Landscape of driver mutations and their clinical effects on Down syndrome-related myeloid neoplasms.

ABSTRACT: Transient abnormal myelopoiesis (TAM) is a common complication in newborns with Down syndrome (DS). It commonly progresses to myeloid leukemia (ML-DS) after spontaneous regression. In contrast to the favorable prognosis of primary ML-DS, patients with refractory/relapsed ML-DS have poor outcomes. However, the molecular basis for refractoriness and relapse and the full spectrum of driver mutations in ML-DS remain largely unknown. We conducted a genomic profiling study of 143 TAM, 204 ML-DS, and 34 non-DS acute megakaryoblastic leukemia cases, including 39 ML-DS cases analyzed by exome sequencing. Sixteen novel mutational targets were identified in ML-DS samples. Of these, inactivations of IRX1 (16.2%) and ZBTB7A (13.2%) were commonly implicated in the upregulation of the MYC pathway and were potential targets for ML-DS treatment with bromodomain-containing protein 4 inhibitors. Partial tandem duplications of RUNX1 on chromosome 21 were also found, specifically in ML-DS samples (13.7%), presenting its essential role in DS leukemia progression. Finally, in 177 patients with ML-DS treated following the same ML-DS protocol (the Japanese Pediatric Leukemia and Lymphoma Study Group acute myeloid leukemia -D05/D11), CDKN2A, TP53, ZBTB7A, and JAK2 alterations were associated with a poor prognosis. Patients with CDKN2A deletions (n = 7) or TP53 mutations (n = 4) had substantially lower 3-year event-free survival (28.6% vs 90.5%; P < .001; 25.0% vs 89.5%; P < .001) than those without these mutations. These findings considerably change the mutational landscape of ML-DS, provide new insights into the mechanisms of progression from TAM to ML-DS, and help identify new therapeutic targets and strategies for ML-DS.

MIZU-KUSSEI1 (MIZ1) and GNOM/MIZ2 control not only positive hydrotropism but also phototropism in Arabidopsis roots.

In response to unilateral blue light illumination, roots of some plant species such as Arabidopsis thaliana exhibit negative phototropism (bending away from light), which is important for light avoidance in nature. MIZU-KUSSEI1 (MIZ1) and GNOM/MIZ2 are essential for positive hydrotropism (i.e. in the presence of a moisture gradient, root bending towards greater water availability). Intriguingly, mutations in these genes also cause a substantial reduction in phototropism. Here, we examined whether the same tissue-specific sites of expression required for MIZ1- and GNOM/MIZ2-regulated hydrotropism in Arabidopsis roots are also required for phototropism. The attenuated phototropic response of miz1 roots was completely restored when a functional MIZ1-green fluorescent protein (GFP) fusion was expressed in the cortex of the root elongation zone but not in other tissues such as root cap, meristem, epidermis, or endodermis. The hydrotropic defect and reduced phototropism of miz2 roots were restored by GNOM/MIZ2 expression in either the epidermis, cortex, or stele, but not in the root cap or endodermis. Thus, the sites in root tissues that are involved in the regulation of MIZ1- and GNOM/MIZ2-dependent hydrotropism also regulate phototropism. These results suggest that MIZ1- and GNOM/MIZ2-mediated pathways are, at least in part, shared by hydrotropic and phototropic responses in Arabidopsis roots.

Isolated Bone Recurrence of Medulloblastoma With MYCN Amplification and TP53 Loss: A Case Report.

Extraneural recurrence of a medulloblastoma is rare with dismal prognosis. A 9-year-old girl with medulloblastoma was treated with gross total resection followed by a combination of chemotherapy and radiotherapy. Fourteen months after treatment completion, she developed multifocal bone metastases. Despite chemotherapy combined with irradiation, she died 18 months after recurrence due to progressive disease. Fluorescence in situ hybridization on formalin-fixed paraffin-embedded tissue sections revealed MYCN amplification and TP53 loss, consistent with the genetic alterations of a rapidly progressive subgroup of recurrent medulloblastomas. In clinical practice, dismal biologic features can be determined using fluorescence in situ hybridization in defective materials.

Association of Multiple Gene Polymorphisms Including Homozygous NUDT15 R139C With Thiopurine Intolerance During the Treatment of Acute Lymphoblastic Leukemia.

