Differences in the translation efficiency and mRNA stability mediated by 5′-UTR splice variants of human SP-A1 and SP-A2 genes

2005 ◽  
Vol 289 (3) ◽  
pp. L497-L508 ◽  
Author(s):  
Guirong Wang ◽  
Xiaoxuan Guo ◽  
Joanna Floros
Keyword(s):  
Gene Expression ◽  
Mrna Stability ◽  
Mrna Decay ◽  
Molecular Mechanisms ◽  
Luciferase Activity ◽  
Splice Variants ◽  
Translational Efficiency ◽  
Translation Efficiency ◽  
Control Vector

Surfactant protein A (SP-A) plays an important role in host defense, modulation of inflammatory processes, and surfactant-related functions of the lung. The human SP-A (hSP-A) locus consists of two functional genes, SP-A1 and SP-A2. Several hSP-A 5′-untranslated region (UTR) splice variants for each gene have been characterized and shown to be translated in vitro and in vivo. In this report, we investigated the role of hSP-A 5′-UTR splice variants on SP-A production and molecular mechanisms involved. We used in vitro transient expression of hSP-A 5′-UTR constructs containing luciferase as the reporter gene and quantitative real-time PCR to study hSP-A 5′-UTR-mediated gene expression. We found that 1) the four (A′D′, ABD, AB′D′, and A′CD′) 5′-UTR splice variants under study enhanced gene expression, by increasing luciferase activity from 2.5- to 19.5-fold and luciferase mRNA from 4.3- to 8.8-fold compared with the control vector that lacked hSP-A 5′-UTR; 2) all four 5′-UTR splice variants studied regulated mRNA stability. The ABD variant exhibited the lowest rate of mRNA decay compared with the other three constructs (A′D′, AB′D′, and A′CD′). These three constructs also exhibited significantly lower rate of mRNA decay compared with the control vector; 3) based on the indexes of translational efficiency (luciferase activity/mRNA), ABD and AB′D′ exhibited higher translational efficiency compared with the control vector, whereas the translational efficiency of each A′D′ and A′CD′ was lower than that of the control vector. These findings indicate that the hSP-A 5′-UTR splice variants play an important role in both SP-A translation and mRNA stability.

scholarly journals Cytokine-mediated regulation of expression of Gfi1 and U2afll4 genes activated by T-cells with different differentiation status in vitro

2016 ◽  
Vol 62 (2) ◽  
pp. 180-186
Author(s):  
K.A. Yurova ◽  
N.A. Sokhonevich ◽  
O.G. Khaziakhmatova ◽  
L.S. Litvinova
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
Molecular Mechanisms ◽  
Splice Variants ◽  
Inhibitory Effect ◽  
Regulation Of Expression ◽  
Dose Dependent Decrease ◽  
Differentiation Status ◽  
Dose Dependent

The dose-dependent effects of cytokines (IL-2, IL-7, and IL-15), which have a common g-chain, on mRNA expression of U2afll4 and GFi1 genes involved in regulation of alternative splicing of the Ptprc gene, have been investigated in vitro using T-lymphocyte cultures with different degrees of differentiation. IL-2, IL-7, and IL-15 caused a similar unidirectional inhibitory effect of various severity on restimulated CD45RO+ T-cells exposed to an antigen-independent activation; they caused a dose-dependent decrease of the U2af1l4 gene expression, and an increase of Gfi1 gene expression. This may suggest formation of active forms of the CD45 receptor, and also limitation of the formation of low-molecular short splice variants of the CD45RO receptor. Under conditions of antigen-independent stimulation of naive CD45RA+-cells rIL-7 and IL-15 exhibited opposite effects on U2af1l4 and Gfi1 gene expression. The increase of IL-7 concentrations in the incubation medium of naive cells was accompanied by a decrease in expression of both genes. IL-15 IL-7 exhibited opposite effects. Cytokines possessing a common g-chain (IL-2, IL-7, and IL-15) prevented antigen-independent differentiation of naive T-cells, by preventing the formation of polyclonal “surrogate“ cells. In general, the study of the molecular mechanisms of genetic control determining homeostatic processes of T-cells in response to exposure to antigenic or non-antigenic treatments may be important for construction of a general model of self-maintenance and differentiation of immune cells

scholarly journals NF-κB-activated SPRY4-IT1 promotes cancer cell metastasis by downregulating TCEB1 mRNA via Staufen1-mediated mRNA decay

