Testicular Function: From Gene Expression to Genetic Manipulation

Abstract Background RNA interference (RNAi) regulates gene expression in most multicellular organisms through binding of small RNA effectors to target transcripts. Exploiting this process is a popular strategy for genetic manipulation and has applications that includes arthropod pest control. RNAi technologies are dependent on delivery method with the most convenient likely being feeding, which is effective in some animals while others are insensitive. The two-spotted spider mite, Tetranychus urticae, is prime candidate for developing RNAi approaches due to frequent occurrence of conventional pesticide resistance. Using a sequencing-based approach, the fate of ingested RNAs was explored to identify features and conditions that affect small RNA biogenesis from external sources to better inform RNAi design. Results Biochemical and sequencing approaches in conjunction with extensive computational assessment were used to evaluate metabolism of ingested RNAs in T. urticae. This chelicerae arthropod shows only modest response to oral RNAi and has biogenesis pathways distinct from model organisms. Processing of synthetic and plant host RNAs ingested during feeding were evaluated to identify active substrates for spider mite RNAi pathways. Through cataloging characteristics of biochemically purified RNA from these sources, trans-acting small RNAs could be distinguished from degradation fragments and their origins documented. Conclusions Using a strategy that delineates small RNA processing, we found many transcripts have the potential to enter spider mite RNAi pathways, however, trans-acting RNAs appear very unstable and rare. This suggests potential RNAi pathway substrates from ingested materials are mostly degraded and infrequently converted into regulators of gene expression. Spider mites infest a variety of plants, and it would be maladaptive to generate diverse gene regulators from dietary RNAs. This study provides a framework for assessing RNAi technology in organisms where genetic and biochemical tools are absent and benefit rationale design of RNAi triggers for T.urticae.

Download Full-text

Genetic manipulation of RPS5 gene expression modulates the initiation of commitment of MEL cells to erythroid maturation: Implications in understanding ribosomopathies

International Journal of Oncology ◽

10.3892/ijo.2015.3017 ◽

2015 ◽

Vol 47 (1) ◽

pp. 303-314 ◽

Cited By ~ 3

Author(s):

IOANNIS S. VIZIRIANAKIS ◽

ELENI T. PAPACHRISTOU ◽

PANAGIOTIS ANDREADIS ◽

ELENA ZOPOUNIDOU ◽

CHRISTINA N. MATRAGKOU ◽

...

Keyword(s):

Gene Expression ◽

Genetic Manipulation ◽

Erythroid Maturation

Download Full-text

Minireview: Transcriptional Regulation in Development of Bone

Endocrinology ◽

10.1210/en.2004-1343 ◽

2005 ◽

Vol 146 (3) ◽

pp. 1012-1017 ◽

Cited By ~ 108

Author(s):

Tatsuya Kobayashi ◽

Henry Kronenberg

Keyword(s):

Gene Expression ◽

Transcriptional Regulation ◽

Transcription Factors ◽

Bone Cells ◽

Genetic Manipulation ◽

Bone Development ◽

Regulation Of Gene Expression ◽

Bone Cell ◽

Cellular Functions ◽

Multiple Differentiation

Regulation of gene expression by transcription factors is one of the major mechanisms for controlling cellular functions. Recent advances in genetic manipulation of model animals has allowed the study of the roles of various genes and their products in physiological settings and has demonstrated the importance of specific transcription factors in bone development. Three lineages of bone cells, chondrocytes, osteoblasts, and osteoclasts, develop and differentiate according to their distinct developmental programs. These cells go through multiple differentiation stages, which are often regulated by specific transcription factors. In this minireview, we will discuss selected transcription factors that have been demonstrated to critically affect bone cell development. Further study of these molecules will lead to deeper understanding in mechanisms that govern development of bone.

Download Full-text

A collection of genetic mouse lines and related tools for inducible and reversible intersectional misexpression

10.1101/754192 ◽

2019 ◽

Author(s):

Elham Ahmadzadeh ◽

N. Sumru Bayin ◽

Xinli Qu ◽

Aditi Singh ◽

Linda Madisen ◽

...

