DEMETER-mediated DNA Demethylation in Gamete Companion Cells and the Endosperm, and its Possible Role in Embryo Development in Arabidopsis

2020 ◽  
Vol 63 (5) ◽  
pp. 321-329
Author(s):  
Kyunghyuk Park ◽  
Seunga Lee ◽  
Hyunjin Yoo ◽  
Yeonhee Choi
Science ◽  
2012 ◽  
Vol 337 (6100) ◽  
pp. 1360-1364 ◽  
Author(s):  
C. A. Ibarra ◽  
X. Feng ◽  
V. K. Schoft ◽  
T.-F. Hsieh ◽  
R. Uzawa ◽  
...  

2012 ◽  
Vol 24 (1) ◽  
pp. 183 ◽  
Author(s):  
J. Teson ◽  
K. Lee ◽  
L. Spate ◽  
R. S. Prather

One of the key regulators of gene expression in mammals is DNA methylation. The Tet family (Tet1–3) is suggested to be involved in regulating the level of methylation by hydroxylating a methyl group from 5-methylcytosine to form 5-hydroxymethylcystosine. This hydroxylation alters the 3-dimensional structure of the DNA and results in altered gene expression. Previous studies conducted in the mouse have shown that Tet1 is important for inner cell mass specification by regulating the apparent level of methylation on a specific promoter region in blastocysts and Tet3 is related to the apparent paternal DNA demethylation after fertilization by hydroxylating the paternal genome. The objective of this study was to investigate the expression profile of the Tet family in porcine oocytes and pre-implantation-stage embryos derived from IVF and somatic cell nuclear transfer (SCNT). The RNA was isolated from donor cells, germinal vesicle (GV), MII and 2-cell and blastocyst stage embryos (20 oocytes or embryos per group). Levels of mRNA for each Tet gene were measured by quantitative real-time RT-PCR. The levels of each mRNA transcript were compared to YWHAG, a housekeeping gene that shows a constant level of expression throughout pre-implantation embryo development and normalized to the GV stage. The analysis was repeated with 3 biological replications and 2 experimental replications. Differences in gene expression were compared by ANOVA and P < 0.05 was considered significant. No difference was found in the levels of the Tet family members between GV and MII stage oocytes. Compared with GV stage oocytes, up-regulation of Tet3 at the 2-cell stage was detected in both IVF and SCNT embryos, 4.7 and 6.2 fold, respectively. A dramatic increase in Tet1 was also observed at the blastocyst stage in IVF and SCNT embryos when compared with the GV stage, 65.7 and 79.7 fold increases, respectively. Interestingly, the level of Tet3 was down-regulated in blastocyst embryos at a 25 or more fold decrease compared with GV. The level of Tet2 remained constant throughout embryo development. Embryos (2-cell and blastocyst) compared from IVF and SCNT showed no difference in Tet expression levels. Donor cells had significantly lower levels of Tet2 and Tet3 when compared with GV. Our results indicate that the Tet family shows a dynamic expression profile during porcine pre-implantation embryo development. High expression of Tet3 in 2-cell stage embryos suggests its importance during the post-activation demethylation process. The increase of Tet1 transcript in blastocysts suggests that Tet1 is involved in regulating the type of methylation at the blastocyst stage. These results are consistent with results from previous mouse studies. There was no misregulated expression of the Tet family in SCNT embryos compared with IVF embryos, thus indicating successful reprogramming of the Tet family after SCNT. Lower levels of Tet2 and Tet3 would indicate that Tet1 is important for maintaining type of methylation in donor cells. This is the first report on the profile of the Tet family during porcine pre-implantation embryo development and further studies are needed to clarify their role during this stage.


Reproduction ◽  
2019 ◽  
Vol 158 (2) ◽  
pp. 125-135 ◽  
Author(s):  
Zhenzhen Zhang ◽  
Changjiu He ◽  
Lu Zhang ◽  
Tianqi Zhu ◽  
Dongying Lv ◽  
...  