Oncogene ◽  
2021 ◽  
Author(s):  
Lin Zhao ◽  
Longyang Jiang ◽  
Ming Zhang ◽  
Qiang Zhang ◽  
Qiutong Guan ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Cancer Cell ◽  
Mrna Stability ◽  
Mrna Decay ◽  
Molecular Mechanisms ◽  
Cancer Metastasis ◽  
Breast And Ovarian Cancer ◽  
Cell Metastasis

AbstractPrevious study demonstrated that most long non-coding RNAs (lncRNAs) function as competing endogenous RNAs or molecular sponges to negatively modulate miRNA and regulate tumor development. However, the molecular mechanisms of lncRNAs in cancer are not fully understood. Our study describes the role of the lncRNA SPRY4 intronic transcript 1 (SPRY4-IT1) in cancer metastasis by mechanisms related to Staufen1 (STAU1)-mediated mRNA decay (SMD). Briefly, we found that, high SPRY4-IT1 expression was associated with aggressiveness and poor outcome in human colorectal, breast and ovarian cancer tissues. In addition, functional assays revealed that SPRY4-IT1 significantly promoted colorectal, breast and ovarian cancer metastasis in vitro and in vivo. Mechanistically, microarray analyses identified several differentially-expressed genes upon SPRY4-IT1 overexpression in HCT 116 colorectal cancer cells. Among them, the 3′-UTR of transcription elongation factor B subunit 1 (TCEB1) mRNA can base-pair with the Alu element in the 3′-UTR of SPRY4-IT1. Moreover, SPRY4-IT1 was found to bind STAU1, promote STAU1 recruitment to the 3′-UTR of TCEB1 mRNA, and affect TCEB1 mRNA stability and expression, resulting in hypoxia-inducible factor 1α (HIF-1α) upregulation, and thereby affecting cancer cell metastasis. In addition, STAU1 depletion abrogated TCEB1 SMD and alleviated the pro-metastatic effect of SPRY4-IT1 overexpression. Significantly, we revealed that SPRY4-IT1 is also transactivated by NF-κB/p65, which activates SPRY4-IT1 to inhibit TCEB1 expression, and subsequently upregulate HIF-1α. In conclusion, our results highlight a novel mechanism of cytoplasmic lncRNA SPRY4-IT1 in which SPRY4-IT1 affecting TCEB1 mRNA stability via STAU1-mediated degradation during cancer metastasis.

scholarly journals Cell–cell coupling and DNA methylation abnormal phenotypes in the after-hours mice

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Federico Tinarelli ◽  
Elena Ivanova ◽  
Ilaria Colombi ◽  
Erica Barini ◽  
Edoardo Balzani ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Neuronal Networks ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Neuronal Cultures ◽  
Functional Impairments ◽  
After Hours ◽  
The Impact

Abstract Background DNA methylation has emerged as an important epigenetic regulator of brain processes, including circadian rhythms. However, how DNA methylation intervenes between environmental signals, such as light entrainment, and the transcriptional and translational molecular mechanisms of the cellular clock is currently unknown. Here, we studied the after-hours mice, which have a point mutation in the Fbxl3 gene and a lengthened circadian period. Methods In this study, we used a combination of in vivo, ex vivo and in vitro approaches. We measured retinal responses in Afh animals and we have run reduced representation bisulphite sequencing (RRBS), pyrosequencing and gene expression analysis in a variety of brain tissues ex vivo. In vitro, we used primary neuronal cultures combined to micro electrode array (MEA) technology and gene expression. Results We observed functional impairments in mutant neuronal networks, and a reduction in the retinal responses to light-dependent stimuli. We detected abnormalities in the expression of photoreceptive melanopsin (OPN4). Furthermore, we identified alterations in the DNA methylation pathways throughout the retinohypothalamic tract terminals and links between the transcription factor Rev-Erbα and Fbxl3. Conclusions The results of this study, primarily represent a contribution towards an understanding of electrophysiological and molecular phenotypic responses to external stimuli in the Afh model. Moreover, as DNA methylation has recently emerged as a new regulator of neuronal networks with important consequences for circadian behaviour, we discuss the impact of the Afh mutation on the epigenetic landscape of circadian biology.