Keyword(s):

Gene Expression ◽

Target Gene ◽

Genetic Manipulation ◽

Cell Types ◽

Biological Processes ◽

Control Of Gene Expression ◽

A Cell ◽

Summary Statement ◽

Spatiotemporal Control ◽

Mouse Lines

AbstractThanks to many advances in genetic manipulation, mouse models have become very powerful in their ability to interrogate biological processes. In order to precisely target expression of a gene of interest to particular cell types, intersectional genetic approaches utilizing two promoter/enhancers unique to a cell type are ideal. Within these methodologies, variants that add temporal control of gene expression are the most powerful. We describe the development, validation and application of an intersectional approach that involves three transgenes, requiring the intersection of two promoter/enhancers to target gene expression to precise cell types. Furthermore, the approach utilizes available lines expressing tTA/rTA to control timing of gene expression based on whether doxycycline is absent or present, respectively. We also show that the approach can be extended to other animal models, using chicken embryos. We generated three mouse lines targeted at the Tigre (Igs7) locus with TRE-loxP-tdTomato-loxP upstream of three genes (p21, DTA and Ctgf) and combined them with Cre and tTA/rtTA lines that target expression to the cerebellum and limbs. Our tools will facilitate unraveling biological questions in multiple fields and organisms.Summary statementAhmadzadeh et al. present a collection of four mouse lines and genetic tools for misexpression-mediated manipulation of cellular activity with high spatiotemporal control, in a reversible manner.

Download Full-text

Novel plasmids for gene expression analysis and for genetic manipulation in the gastric pathogenHelicobacter pylori

FEMS Immunology & Medical Microbiology ◽

10.1016/j.femsim.2004.10.016 ◽

2005 ◽

Vol 44 (2) ◽

pp. 157-162 ◽

Cited By ~ 6

Author(s):

Stefan Bereswill ◽

Ruth SchÃ¶nenberger ◽

Arnoud H.M. Vliet ◽

Johannes G. Kusters ◽

Manfred Kist

Keyword(s):

Gene Expression ◽

Expression Analysis ◽

Gene Expression Analysis ◽

Genetic Manipulation

Download Full-text

HIF1α is not a target of 14q deletion in clear cell renal cancer

Scientific Reports ◽

10.1038/s41598-020-74631-7 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Niraj Shenoy

Keyword(s):

Gene Expression ◽

Tumor Suppressor ◽

Protein Expression ◽

Survival Data ◽

Clear Cell ◽

Genetic Manipulation ◽

The Cancer Genome Atlas ◽

Cell Renal Cell Carcinoma ◽

Survival Difference

Abstract HIF1α has been termed a tumor-suppressor in clear cell renal cell carcinoma (ccRCC), primarily based on functional proliferation studies in cell lines (in vitro and in vivo) with genetic manipulation, and the adverse prognosis of 14q-deleted ccRCC patients. In other malignancies, however, HIF1α has an established tumor-promoting role. Therefore, this study sought to further examine the role of HIF1α in ccRCC using bioinformatic analyses of 530 ccRCC patients from The Cancer Genome Atlas (TCGA) and The Cancer Proteome Atlas (TCPA) registries. Although lower copy numbers of HIF1A (encoding HIF1α, located at 14q23.2) was associated with worse survival, there was no survival difference based on either HIF1A mRNA or HIF1α protein expression. Interestingly, L2HGDH (L-2-Hydroxyglutarate Dehydrogenase), a recently characterized epigenetic modulating ccRCC tumor-suppressor with a marked impact on survival, was found to be located only ~ 11.5Mbp from HIF1A on 14q (at 14q21.3). L2HGDH was therefore co-deleted in ~ 95% of 14q deletions involving HIF1A locus. Remarkably, HIF1A CNV had a markedly stronger correlation with L2HGDH expression (Rho = 0.55) than its own gene expression (Rho = 0.27), indicating high preserved-allele compensation of HIF1A. Genetic loss of HIF1A was therefore associated with a much greater reduction of L2HGDH gene expression than its own gene expression, providing a possible explanation for survival differences based on HIF1A CNV and mRNA expression. Furthermore, in 14q-deleted ccRCC patients with complete (uncensored) survival data, in the relatively rare cases where genetic loss of HIF1A occurred without genetic loss of L2HGDH (n = 5), the survival was significantly greater than where there was simultaneous genetic loss of both (n = 87) (mean survival 1670.8 ± 183.5 days vs 885.1 ± 78.4 days; p = 0.007). In addition, there was no correlation between HIF1A mRNA and HIF1α protein expression in ccRCC (R = 0.02), reflecting the primarily post-translational regulation of HIF1α. Lastly, even between L2HGDH and HIF1A loci, 14q was found to have several other yet-to-be-characterized potential ccRCC tumor-suppressors. Taken together, the data indicate that HIF1α is not a target of 14q deletion in ccRCC and that it is not a tumor-suppressor in this malignancy.