Although thiopurine is a crucial drug for treating acute lymphoblastic leukemia, individual variations in intolerance are observed due to gene polymorphisms. A 3-year-old boy with B-cell precursor acute lymphoblastic leukemia who was administered thiopurine developed mucositis, sepsis, and hemophagocytic lymphohistiocytosis due to prolonged hematologic toxicity, chronic disseminated candidiasis, and infective endocarditis that triggered multiple brain infarctions. The patient was found to harbor 3 gene polymorphisms associated with thiopurine intolerance including homozygous NUDT15 R139C, heterozygous ITPA C94A, and homozygous MTHFR C677T and heterozygous RFC1 G80A. Thus, the combined effect of intolerance via multiple gene polymorphisms should be considered in case of unexpected adverse reactions.

Clinical, cytogenetic, and molecular analyses of 17 neonates with transient abnormal myelopoiesis and nonconstitutional trisomy 21.

BACKGROUND: Transient abnormal myelopoiesis (TAM) is a unique myeloproliferative disorder that occurs in neonates with constitutional trisomy 21/Down syndrome (DS). Although TAM also develops in neonates without constitutional trisomy 21, the clinical, cytogenetic, and molecular characteristics of those patients are not fully understood. PROCEDURE: We retrospectively evaluated the clinical and cytogenetic findings and GATA1 mutation status of 17 neonates with TAM and nonconstitutional trisomy 21 tested for GATA1 mutations at our institute, and compared the findings with those of 64 neonates with TAM and constitutional trisomy 21/DS. RESULTS: DS clinical features were observed in five of the 17 (29%) patients. In all patients, both trisomy 21 and GATA1 mutations were detected in diagnostic samples. Over a median follow-up of 33 (range, 0-139) months, early death (< 6 months of age) occurred in four patients (24%). Overall and event-free survivals were not significantly different between the patients with TAM and nonconstitutional trisomy 21 and those with TAM and constitutional trisomy 21/DS (five-year overall survival: 76% ± 10% vs 53% ± 13%, P = 0.40; five-year event-free survival: 55% ± 13% vs 48% ± 12%, P = 0.90). The five-year cumulative incidence of progression to myeloid leukemia of DS was also similar between the groups (21% vs 24%, P = 0.80). CONCLUSIONS: Patients with TAM and nonconstitutional trisomy 21 exhibited similar biology and outcomes to those with TAM and constitutional trisomy 21/DS. The possibility of TAM should be considered even in phenotypically normal neonates with TAM symptoms, for appropriate management.

Centromere-targeted de novo integrations of an LTR retrotransposon of Arabidopsis lyrata.

The plant genome evolves with rapid proliferation of LTR-type retrotransposons, which is associated with their clustered accumulation in gene-poor regions, such as centromeres. Despite their major role for plant genome evolution, no mobile LTR element with targeted integration into gene-poor regions has been identified in plants. Here, we report such targeted integrations de novo. We and others have previously shown that an ATCOPIA93 family retrotransposon in Arabidopsis thaliana is mobilized when the DNA methylation machinery is compromised. Although ATCOPIA93 family elements are low copy number in the wild-type A. thaliana genome, high-copy-number related elements are found in the wild-type Arabidopsis lyrata genome, and they show centromere-specific localization. To understand the mechanisms for the clustered accumulation of the A. lyrata elements directly, we introduced one of them, named Tal1 (Transposon of Arabidopsis lyrata 1), into A. thaliana by transformation. The introduced Tal1 was retrotransposed in A. thaliana, and most of the retrotransposed copies were found in centromeric repeats of A. thaliana, suggesting targeted integration. The targeted integration is especially surprising because the centromeric repeat sequences differ considerably between A. lyrata and A. thaliana. Our results revealed unexpectedly dynamic controls for evolution of the transposon-rich heterochromatic regions.

Plasma Membrane-Associated Ca2+-Binding Protein PCaP1 is Involved in Root Hydrotropism of Arabidopsis thaliana.