scholarly journals Inter-species functional compatibility of the Theobroma cacao and Arabidopsis FT orthologs: 90 million years of functional conservation of meristem identity genes

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
S. F. Prewitt ◽  
A. Shalit-Kaneh ◽  
S. N. Maximova ◽  
M. J. Guiltinan
Keyword(s):  
Gene Expression ◽  
Tissue Culture ◽  
Gene Family ◽  
Flowering Time ◽  
Molecular Mechanisms ◽  
Regulatory Pathways ◽  
Late Flowering ◽  
Precocious Flowering ◽  
Transition To Flowering

Abstract Background In angiosperms the transition to flowering is controlled by a complex set of interacting networks integrating a range of developmental, physiological, and environmental factors optimizing transition time for maximal reproductive efficiency. The molecular mechanisms comprising these networks have been partially characterized and include both transcriptional and post-transcriptional regulatory pathways. Florigen, encoded by FLOWERING LOCUS T (FT) orthologs, is a conserved central integrator of several flowering time regulatory pathways. To characterize the molecular mechanisms involved in controlling cacao flowering time, we have characterized a cacao candidate florigen gene, TcFLOWERING LOCUS T (TcFT). Understanding how this conserved flowering time regulator affects cacao plant’s transition to flowering could lead to strategies to accelerate cacao breeding. Results BLAST searches of cacao genome reference assemblies identified seven candidate members of the CENTRORADIALIS/TERMINAL FLOWER1/SELF PRUNING gene family including a single florigen candidate. cDNA encoding the predicted cacao florigen was cloned and functionally tested by transgenic genetic complementation in the Arabidopsis ft-10 mutant. Transgenic expression of the candidate TcFT cDNA in late flowering Arabidopsis ft-10 partially rescues the mutant to wild-type flowering time. Gene expression studies reveal that TcFT is spatially and temporally expressed in a manner similar to that found in Arabidopsis, specifically, TcFT mRNA is shown to be both developmentally and diurnally regulated in leaves and is most abundant in floral tissues. Finally, to test interspecies compatibility of florigens, we transformed cacao tissues with AtFT resulting in the remarkable formation of flowers in tissue culture. The morphology of these in vitro flowers is normal, and they produce pollen that germinates in vitro with high rates. Conclusion We have identified the cacao CETS gene family, central to developmental regulation in angiosperms. The role of the cacao’s single FT-like gene (TcFT) as a general regulator of determinate growth in cacao was demonstrated by functional complementation of Arabidopsis ft-10 late-flowering mutant and through gene expression analysis. In addition, overexpression of AtFT in cacao resulted in precocious flowering in cacao tissue culture demonstrating the highly conserved function of FT and the mechanisms controlling flowering in cacao.

Serial analysis of gene expression (SAGE) during porcine embryo development

2004 ◽  
Vol 16 (2) ◽  
pp. 87 ◽  
Author(s):  
Le Ann Blomberg ◽  
Kurt A. Zuelke
Keyword(s):  
Gene Expression ◽  
Functional Genomics ◽  
Embryo Development ◽  
Molecular Mechanisms ◽  
Physiological Role ◽  
Transgenic Animals ◽  
Specific Gene ◽  
Research Strategies

Functional genomics provides a powerful means for delving into the molecular mechanisms involved in pre-implantation development of porcine embryos. High rates of embryonic mortality (30%), following either natural mating or artificial insemination, emphasise the need to improve the efficiency of reproduction in the pig. The poor success rate of live offspring from in vitro-manipulated pig embryos also hampers efforts to generate transgenic animals for biotechnology applications. Previous analysis of differential gene expression has demonstrated stage-specific gene expression for in vivo-derived embryos and altered gene expression for in vitro-derived embryos. However, the methods used to date examine relatively few genes simultaneously and, thus, provide an incomplete glimpse of the physiological role of these genes during embryogenesis. The present review will focus on two aspects of applying functional genomics research strategies for analysing the expression of genes during elongation of pig embryos between gestational day (D) 11 and D12. First, we compare and contrast current methodologies that are being used for gene discovery and expression analysis during pig embryo development. Second, we establish a paradigm for applying serial analysis of gene expression as a functional genomics tool to obtain preliminary information essential for discovering the physiological mechanisms by which distinct embryonic phenotypes are derived.

