Identification of genes preventing transgenerational transmission of stress-induced epigenetic states

Abstract:

Examples of transgenerational transmission of environmentally induced epigenetic traits remain rare and disputed. Abiotic stress can release the transcription of epigenetically suppressed transposons and, noticeably, this activation is only transient. Therefore, it is likely that mechanisms countering the mitotic and meiotic inheritance of stress-triggered chromatin changes must exist but are undefined. To reveal thesemechanisms, we screened for Arabidopsis mutants impaired in the resetting of stress-induced loss of epigenetic silencing and found that two chromatin regulators, Decrease in DNA methylation1 (DDM1) andMorpheus’Molecule1 (MOM1), act redundantly to restore prestress state and thus erase “epigenetic stress memory”. In ddm1 mutants, stress hyperactivates heterochromatic transcription and transcription persists longer than in the wild type. However, this newly acquired state is not transmitted to the progeny.
Strikingly, although stress-induced transcription in mom1 mutants is as rapidly silenced as in wild type, in ddm1mom1double mutants, transcriptional signatures of stress are able to persist and are found
in the progeny of plants stressed as small seedlings. Our results reveal an important, previously unidentified function of DDM1 and MOM1 in rapid resetting of stress induced epigenetic states, and therefore also in preventing their mitotic propagation and transgenerational inheritance.
Intro
Although environmentally induced traits and their transgenerational transmission in plants have been described repeatedly, trait stability and the involvement of epigenetic mechanisms in their generation remain controversial (1–3). In contrast, it has been well documented that environmental challenges such as elevated temperature can transcriptionally activate chromosomal loci normally silenced by repressive chromatin (4–7). However, this release of epigenetic silencing, unaccompanied by changes in DNA methylation or histone modifications, is only transient (4, 5). Such noncanonical release of transcriptional gene silencing (TGS) is similar to alterations in epigenetic regulation observed in the mom1 mutant, where release of TGS occurs without major changes in epigenetic marks (8–13). Although, molecular mechanisms used by MOM1 in TGS regulation are not well understood, genetic studies have linked MOM1 activity to small interfering RNAs (14) and RNA processing (15). In addition, structure/function studies have suggested that a conserved domain ofMOM1 forms a homodimer, which is possibly required as a binding platform
for additional silencing factors (13, 16). The rapid resilencing of heterochromatic transcription induced by heat stress seems to involve changes in nucleosome occupancy and resilencing is delayed in mutants with impaired chromatin assembly (5). These observations suggest that suppressive chromatin has certain plasticity in response to stress, but also a robust buffering system that resets its prestress state. This counters the persistence of stress-induced epigenetic alterations during subsequent development and thus their transmission to the progeny.
Results and Discussion
To identify factors involved in the erasure of “epigenetic stress
memory,” we performed a genetic screen using Arabidopsis line
LUC25 carrying a transcriptionally silenced transgene encoding
firefly luciferase (LUC) (14), which as an endogenous chromosomal
TGS target loci can be transiently activated after heat
stress. First, we introduced the mom1 mutation into LUC25 (mom1
LUC25). The mom1 mutation partially releases silencing of LUC25,
producing weak luciferase signals in roots but not aerial parts, where
the LUC transgene remains silenced. Importantly, the LUC transgene
in mom1 LUC25 is strongly activated by heat stress, similar to
LUC25 (Fig. S1A). We presumed that the introduction of the mom1
mutation would enhance stress-induced luciferase signals, increasing
clarity and thus the efficiency of the mutant screen. Moreover,
although themom1mutation does not directly influence the kinetics
of stress-induced TGS release, MOM1 is involved in buffering
epigenetic states of chromatin (11).We hypothesized, therefore,
that any deficiency in such buffering would facilitate phenotypic
detection of additional epigenetic regulators involved in the
rapid restoration of TGS after stress and, thus, in the erasure of
epigenetic stress memory.
M2 seedlings of mutagenized mom1 LUC25 were germinated
for 5 d and individuals showing enhanced luciferase signals before
stress treatment were removed, because these plants release TGS
constitutively. The remaining seedlings were subjected to heat stress
and plants showing significantly stronger and/or longer-lasting
luciferase signals were selected and grown to maturity (Fig. S1B).
We examined their M3 progeny to determine whether selected