Root hydrotropism is an essential growth response to water potential gradients in plants. To understand the mechanism, fundamental elements such as MIZU-KUSSEI 1 (MIZ1) have been investigated extensively. We investigated the physiological role of a plasma membrane-associated cation-binding protein (PCaP1) and examined the effect of PCaP1 loss-of-function mutations on root hydrotropism. pcap1 knockout mutants showed a defect in root bending as a hydrotropic response, although gravitropism was normal in pcap1 mutants. When pcap1 seedlings were treated with abscisic acid, a negative regulator of gravitropism, the seedlings showed normal gravitropism. The hydrotropism defect in pcap1 mutants was clearly rescued by introducing the genomic sequence of PCaP1 with an endodermis-specific promoter. Analysis of PCaP1-greenfluorescent protein-expressing roots by confocal laser scanning microscopy revealed that PCaP1 was stably associated with the plasma membrane in most cells, but in the cytoplasm of endodermal cells at the bending region. Furthermore, we prepared a transgenic line overexpressing MIZ1 on the pcap1 background and found that the pcap1 hydrotropism defect was rescued. Our results indicate that PCaP1 in the endodermal cells of the root elongation zone is involved in the hydrotropic response. We suggest that PCaP1 contributes to hydrotropism through a MIZ1-independent pathway or as one of the upstream components that transduce water potential signals to MIZ1.

Circumnutational movement in rice coleoptiles involves the gravitropic response: analysis of an agravitropic mutant and space-grown seedlings.

Plants exhibit helical growth movements known as circumnutation in growing organs. Some studies indicate that circumnutation involves the gravitropic response, but this notion is a matter of debate. Here, using the agravitropic rice mutant lazy1 and space-grown rice seedlings, we found that circumnutation was reduced or lost during agravitropic growth in coleoptiles. Coleoptiles of wild-type rice exhibited circumnutation in the dark, with vigorous oscillatory movements during their growth. The gravitropic responses in lazy1 coleoptiles differed depending on the growth stage, with gravitropic responses detected during early growth and agravitropism during later growth. The nutation-like movements observed in lazy1 coleoptiles at the early stage of growth were no longer detected with the disappearance of the gravitropic response. To verify the relationship between circumnutation and gravitropic responses in rice coleoptiles, we conducted spaceflight experiments in plants under microgravity conditions on the International Space Station. Wild-type rice seeds were germinated, and the resulting seedlings were grown under microgravity or a centrifuge-generated 1 g environment in space. We began filming the seedlings 2 days after seed imbibition and obtained images of seedling growth every 15 min. The seed germination rate in space was 92-100% under both microgravity and 1 g conditions. LED-synchronized flashlight photography induced an attenuation of coleoptile growth and circumnutational movement due to cumulative light exposure. Nevertheless, wild-type rice coleoptiles still showed circumnutational oscillations under 1 g but not microgravity conditions. These results support the idea that the gravitropic response is involved in plant circumnutation.

Two siblings with familial neuroblastoma with distinct clinical phenotypes harboring an ALK germline mutation.

The authors report two siblings with familial neuroblastoma with a germline R1275Q mutation of the tyrosine kinase domain of ALK. Whole exome sequencing and copy number variation assay were performed to investigate genetic alterations in the two cases. No common somatic mutations or gene polymorphisms related to the tumorigenesis of neuroblastoma were detected. A distinct pattern involving both segmental chromosomal alteration and MYCN amplification was detected. The diversity of biological behavior of familial neuroblastoma harboring a germline ALK mutation may depend on conventional prognostic factors, such as segmental chromosomal alterations and MYCN amplification, rather than additional acquired mutations.

Gravitropism interferes with hydrotropism via counteracting auxin dynamics in cucumber roots: clinorotation and spaceflight experiments.

Roots of land plants show gravitropism and hydrotropism in response to gravity and moisture gradients, respectively, for controlling their growth orientation. Gravitropism interferes with hydrotropism, although the mechanistic aspects are poorly understood. Here, we differentiated hydrotropism from gravitropism in cucumber roots by conducting clinorotation and spaceflight experiments. We also compared mechanisms regulating hydrotropism and auxin-regulated gravitropism. Clinorotated or microgravity (µG)-grown cucumber seedling roots hydrotropically bent toward wet substrate in the presence of moisture gradients, but they grew straight in the direction of normal gravitational force at the Earth's surface (1G) on the ground or centrifuge-generated 1G in space. The roots appeared to become hydrotropically more sensitive to moisture gradients under µG conditions in space. Auxin transport inhibitors significantly reduced the hydrotropic response of clinorotated seedling roots. The auxin efflux protein CsPIN5 was differentially expressed in roots of both clinorotated and µG-grown seedlings; with higher expression in the high-humidity (concave) side than the low-humidity (convex) side of hydrotropically responding roots. Our results suggest that roots become hydrotropically sensitive in µG, and CsPIN5-mediated auxin transport has an important role in inducing root hydrotropism. Thus, hydrotropic and gravitropic responses in cucumber roots may compete via differential auxin dynamics established in response to moisture gradients and gravity.