scholarly journals Regulation of Multiple Core Spliceosomal Proteins by Alternative Splicing-Coupled Nonsense-Mediated mRNA Decay

2008 ◽  
Vol 28 (13) ◽  
pp. 4320-4330 ◽  
Author(s):  
Arneet L. Saltzman ◽  
Yoon Ki Kim ◽  
Qun Pan ◽  
Matthew M. Fagnani ◽  
Lynne E. Maquat ◽  
...  
Keyword(s):  
Gene Expression ◽  
Alternative Splicing ◽  
Mrna Decay ◽  
Splice Variants ◽  
Cellular Functions ◽  
Regulate Gene Expression ◽  
Premature Termination Codons ◽  
Microarray Profiling ◽  
Nonsense Mediated Mrna Decay ◽  
Regulate Gene

ABSTRACT Alternative splicing (AS) can regulate gene expression by introducing premature termination codons (PTCs) into spliced mRNA that subsequently elicit transcript degradation by the nonsense-mediated mRNA decay (NMD) pathway. However, the range of cellular functions controlled by this process and the factors required are poorly understood. By quantitative AS microarray profiling, we find that there are significant overlaps among the sets of PTC-introducing AS events affected by individual knockdown of the three core human NMD factors, Up-Frameshift 1 (UPF1), UPF2, and UPF3X/B. However, the levels of some PTC-containing splice variants are less or not detectably affected by the knockdown of UPF2 and/or UPF3X, compared with the knockdown of UPF1. The intron sequences flanking the affected alternative exons are often highly conserved, suggesting important regulatory roles for these AS events. The corresponding genes represent diverse cellular functions, and surprisingly, many encode core spliceosomal proteins and assembly factors. We further show that conserved, PTC-introducing AS events are enriched in genes that encode core spliceosomal proteins. Where tested, altering the expression levels of these core spliceosomal components affects the regulation of PTC-containing splice variants from the corresponding genes. Together, our results show that AS-coupled NMD can have different UPF factor requirements and is likely to regulate many general components of the spliceosome. The results further implicate general spliceosomal components in AS regulation.

Translational regulation of human beta interferon mRNA: association of the 3' AU-rich sequence with the poly(A) tail reduces translation efficiency in vitro

1993 ◽  
Vol 13 (6) ◽  
pp. 3487-3493
Author(s):  
G Grafi ◽  
I Sela ◽  
G Galili
Keyword(s):  
Translational Regulation ◽  
Binding Activity ◽  
Rnase H ◽  
Translational Efficiency ◽  
Translation Efficiency ◽  
Protection Assay ◽  
Rich Region ◽  
Reticulocyte Lysate ◽  
Rna Probe

The 3' AU-rich region of human beta-1 interferon (hu-IFN beta) mRNA was found to act as a translational inhibitory element. The translational regulation of this 3' AU-rich sequence and the effect of its association with the poly(A) tail were studied in cell-free rabbit reticulocyte lysate. A poly(A)-rich hu-IFN beta mRNA (110 A residues) served as an inefficient template for protein synthesis. However, translational efficiency was considerably improved when the poly(A) tract was shortened (11 A residues) or when the 3' AU-rich sequence was deleted, indicating that interaction between these two regions was responsible for the reduced translation of the poly(A)-rich hu-IFN beta mRNA. Differences in translational efficiency of the various hu-IFN beta mRNAs correlated well with their polysomal distribution. The poly(A)-rich hu-IFN beta mRNA failed to form large polysomes, while its counterpart bearing a short poly(A) tail was recruited more efficiently into large polysomes. The AU-rich sequence-binding activity was reduced when the RNA probe contained both the 3' AU-rich sequence and long poly(A) tail, supporting a physical association between these two regions. Further evidence for this interaction was achieved by RNase H protection assay. We suggest that the 3' AU-rich sequence may regulate the translation of hu-IFN beta mRNA by interacting with the poly(A) tail.