α-Ketoglutarate (α-KG) is an intermediary metabolite in the tricarboxylic acid (TCA) cycle and functions to inhibit ATPase and maintain the pluripotency of embryonic stem cells (ESCs); however, little is known regarding the effects of α-KG on the development of preimplantation embryos. Herein, we report that α-KG (150 μM) treatment significantly promoted the blastocyst rate, the number of inner cell mass (ICM) cells and foetal growth after embryo transfer. Mechanistic studies revealed two important pathways involved in the α-KG effects on embryo development. First, α-KG modulates mitochondria function by inducing relatively low ATP production without modification of mitochondrial copy number. The relatively low energy metabolism preserves the pluripotency and competence of the ICM. Second, α-KG modifies epigenetics in embryos cultured in vitro by affecting the activity of the DNA demethylation enzyme TET and the DNA methylation gene Dnmt3a to increase the ratio of 5hmC/5mC ratio. Elevation of the 5hmC/5mC ratio not only promotes the pluripotency of the ICM but also leads to a methylation level in an in vitro embryo close to that in an in vivo embryo. All these functions of α-KG collectively contribute to an increase in the number of ICM cells, leading to greater adaptation of cultured embryos to in vitro conditions and promoting foetal growth after embryo transfer. Our findings provide basic knowledge regarding the mechanisms by which α-KG affects embryo development and cell differentiation.


2017 ◽  
Vol 114 (8) ◽  
pp. 2078-2083 ◽  
Author(s):  
Jin-Sup Park ◽  
Jennifer M. Frost ◽  
Kyunghyuk Park ◽  
Hyonhwa Ohr ◽  
Guen Tae Park ◽  
...  

The DEMETER (DME) DNA glycosylase initiates active DNA demethylation via the base-excision repair pathway and is vital for reproduction inArabidopsis thaliana. DME-mediated DNA demethylation is preferentially targeted to small, AT-rich, and nucleosome-depleted euchromatic transposable elements, influencing expression of adjacent genes and leading to imprinting in the endosperm. In the female gametophyte,DMEexpression and subsequent genome-wide DNA demethylation are confined to the companion cell of the egg, the central cell. Here, we show that, in the male gametophyte,DMEexpression is limited to the companion cell of sperm, the vegetative cell, and to a narrow window of time: immediately after separation of the companion cell lineage from the germline. We define transcriptional regulatory elements ofDMEusing reporter genes, showing that a small region, which surprisingly lies within theDMEgene, controls its expression in male and female companion cells.DMEexpression from this minimal promoter is sufficient to rescue seed abortion and the aberrant DNA methylome associated with the nulldme-2mutation. Within this minimal promoter, we found short, conserved enhancer sequences necessary for the transcriptional activities ofDMEand combined predicted binding motifs with published transcription factor binding coordinates to produce a list of candidate upstream pathway members in the genetic circuitry controlling DNA demethylation in gamete companion cells. These data show how DNA demethylation is regulated to facilitate endosperm gene imprinting and potential transgenerational epigenetic regulation, without subjecting the germline to potentially deleterious transposable element demethylation.


Author(s):  
James Cronshaw

Long distance transport in plants takes place in phloem tissue which has characteristic cells, the sieve elements. At maturity these cells have sieve areas in their end walls with specialized perforations. They are associated with companion cells, parenchyma cells, and in some species, with transfer cells. The protoplast of the functioning sieve element contains a high concentration of sugar, and consequently a high hydrostatic pressure, which makes it extremely difficult to fix mature sieve elements for electron microscopical observation without the formation of surge artifacts. Despite many structural studies which have attempted to prevent surge artifacts, several features of mature sieve elements, such as the distribution of P-protein and the nature of the contents of the sieve area pores, remain controversial.


2014 ◽  
Vol 4 (1_suppl) ◽  
pp. s-0034-1376587-s-0034-1376587
Author(s):  
N. Chutkan ◽  
R. Sangani ◽  
H. Zhou ◽  
S. Fulzele

2014 ◽  
Author(s):  
Akihiko Sakashita ◽  
Yosuke Iseki ◽  
Mei Nakajima ◽  
Takuya Wakai ◽  
Hisato Kobayashi ◽  
...  

2014 ◽  
Author(s):  
Xing Duan ◽  
Zhen-Bo Wang ◽  
Xiang-Shun Cui ◽  
Nam-Hyung Kim ◽  
Shao-Chen Sun

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