Auxin transport and response requirements for root hydrotropism differ between plant species.

The direction of auxin transport changes in gravistimulated roots, causing auxin accumulation in the lower side of horizontally reoriented roots. This study found that auxin was similarly involved in hydrotropism and gravitropism in rice and pea roots, but hydrotropism in Lotus japonicus roots was independent of both auxin transport and response. Application of either auxin transport inhibitors or an auxin response inhibitor decreased both hydrotropism and gravitropism in rice roots, and reduced hydrotropism in pea roots. However, Lotus roots treated with these inhibitors showed reduced gravitropism but an unaltered or an enhanced hydrotropic response. Inhibiting auxin biosynthesis substantially reduced both tropisms in rice and Lotus roots. Removing the final 0.2 mm (including the root cap) from the root tip inhibited gravitropism but not hydrotropism in rice seedling roots. These results suggested that modes of auxin involvement in hydrotropism differed between plant species. In rice roots, although auxin transport and responses were required for both gravitropism and hydrotropism, the root cap was involved in the auxin regulation of gravitropism but not hydrotropism. Hydrotropism in Lotus roots, however, may be regulated by a novel mechanism that is independent of both auxin transport and the TIR1/AFBs auxin response pathway.

Gravitropic response and circumnutation in pea (Pisum sativum) seedling roots.

Plant circumnutation is a helical movement of growing organs such as shoots and roots. Gravitropic response is hypothesized to act as an external oscillator in shoot circumnutation, although this is subject to debate. The relationship between circumnutational movement and gravitropic response in roots remains unknown. In this study, we analyzed circumnutation of agravitropic roots using the ageotropum pea (Pisum sativum) mutant, and compared it with that of wild-type (cv. Alaska) pea roots. We further examined the relationship of gravitropic response to circumnutation of Alaska seedling roots by removing the gravisensing tissue (the root cap) and by treating the roots with auxin transport inhibitors. Alaska roots displayed circumnutational movements with a period of approximately 150 min, whereas ageotropum roots did not exhibit distinct circumnutational movement. Removal of the root cap in Alaska roots reduced gravitropic response and circumnutational movements. Treatment of Alaska roots with auxin transport inhibitors, 2,3,5-triiodobenzoic acid (TIBA) and N-(1-naphthyl)phthalamic acid (NPA), dramatically reduced gravitropic response and circumnutational movements. These results suggest that a gravity-regulated auxin transport is involved in circumnutation of pea seedling roots.

Bursts of retrotransposition reproduced in Arabidopsis.

Retrotransposons, which proliferate by reverse transcription of RNA intermediates, comprise a major portion of plant genomes. Plants often change the genome size and organization during evolution by rapid proliferation and deletion of long terminal repeat (LTR) retrotransposons. Precise transposon sequences throughout the Arabidopsis thaliana genome and the trans-acting mutations affecting epigenetic states make it an ideal model organism with which to study transposon dynamics. Here we report the mobilization of various families of endogenous A. thaliana LTR retrotransposons identified through genetic and genomic approaches with high-resolution genomic tiling arrays and mutants in the chromatin-remodelling gene DDM1 (DECREASE IN DNA METHYLATION 1). Using multiple lines of self-pollinated ddm1 mutant, we detected an increase in copy number, and verified this for various retrotransposons in a gypsy family (ATGP3) and copia families (ATCOPIA13, ATCOPIA21, ATCOPIA93), and also for a DNA transposon of a Mutator family, VANDAL21. A burst of retrotransposition occurred stochastically and independently for each element, suggesting an additional autocatalytic process. Furthermore, comparison of the identified LTR retrotransposons in related Arabidopsis species revealed that a lineage-specific burst of retrotransposition of these elements did indeed occur in natural Arabidopsis populations. The recent burst of retrotransposition in natural population is targeted to centromeric repeats, which is presumably less harmful than insertion into genes. The ddm1-induced retrotransposon proliferations and genome rearrangements mimic the transposon-mediated genome dynamics during evolution and provide experimental systems with which to investigate the controlling molecular factors directly.

Role of Abscisic Acid, Reactive Oxygen Species, and Ca2+ Signaling in Hydrotropism-Drought Avoidance-Associated Response of Roots.

