physiology and the mechanism of clock-controlled gene regulation.

scholarly journals Distinct Expression Patterns Predict Differential Roles of the miRNA-Binding Proteins, Lin28 and Lin28b, in the Mouse Testis: Studies During Postnatal Development and in a Model of Hypogonadotropic Hypogonadism

Endocrinology ◽  
2013 ◽  
Vol 154 (3) ◽  
pp. 1321-1336 ◽  
Author(s):  
Francisco Gaytan ◽  
Susana Sangiao-Alvarellos ◽  
María Manfredi-Lozano ◽  
David García-Galiano ◽  
Francisco Ruiz-Pino ◽  
...  
Keyword(s):  
Postnatal Development ◽  
Binding Proteins ◽  
Rna Binding ◽  
Hypogonadotropic Hypogonadism ◽  
Expression Patterns ◽  
Mouse Testis ◽  
Null Mouse ◽  
Cellular Distribution ◽  
Protein Distribution ◽  
Postnatal Maturation

Abstract Lin28 (also termed Lin28a) and Lin28b are related RNA-binding proteins, involved in the control of microRNA synthesis, especially of the let-7 family, with putative functions in early (embryo) development. However, their roles during postnatal maturation remain ill defined. Despite the general assumption that Lin28 and Lin28b share similar targets and functions, conclusive demonstration of such redundancy is still missing. In addition, recent observations suggest a role of Lin28 proteins in mammalian reproduction, which is yet to be defined. We document herein the patterns of RNA expression and protein distribution of Lin28 and Lin28b in mouse testis during postnatal development and in a model of hypogonadotropic hypogonadism as a result of inactivation of the kisspeptin receptor, Gpr54. Lin28 and Lin28b mRNAs were expressed in mouse testis across postnatal maturation, but their levels disparately varied between neonatal and pubertal periods, with peak Lin28 levels in infantile testes and sustained elevation of Lin28b mRNA in young adult male gonads, where relative levels of let-7a and let-7b miRNAs were significantly suppressed. In addition, Lin28 peptides displayed totally different patterns of cellular distribution in mouse testis: Lin28 was located in undifferentiated and type-A1 spermatogonia, whereas Lin28b was confined to spermatids and interstitial Leydig cells. These profiles were perturbed in Gpr54 null mouse testis, which showed preserved but irregular Lin28 signal and absence of Lin28b peptide, which was rescued by administration of gonadotropins, mainly hCG (as super-agonist of LH). In addition, increased relative levels of Lin28, but not Lin28b, mRNA and of let-7a/let-7b miRNAs were observed in Gpr54 KO mouse testes. Altogether, our data are the first to document the divergent patterns of cellular distribution and mRNA expression of Lin28 and Lin28b in the mouse testis along postnatal maturation and their alteration in a model of congenital hypogonadotropic hypogonadism. Our findings suggest distinct functional roles of these two related, but not overlapping, miRNA-binding proteins in the male gonad.

scholarly journals Compendium of Methods to Uncover RNA-Protein Interactions In Vivo

2021 ◽  
Vol 4 (1) ◽  
pp. 22
Author(s):  
Mrinmoyee Majumder ◽  
Viswanathan Palanisamy
Keyword(s):  
Gene Expression ◽  
Protein Interactions ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Expression Patterns ◽  
Control Of Gene Expression ◽  
Regulatory Pathways ◽  
Advantages And Disadvantages ◽  
Protein Nucleic Acid

Control of gene expression is critical in shaping the pro-and eukaryotic organisms’ genotype and phenotype. The gene expression regulatory pathways solely rely on protein–protein and protein–nucleic acid interactions, which determine the fate of the nucleic acids. RNA–protein interactions play a significant role in co- and post-transcriptional regulation to control gene expression. RNA-binding proteins (RBPs) are a diverse group of macromolecules that bind to RNA and play an essential role in RNA biology by regulating pre-mRNA processing, maturation, nuclear transport, stability, and translation. Hence, the studies aimed at investigating RNA–protein interactions are essential to advance our knowledge in gene expression patterns associated with health and disease. Here we discuss the long-established and current technologies that are widely used to study RNA–protein interactions in vivo. We also present the advantages and disadvantages of each method discussed in the review.

scholarly journals Single cell RNA-seq reveals genes vital to in vitro fertilized embryos and parthenotes in pigs

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Zhi-Qiang Du ◽  
Hao Liang ◽  
Xiao-Man Liu ◽  
Yun-Hua Liu ◽  
Chonglong Wang ◽  
...  
Keyword(s):  
Embryo Development ◽  
Gene Networks ◽  
Regulatory Networks ◽  
Rna Binding ◽  
Expression Patterns ◽  
Early Embryo ◽  
Specific Factor ◽  
Early Embryos ◽  
Genome Activation

