7 research outputs found
Azacitidine (5-AzaC)-treatment and mutations in DNA methylase genes affect embryogenic response and expression of the genes that are involved in somatic embryogenesis in Arabidopsis
Epigenetic processes including DNA methylation play a pivotal role in regulating the genes that control plant development. In contrast to in planta development, the contribution of DNA methylation to the morphogenic processes that are induced in vitro are much less recognised. Hence, in the present study, we analysed the impact of DNA methylation on somatic embryogenesis (SE) that was induced in Arabidopsis. The results demonstrated a decrease in the global DNA methylation level during SE that contrasted with the up-regulation of MET1 and CMT3 DNA methylases and the down-regulation of DNA demethylases (ROS1, DME and DML2). Hence, the global DNA methylation level appears not to correlate with the transcriptional activity of the genes encoding DNA methylases/demethylases, thereby implying the complexity of the regulatory
mechanism that controls the DNA methylation status of the SE-epigenome. Moreover, distinct changes in the expression level of the SE-regulatory genes were indicated in the 5-AzaC-treated and DNA methylase mutant cultures. Accordingly, a significant repression of the LEC2, LEC1 and BBM genes was found in the 5-AzaC-treated culture that was incapable of SE induction. In contrast, the distinct up-regulation of these genes was observed in the drm1drm2 and drm1drm2cmt3 mutant cultures with an improved embryogenic response. The modulated expression of DNA methylase genes and the significantly modified embryogenic response of the met1 and drm mutants imply that both the maintenance and the de novo pathway of DNA methylation are engaged in the regulation of SE in Arabidopsis
Trichostatin A Triggers an Embryogenic Transition in Arabidopsis Explants via an Auxin-Related Pathway
Auxin is an important regulator of plant ontogenies including embryo development and
the exogenous application of this phytohormone has been found to be necessary for
the induction of the embryogenic response in plant explants that have been cultured
in vitro. However, in the present study, we show that treatment of Arabidopsis explants
with trichostatin A (TSA), which is a chemical inhibitor of histone deacetylases, induces
somatic embryogenesis (SE) without the exogenous application of auxin. We found that
the TSA-treated explants generated somatic embryos that developed efficiently on the
adaxial side of the cotyledons, which are the parts of an explant that are involved in
auxin-induced SE. A substantial reduction in the activity of histone deacetylase (HDAC)
was observed in the TSA-treated explants, thus confirming a histone acetylationrelated
mechanism of the TSA-promoted embryogenic response. Unexpectedly, the
embryogenic effect of TSA was lower on the auxin-supplemented media and this finding
further suggests an auxin-related mechanism of TSA-induced SE. Congruently, we
found a significantly increased content of indolic compounds, which is indicative of IAA
and an enhanced DR5::GUS signal in the TSA-treated explants. In line with these results,
two of the YUCCA genes (YUC1 and YUC10), which are involved in auxin biosynthesis,
were found to be distinctly up-regulated during TSA-induced SE and their expression
was colocalised with the explant sites that are involved in SE. Beside auxin, ROS were
extensively accumulated in response to TSA, thereby indicating that a stress-response
is involved in TSA-triggered SE. Relevantly, we showed that the genes encoding the
transcription factors (TFs) that have a regulatory function in auxin biosynthesis including
LEC1, LEC2, BBM, and stress responses (MYB118) were highly up-regulated in the
TSA-treated explants. Collectively, the results provide several pieces of evidence about
the similarities between the molecular pathways of SE induction that are triggered by
TSA and 2,4-D that involve the activation of the auxin-responsive TF genes that have
a regulatory function in auxin biosynthesis and stress responses. The study suggests
the involvement of histone acetylation in the auxin-mediated release of the embryogenic
program of development in the somatic cells of Arabidopsis
AGL15 Controls the Embryogenic Reprogramming of Somatic Cells in Arabidopsis through the Histone Acetylation-Mediated Repression of the miRNA Biogenesis Genes
The embryogenic transition of somatic cells requires an extensive reprogramming of
the cell transcriptome. Relevantly, the extensive modulation of the genes that have a regulatory
function, in particular the genes encoding the transcription factors (TFs) and miRNAs, have been
indicated as controlling somatic embryogenesis (SE) that is induced in vitro in the somatic cells of
plants. Identifying the regulatory relationships between the TFs and miRNAs during SE induction
is of central importance for understanding the complex regulatory interplay that fine-tunes a
cell transcriptome during the embryogenic transition. Hence, here, we analysed the regulatory
relationships between AGL15 (AGAMOUS-LIKE 15) TF and miR156 in an embryogenic culture of
Arabidopsis. Both AGL15 and miR156 control SE induction and AGL15 has been reported to target