Plant roots exert hydrotropism in response to moisture gradients to avoid drought stress. The regulatory mechanism underlying hydrotropism involves novel regulators such as MIZ1 and GNOM/MIZ2 as well as abscisic acid (ABA), reactive oxygen species (ROS), and Ca2+ signaling. ABA, ROS, and Ca2+ signaling are also involved in plant responses to drought stress. Although the mechanism of moisture gradient perception remains largely unknown, the sensory apparatus has been reported to reside in the root elongation zone rather than in the root cap. In Arabidopsis roots, hydrotropism is mediated by the action of MIZ1 and ABA in the cortex of the elongation zone, the accumulation of ROS at the root curvature, and the variation in the cytosolic Ca2+ concentration in the entire root tip including the root cap and stele of the elongation zone. Moreover, root exposure to moisture gradients has been proposed to cause asymmetric ABA distribution or Ca2+ signaling, leading to the induction of the hydrotropic response. A comprehensive and detailed analysis of hydrotropism regulators and their signaling network in relation to the tissues required for their function is apparently crucial for understanding the mechanisms unique to root hydrotropism. Here, referring to studies on plant responses to drought stress, we summarize the recent findings relating to the role of ABA, ROS, and Ca2+ signaling in hydrotropism, discuss their functional sites and plausible networks, and raise some questions that need to be answered in future studies.

Antileukemic effect of azacitidine, a DNA methyltransferase inhibitor, on cell lines of myeloid leukemia associated with Down syndrome.

Myeloid leukemia associated with Down syndrome (ML-DS) responds well to chemotherapy and has a favorable prognosis, but the clinical outcome of patients with refractory or relapsed ML-DS is dismal. We recently reported a case of relapsed ML-DS with an effective response to a DNA methyltransferase inhibitor, azacitidine (AZA). However, the efficacy of AZA for refractory or relapsed ML-DS remains uncertain. Here, we investigated the effects and mechanism of action of AZA on three ML-DS cell lines derived from relapsed cases. AZA inhibited the proliferation of all examined ML-DS cell lines to the same extent as that of AZA-sensitive acute myeloid leukemia non-Down syndrome cell lines. Transient low-dose AZA treatment exerted durable antileukemic effects on ML-DS cells. The inhibitory effect included cell cycle arrest, apoptosis, and reduction of aldehyde dehydrogenase activity. Comprehensive differential gene expression analysis showed that AZA induced megakaryocytic differentiation in all ML-DS cell lines examined. Furthermore, AZA induced activation of type I interferon-stimulated genes, primarily involved in antiproliferation signaling, without stimulation of the interferon receptor-mediated autocrine system. Activation of the type I interferon pathway by stimulation with interferon-α exerted antiproliferative effects on ML-DS cells, suggesting that AZA exerts its antileukemic effects on ML-DS cells at least partially through the type I interferon pathway. Moreover, the effect of AZA on normal hematopoiesis did not differ significantly between individuals with non-Down syndrome and Down syndrome. In summary, this study suggests that AZA is a potentially effective treatment option for ML-DS disease control, including relapsed cases, and has reduced side effects.

RNAi-independent de novo DNA methylation revealed in Arabidopsis mutants of chromatin remodeling gene DDM1.

Methylation of histone H3 lysine 9 (H3K9me) and small RNAs are associated with constitutively silent chromatin in diverse eukaryotes including plants. In plants, silent transposons are also marked by cytosine methylation, especially at non-CpG sites. Transposon-specific non-CpG methylation in plants is controlled by small RNAs and H3K9me. Although it is often assumed that small RNA directs H3K9me, interaction between small RNA and H3K9me has not been directly demonstrated in plants. We have previously shown that a mutation in the chromatin remodeling gene DDM1 (DECREASE IN DNA METHYLATION 1) induces a global decrease but a local increase of cytosine methylation and accumulation of small RNA at a locus called BONSAI. Here we show that de novo BONSAI methylation does not depend on RNAi but does depend on H3K9me. In mutants of H3K9 methyltransferase gene KRYPTONITE or the H3K9me-dependent DNA methyltransferase gene CHROMOMETHYALSE3, the ddm1-induced de novo cytosine methylation was abolished for all three contexts (CpG, CpHpG and CpHpH). Furthermore, RNAi mutants showed strong developmental defects when combined with the ddm1 mutation. Our results revealed unexpected interactions of epigenetic modifications that may be conserved among diverse eukaryotes.

An Arabidopsis jmjC domain protein protects transcribed genes from DNA methylation at CHG sites.