AbstractSuccessful early embryo development requires the correct reprogramming and configuration of gene networks by the timely and faithful execution of zygotic genome activation (ZGA). However, the regulatory principle of molecular elements and circuits fundamental to embryo development remains largely obscure. Here, we profiled the transcriptomes of single zygotes and blastomeres, obtained from in vitro fertilized (IVF) or parthenogenetically activated (PA) porcine early embryos (1- to 8-cell), focusing on the gene expression dynamics and regulatory networks associated with maternal-to-zygote transition (MZT) (mainly maternal RNA clearance and ZGA). We found that minor and major ZGAs occur at 1-cell and 4-cell stages for both IVF and PA embryos, respectively. Maternal RNAs gradually decay from 1- to 8-cell embryos. Top abundantly expressed genes (CDV3, PCNA, CDR1, YWHAE, DNMT1, IGF2BP3, ARMC1, BTG4, UHRF2 and gametocyte-specific factor 1-like) in both IVF and PA early embryos identified are of vital roles for embryo development. Differentially expressed genes within IVF groups are different from that within PA groups, indicating bi-parental and maternal-only embryos have specific sets of mRNAs distinctly decayed and activated. Pathways enriched from DEGs showed that RNA associated pathways (RNA binding, processing, transport and degradation) could be important. Moreover, mitochondrial RNAs are found to be actively transcribed, showing dynamic expression patterns, and for DNA/H3K4 methylation and transcription factors as well. Taken together, our findings provide an important resource to investigate further the epigenetic and genome regulation of MZT events in early embryos of pigs.

scholarly journals To Divide or Not to Divide? How Deuterium Affects Growth and Division of Chlamydomonas reinhardtii

Biomolecules ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 861
Author(s):  
Veronika Kselíková ◽  
Vilém Zachleder ◽  
Kateřina Bišová
Keyword(s):  
Cell Cycle ◽  
Chlamydomonas Reinhardtii ◽  
Cell Cycle Progression ◽  
Model Organism ◽  
Cycle Progression ◽  
Synchronous Cultures ◽  
Multiple Fission ◽  
First Choice ◽  
High Doses

Extensive in vivo replacement of hydrogen by deuterium, a stable isotope of hydrogen, induces a distinct stress response, reduces cell growth and impairs cell division in various organisms. Microalgae, including Chlamydomonas reinhardtii, a well-established model organism in cell cycle studies, are no exception. Chlamydomonas reinhardtii, a green unicellular alga of the Chlorophyceae class, divides by multiple fission, grows autotrophically and can be synchronized by alternating light/dark regimes; this makes it a model of first choice to discriminate the effect of deuterium on growth and/or division. Here, we investigate the effects of high doses of deuterium on cell cycle progression in C. reinhardtii. Synchronous cultures of C. reinhardtii were cultivated in growth medium containing 70 or 90% D2O. We characterize specific deuterium-induced shifts in attainment of commitment points during growth and/or division of C. reinhardtii, contradicting the role of the “sizer” in regulating the cell cycle. Consequently, impaired cell cycle progression in deuterated cultures causes (over)accumulation of starch and lipids, suggesting a promising potential for microalgae to produce deuterated organic compounds.

scholarly journals Poplar protease inhibitor expression differs in an herbivore specific manner

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Franziska Eberl ◽  
Thomas Fabisch ◽  
Katrin Luck ◽  
Tobias G. Köllner ◽  
Heiko Vogel ◽  
...  
Keyword(s):  
Protease Inhibitors ◽  
Molecular Mechanisms ◽  
Expression Patterns ◽  
Transcriptional Level ◽  
Species Identity ◽  
Defense Proteins ◽  
Woody Perennials ◽  
Cdna Sequences ◽  
Black Poplar ◽  
Leaf Area Loss

Abstract Background Protease inhibitors are defense proteins widely distributed in the plant kingdom. By reducing the activity of digestive enzymes in insect guts, they reduce the availability of nutrients and thus impair the growth and development of the attacking herbivore. One well-characterized class of protease inhibitors are Kunitz-type trypsin inhibitors (KTIs), which have been described in various plant species, including Populus spp. Long-lived woody perennials like poplar trees encounter a huge diversity of herbivores, but the specificity of tree defenses towards different herbivore species is hardly studied. We therefore aimed to investigate the induction of KTIs in black poplar (P. nigra) leaves upon herbivory by three different chewing herbivores, Lymantria dispar and Amata mogadorensis caterpillars, and Phratora vulgatissima beetles. Results We identified and generated full-length cDNA sequences of 17 KTIs that are upregulated upon herbivory in black poplar leaves, and analyzed the expression patterns of the eight most up-regulated KTIs via qRT-PCR. We found that beetles elicited higher transcriptional induction of KTIs than caterpillars, and that both caterpillar species induced similar KTI expression levels. Furthermore, KTI expression strongly correlated with the trypsin-inhibiting activity in the herbivore-damaged leaves, but was not dependent on damage severity, i.e. leaf area loss, for most of the genes. Conclusions We conclude that the induction of KTIs in black poplar is controlled at the transcriptional level in a threshold-based manner and is strongly influenced by the species identity of the herbivore. However, the underlying molecular mechanisms and ecological consequences of these patterns remain to be investigated.

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