Tissue specificity of type I collagen gene expression is determined at both transcriptional and post-transcriptional levels

1984 ◽  
Vol 4 (9) ◽  
pp. 1843-1852
Author(s):  
R J Focht ◽  
S L Adams
Keyword(s):  
Gene Expression ◽  
Rna Structure ◽  
Type I Collagen ◽  
Translational Efficiency ◽  
Type I ◽  
Collagen Gene ◽  
Differentiated Cells ◽  
Collagen Gene Expression

We analyzed the control of type I collagen synthesis in four kinds of differentiated cells from chicken embryos which synthesize very different amounts of the protein. Tendon, skin, and smooth muscle cells were found to have identical amounts of type I collagen RNAs; however, the RNAs had inherently different translatabilities, which were observed both in vivo and in vitro. Chondrocytes also had substantial amounts of type I collagen RNAs, even though they directed no detectable synthesis of the protein either in vivo or in vitro. Type I collagen RNAs in chondrocytes display altered electrophoretic mobilities, suggesting that in these cells the reduction in translational efficiency may be mediated in part by changes in the RNA structure. These data indicate that control of type I collagen gene expression is a complex process which is exerted at both transcriptional and post-transcriptional levels.

scholarly journals Differential regulation of mRNA fate by the human Ccr4-Not complex is driven by coding sequence composition and mRNA localization

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Sarah L. Gillen ◽  
Chiara Giacomelli ◽  
Kelly Hodge ◽  
Sara Zanivan ◽  
Martin Bushell ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Synthesis ◽  
Mrna Stability ◽  
Mrna Localization ◽  
Synthesis Rate ◽  
Protein Synthesis Rate ◽  
Translational Efficiency ◽  
Mrna Levels ◽  
Control Of Gene Expression ◽  
Mrna Fate

Abstract Background Regulation of protein output at the level of translation allows for a rapid adaptation to dynamic changes to the cell’s requirements. This precise control of gene expression is achieved by complex and interlinked biochemical processes that modulate both the protein synthesis rate and stability of each individual mRNA. A major factor coordinating this regulation is the Ccr4-Not complex. Despite playing a role in most stages of the mRNA life cycle, no attempt has been made to take a global integrated view of how the Ccr4-Not complex affects gene expression. Results This study has taken a comprehensive approach to investigate post-transcriptional regulation mediated by the Ccr4-Not complex assessing steady-state mRNA levels, ribosome position, mRNA stability, and protein production transcriptome-wide. Depletion of the scaffold protein CNOT1 results in a global upregulation of mRNA stability and the preferential stabilization of mRNAs enriched for G/C-ending codons. We also uncover that mRNAs targeted to the ER for their translation have reduced translational efficiency when CNOT1 is depleted, specifically downstream of the signal sequence cleavage site. In contrast, translationally upregulated mRNAs are normally localized in p-bodies, contain disorder-promoting amino acids, and encode nuclear localized proteins. Finally, we identify ribosome pause sites that are resolved or induced by the depletion of CNOT1. Conclusions We define the key mRNA features that determine how the human Ccr4-Not complex differentially regulates mRNA fate and protein synthesis through a mechanism linked to codon composition, amino acid usage, and mRNA localization.

scholarly journals GENE EXPRESSION DURING INDIRECT SOMATIC EMBRYOGENESIS OF PLANTS

2004 ◽  
Vol 42 (2) ◽  
Author(s):  
Tammy Estabrooks ◽  
Zhongmin Dong
Keyword(s):  
Gene Expression ◽  
Somatic Embryogenesis ◽  
Embryo Development ◽  
Current Literature ◽  
Molecular Mechanisms ◽  
Indirect Somatic Embryogenesis ◽  
Experimental Tool ◽  
Plant Embryo ◽  
Plant Embryo Development

Somatic embryogenesis is the process by which somatic cells are induced into an embryogenic state, followed by differentiation into embryos. Somatic embryogenesis, in addition to being a method of propagation, can serve as an experimental tool for research into plant embryo development. This is a review of the current literature on in vitro plant somatic embryogenesis and the molecular advances made to identify genes expressed during the various stages of this process. Some factors hindering the elucidation of the molecular mechanisms underlying somatic embryogenesis are discussed.L’embryogenèse somatique est le processus par lequel les cellules somatiques passent à l’état embryogène et se différencient en embryons. En plus de constituer une méthode de propagation, elle peut servir d’outil expérimental de recherche pour développer des embryons de plantes. Le présent document est une revue de la documentation sur l’embryogenèse somatique végétale in vitro et sur les progrès réalisés à l’échelle moléculaire pour identifier les gènes exprimés au cours des divers stades du processus. On examine aussi certains facteurs qui rendent difficile l’élucidation des mécanismes moléculaires de l’embryogenèse somatique.

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