Differential cytosine methylation of genes and transposons is important for maintaining integrity of plant genomes. In Arabidopsis, transposons are heavily methylated at both CG and non-CG sites, whereas the non-CG methylation is rarely found in active genes. Our previous genetic analysis suggested that a jmjC domain-containing protein IBM1 (increase in BONSAI methylation 1) prevents ectopic deposition of non-CG methylation, and this process is necessary for normal Arabidopsis development. Here, we directly determined the genomic targets of IBM1 through high-resolution genome-wide analysis of DNA methylation. The ibm1 mutation induced extensive hyper-methylation in thousands of genes. Transposons were unaffected. Notably, long transcribed genes were most severely affected. Methylation of genes is limited to CG sites in wild type, but CHG sites were also methylated in the ibm1 mutant. The ibm1-induced hyper-methylation did not depend on previously characterized components of the RNAi-based DNA methylation machinery. Our results suggest novel transcription-coupled mechanisms to direct genic methylation not only at CG but also at CHG sites. IBM1 prevents the CHG methylation in genes, but not in transposons.

Molecular mechanisms of hydrotropism in seedling roots of Arabidopsis thaliana (Brassicaceae).

Roots show positive hydrotropism in response to moisture gradients, which is believed to contribute to plant water acquisition. This article reviews the recent advances of the physiological and molecular genetic studies on hydrotropism in seedling roots of Arabidopsis thaliana. We identified MIZU-KUSSEI1 (MIZ1) and MIZ2, essential genes for hydrotropism in roots; the former encodes a protein of unknown function, and the latter encodes an ARF-GEF (GNOM) protein involved in vesicle trafficking. Because both mutants are defective in hydrotropism but not in gravitropism, these mutations might affect a molecular mechanism unique to hydrotropism. MIZ1 is expressed in the lateral root cap and cortex of the root proper. It is localized as a soluble protein in the cytoplasm and in association with the cytoplasmic face of endoplasmic reticulum (ER) membranes in root cells. Light and ABA independently regulate MIZ1 expression, which influences the ultimate hydrotropic response. In addition, MIZ1 overexpression results in an enhancement of hydrotropism and an inhibition of lateral root formation. This phenotype is likely related to the alteration of auxin content in roots. Specifically, the auxin level in the roots decreases in the MIZ1 overexpressor and increases in the miz1 mutant. Unlike most gnom mutants, miz2 displays normal morphology, growth, and gravitropism, with normal localization of PIN proteins. It is probable that MIZ1 plays a crucial role in hydrotropic response by regulating the endogenous level of auxin in Arabidopsis roots. Furthermore, the role of GNOM/MIZ2 in hydrotropism is distinct from that of gravitropism.

The effects of heat induction and the siRNA biogenesis pathway on the transgenerational transposition of ONSEN, a copia-like retrotransposon in Arabidopsis thaliana.

Environmental stress influences genetic and epigenetic regulation in plant genomes. We previously reported that heat stress activated a copia-like retrotransposon named ONSEN. To investigate the heat sensitivity and transgenerational activation of ONSEN, we analyzed the stress response by temperature shift and multiple heat stress treatments. ONSEN was activated at 37°C, and the newly inserted ONSEN was transcriptionally active and mobile to the next generation subjected to heat stress, indicating that the regulation of ONSEN is independent of positional effects on the chromosome. Reciprocal crosses with activated ONSEN revealed that the transgenerational transposition was inherited from both sexes, indicating that the transposition is suppressed independently of gametophytic regulation. We showed previously that ONSEN was transposed in mutants deficient in small interfering RNA (siRNA) biogenesis, including nrpd2 and rdr2, but not dcl3. To define the functional redundancy of Dicer-like (DCL) proteins in Arabidopsis, we analyzed ONSEN activation in mutants deficient in DCL proteins, including dcl2, dcl3 and dcl4. ONSEN was nearly immobile in a single Dicer mutant; however, some transgenerational transpositions were observed in dcl2/dcl3/dcl4 triple mutants subjected to heat stress. This indicated that the Dicer family is redundant for ONSEN transposition. To examine the activation of ONSEN in undifferentiated cells, ONSEN transcripts and synthesized DNA were analyzed in heat-stressed callus tissue. In contrast to vegetative tissue, high accumulation of the transcripts and amplified DNA copies of ONSEN were detected in callus. This result indicated that ONSEN activation is controlled by cell-specific regulatory mechanisms.