Download Full-text

Rex1 (Zfp42) null mice show impaired testicular function, abnormal testis morphology, and aberrant gene expression

Developmental Biology ◽

10.1016/j.ydbio.2011.05.664 ◽

2011 ◽

Vol 356 (2) ◽

pp. 370-382 ◽

Cited By ~ 10

Author(s):

Naira C. Rezende ◽

Mi-Young Lee ◽

Sébastien Monette ◽

Willie Mark ◽

Ailan Lu ◽

...

Keyword(s):

Gene Expression ◽

Testicular Function ◽

Aberrant Gene Expression ◽

Aberrant Gene

Download Full-text

Murine gastrulation requires HNF-4 regulated gene expression in the visceral endoderm: tetraploid rescue of Hnf-4(−/−) embryos

Development ◽

10.1242/dev.124.2.279 ◽

1997 ◽

Vol 124 (2) ◽

pp. 279-287 ◽

Cited By ~ 11

Author(s):

S.A. Duncan ◽

A. Nagy ◽

W. Chan

Keyword(s):

Gene Expression ◽

Genetic Manipulation ◽

Es Cells ◽

Critical Role ◽

Embryonic Stem ◽

Retinol Binding Protein ◽

Specific Gene ◽

Visceral Endoderm ◽

Regulated Gene Expression

Immediately prior to gastrulation the murine embryo consists of an outer layer of visceral endoderm (VE) and an inner layer of ectoderm. Differentiation and migration of the ectoderm then occurs to produce the three germ layers (ectoderm, embryonic endoderm and mesoderm) from which the fetus is derived. An indication that the VE might have a critical role in this process emerged from studies of Hnf-4(−/−) mouse embryos which fail to undergo normal gastrulation. Since expression of the transcription factor HNF-4 is restricted to the VE during this phase of development, we proposed that HNF-4-regulated gene expression in the VE creates an environment capable of supporting gastrulation. To address this directly we have exploited the versatility of embryonic stem (ES) cells which are amenable to genetic manipulation and can be induced to form VE in vitro. Moreover, embryos derived solely from ES cells can be generated by aggregation with tetraploid morulae. Using Hnf-4(−/−) ES cells we demonstrate that HNF-4 is a key regulator of tissue-specific gene expression in the VE, required for normal expression of secreted factors including alphafetoprotein, apolipoproteins, transthyretin, retinol binding protein, and transferrin. Furthermore, specific complementation of Hnf-4(−/−) embryos with tetraploid-derived Hnf-4(+/+) VE rescues their early developmental arrest, showing conclusively that a functional VE is mandatory for gastrulation.

Download Full-text

Wheat Transcription Factor TaSNAC11-4B Positively Regulates Leaf Senescence through Promoting ROS Production in Transgenic Arabidopsis

International Journal of Molecular Sciences ◽

10.3390/ijms21207672 ◽

2020 ◽

Vol 21 (20) ◽

pp. 7672

Author(s):

Zenglin Zhang ◽

Chen Liu ◽

Yongfeng Guo

Keyword(s):

Gene Expression ◽

Stress Tolerance ◽

Leaf Senescence ◽

Transcriptional Activation ◽

Genetic Manipulation ◽

Ectopic Expression ◽

Luciferase Assay ◽

Yield Increase ◽

C Terminus ◽

Functional Homolog

Senescence is the final stage of leaf development which is accompanied by highly coordinated and complicated reprogramming of gene expression. Genetic manipulation of leaf senescence in major crops including wheat has been shown to be able to increase stress tolerance and grain yield. NAC(No apical meristem (NAM), ATAF1/2, and cup-shaped cotyledon (CUC)) transcription factors (TFs) play important roles in regulating gene expression changes during leaf senescence and in response to abiotic stresses. Here, we report the characterization of TaSNAC11-4B (Uniprot: A0A1D5XI64), a wheat NAC family member that acts as a functional homolog of AtNAP, a key regulator of leaf senescence in Arabidopsis. The expression of TaSNAC11-4B was up-regulated with the progression of leaf senescence, in response to abscisic acid (ABA) and drought treatments in wheat. Ectopic expression of TaSNAC11-4B in Arabidopsis promoted ROS accumulation and significantly accelerated age-dependent as well as drought- and ABA-induced leaf senescence. Results from transcriptional activity assays indicated that the TaSNAC11-4B protein displayed transcriptional activation activities that are dependent on its C terminus. Furthermore, qRT-PCR and dual-Luciferase assay results suggested that TaSNAC11-4B could positively regulate the expression of AtrbohD and AtrbohF, which encode catalytic subunits of the ROS-producing NADPH oxidase. Further analysis of TaSNAC11-4B in wheat senescence and the potential application of this gene in manipulating leaf senescence with the purpose of yield increase and stress tolerance is discussed.

Download Full-text