the MIR156 genes in planta. The results showed that AGL15 contributes to the regulation of miR156
in an embryogenic culture at two levels that involve the activation of the MIR156 transcription and
the containment of the abundance of mature miR156 by repressing the miRNA biogenesis genes
DCL1 (DICER-LIKE1), SERRATE and HEN1 (HUA-ENHANCER1). To repress the miRNA biogenesis
genes AGL15 seems to co-operate with the TOPLESS co-repressors (TPL and TPR1-4), which are
components of the SIN3/HDAC silencing complex. The impact of TSA (trichostatin A), an inhibitor
of the HDAC histone deacetylases, on the expression of the miRNA biogenesis genes together with
the ChIP results implies that histone deacetylation is involved in the AGL15-mediated repression
of miRNA processing. The results indicate that HDAC6 and HDAC19 histone deacetylases might
co-operate with AGL15 in silencing the complex that controls the abundance of miR156 during
embryogenic induction. This study provides new evidence about the histone acetylation-mediated
control of the miRNA pathways during the embryogenic reprogramming of plant somatic cells and
the essential role of AGL15 in this regulatory mechanism
Insights into the Histone Acetylation-Mediated Regulation of the Transcription Factor Genes That Control the Embryogenic Transition in the Somatic Cells of Arabidopsis
Somatic embryogenesis (SE), which is a process that involves the in vitro-induced embryogenic
reprogramming of plant somatic cells, requires dynamic changes in the cell transcriptome. These
changes are fine-tuned by many genetic and epigenetic factors, including posttranslational histone
modifications such as histone acetylation. Antagonistically acting enzymes, histone acetyltransferases
(HATs) and deacetylases (HDACs), which control histone acetylation in many developmental processes,
are believed to control SE. However, the function of specific HAT/HDACs and the genes
that are subjected to histone acetylation-mediated regulation during SE have yet to be revealed.
Here, we present the global and gene-specific changes in histone acetylation in Arabidopsis explants
that are undergoing SE. In the TSA (trichostatin A)-induced SE, we demonstrate that H3 and H4
acetylation might control the expression of the critical transcription factor (TF) genes of a vital role in
SE, including LEC1, LEC2 (LEAFY COTYLEDON 1; 2), FUS3 (FUSCA 3) and MYB118 (MYB DOMAIN
PROTEIN 118). Within the HATs and HDACs, which mainly positively regulate SE, we identified
HDA19 as negatively affecting SE by regulating LEC1, LEC2 and BBM. Finally, we provide some
evidence on the role of HDA19 in the histone acetylation-mediated regulation of LEC2 during SE. Our
results reveal an essential function of histone acetylation in the epigenetic mechanisms that control
the TF genes that play critical roles in the embryogenic reprogramming of plant somatic cells. The
results implicate the complexity of Hac-related gene regulation in embryogenic induction and point
to differences in the regulatory mechanisms that are involved in auxin- and TSA-induced SE
Use of molecular biology techniques to identify genetically modified organisms (GMOs) in food products – do you know what you eat?
tekst w j. pol. i ang.Ocena poprawności oznaczeń produktów spożywczych pod kątem zawartości genetycznie modyfikowanych organizmów (GMO)
miR172 Regulates WUS during Somatic Embryogenesis in Arabidopsis via AP2
In plants, the embryogenic transition of somatic cells requires the reprogramming of the
cell transcriptome, which is under the control of genetic and epigenetic factors. Correspondingly,
the extensive modulation of genes encoding transcription factors and miRNAs has been indicated
as controlling the induction of somatic embryogenesis in Arabidopsis and other plants. Among
the MIRNAs that have a differential expression during somatic embryogenesis, members of the
MIRNA172 gene family have been identified, which implies a role of miR172 in controlling the
embryogenic transition in Arabidopsis. In the present study, we found a disturbed expression of
both MIRNA172 and candidate miR172-target genes, including AP2, TOE1, TOE2, TOE3, SMZ and
SNZ, that negatively affected the embryogenic response of transgenic explants. Next, we examined
the role of AP2 in the miR172-mediated mechanism that controls the embryogenic response. We
found some evidence that by controlling AP2, miR172 might repress the WUS that has an important
function in embryogenic induction. We showed that the mechanism of the miR172-AP2-controlled
repression of WUS involves histone acetylation. We observed the upregulation of the WUS transcripts
in an embryogenic culture that was overexpressing AP2 and treated with trichostatin A (TSA),
which is an inhibitor of HDAC histone deacetylases. The increased expression of the WUS gene
in the embryogenic culture of the hdac mutants further confirmed the role of histone acetylation in
WUS control during somatic embryogenesis. A chromatin-immunoprecipitation analysis provided
evidence about the contribution of HDA6/19-mediated histone deacetylation to AP2-controlled WUS
repression during embryogenic induction. The upstream regulatory elements of the miR172-AP2-
WUS pathway might involve the miR156-controlled SPL9/SPL10, which control the level of mature
miR172 in an embryogenic culture