Smc3 Deacetylation by Hos1 Facilitates Efficient Dissolution of Sister Chromatid Cohesion during Early Anaphase.

Cohesins establish sister chromatid cohesion during S phase and are removed when cohesin Scc1 is cleaved by separase at anaphase onset. During this process, cohesin Smc3 undergoes a cycle of acetylation: Smc3 acetylation by Eco1 in S phase stabilizes cohesin association with chromosomes, and its deacetylation by Hos1 in anaphase allows re-use of Smc3 in the next cell cycle. Here we find that Smc3 deacetylation by Hos1 has a more immediate effect in the early anaphase of budding yeast. Hos1 depletion significantly delayed sister chromatid separation and segregation. Smc3 deacetylation facilitated removal of cohesins from chromosomes without changing Scc1 cleavage efficiency, promoting dissolution of cohesion. This action is probably due to disengagement of Smc1-Smc3 heads prompted by de-repression of their ATPase activity. We suggest Scc1 cleavage per se is insufficient for efficient dissolution of cohesion in early anaphase; subsequent Smc3 deacetylation, triggered by Scc1 cleavage, is also required.

HISTONE DEACETYLASE 9 transduces heat signal in plant cells.

Heat stress limits plant growth, development, and crop yield, but how plant cells precisely sense and transduce heat stress signals remains elusive. Here, we identified a conserved heat stress response mechanism to elucidate how heat stress signal is transmitted from the cytoplasm into the nucleus for epigenetic modifiers. We demonstrate that HISTONE DEACETYLASE 9 (HDA9) transduces heat signals from the cytoplasm to the nucleus to play a positive regulatory role in heat responses in Arabidopsis. Heat specifically induces HDA9 accumulation in the nucleus. Under heat stress, the phosphatase PP2AB'ß directly interacts with and dephosphorylates HDA9 to protect HDA9 from 26S proteasome-mediated degradation, leading to the translocation of nonphosphorylated HDA9 to the nucleus. This heat-induced enrichment of HDA9 in the nucleus depends on the nucleoporin HOS1. In the nucleus, HDA9 binds and deacetylates the target genes related to signaling transduction and plant development to repress gene expression in a transcription factor YIN YANG 1-dependent and -independent manner, resulting in rebalance of plant development and heat response. Therefore, we uncover an HDA9-mediated positive regulatory module in the heat shock signal transduction pathway. More important, this cytoplasm-to-nucleus translocation of HDA9 in response to heat stress is conserved in wheat and rice, which confers the mechanism significant implication potential for crop breeding to cope with global climate warming.

HOS1 promotes plant tolerance to low-energy stress via the SnRK1 protein kinase.

Plants need to integrate internal and environmental signals to mount adequate stress responses. The NUCLEAR PORE COMPLEX (NPC) component HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES 1 (HOS1) is emerging as such an integrator, affecting responses to cold, heat, light, and salinity. Stress conditions often converge in a low-energy signal that activates SUCROSE NON-FERMENTING 1-RELATED KINASE 1 (SnRK1) to promote stress tolerance and survival. Here, we explored the role of HOS1 in the SnRK1-dependent response to low-energy stress in Arabidopsis thaliana, using darkness as a treatment and a combination of genetic, biochemical, and phenotypic assays. We show that the induction of starvation genes and plant tolerance to prolonged darkness are defective in the hos1 mutant. HOS1 interacts physically with the SnRK1α1 catalytic subunit in yeast two-hybrid assays and in planta, and the nuclear accumulation of SnRK1α1 is reduced in the hos1 mutant. Likewise, another NPC mutant, nup160, exhibits lower activation of starvation genes and decreased tolerance to prolonged darkness. Importantly, defects in low-energy responses in the hos1 background are rescued by fusing SnRK1α1 to a potent nuclear localization signal or by sugar supplementation during the dark treatment. Altogether, this work demonstrates the importance of HOS1 for the nuclear accumulation of SnRK1α1, which is key for plant tolerance to low-energy conditions.

Responsiveness to long days for flowering is reduced in Arabidopsis by yearly variation in growing season temperatures.

Conservative flowering behaviours, such as flowering during long days in summer or late flowering at a high leaf number, are often proposed to protect against variable winter and spring temperatures which lead to frost damage if premature flowering occurs. Yet, due the many factors in natural environments relative to the number of individuals compared, assessing which climate characteristics drive these flowering traits has been difficult. We applied a multidisciplinary approach to 10 winter-annual Arabidopsis thaliana populations from a wide climactic gradient in Norway. We used a variable reduction strategy to assess which of 100 climate descriptors from their home sites correlated most to their flowering behaviours when tested for responsiveness to photoperiod after saturation of vernalization; then, assessed sequence variation of 19 known environmental-response flowering genes. Photoperiod responsiveness inversely correlated with interannual variation in timing of growing season onset. Time to flowering appeared driven by growing season length, curtailed by cold fall temperatures. The distribution of FLM, TFL2 and HOS1 haplotypes, genes involved in ambient temperature response, correlated with growing-season climate. We show that long-day responsiveness and late flowering may be driven not by risk of spring frosts, but by growing season temperature and length, perhaps to opportunistically maximize growth.

Balancing selection maintains diversity in a cold tolerance gene in broadly distributed live oaks.

Cold poses major physiological challenges to plants, especially long-lived trees. In trees occurring along variable temperature clines, the expected direction and consequences of selection on cold acclimation ability and freezing tolerance are not straightforward. Here we estimated selection in cold acclimation genes at two evolutionary timescales in all seven species of the American live oaks (Quercus subsection Virentes). Two cold response candidate genes were chosen: ICE1, a key gene in the cold acclimation pathway, and HOS1, which modulates cold response by negatively regulating ICE1. Two housekeeping genes, GAPDB and CHR11, were also analyzed. At the shallow evolutionary timescale, we demonstrate that HOS1 experienced recent balancing selection in the two most broadly distributed species, Q. virginiana and Q. oleoides. At a deeper evolutionary scale, a codon-based model of evolution revealed the signature of negative selection in ICE1. In contrast, three positively selected codons have been identified in HOS1, possibly a signature of the diversification of Virentes into warmer climates from a freezing adapted lineage of oaks. Our findings indicate that evolution has favored diversity in cold tolerance modulation through balancing selection in HOS1 while maintaining core cold acclimation ability, as evidenced by purifying selection in ICE1.

Exploring the pleiotropy of hos1.

Understanding of the roles that HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENE 1 (HOS1) plays in the plant's ability to sense and respond to environmental signals has grown dramatically. Mechanisms through which HOS1 affects plant development have been uncovered, and the broader consequences of hos1 on the plant's ability to perceive and respond to its environment have been investigated. As such, it has been possible to place HOS1 as a key integrator of temperature information in response to both acute signals and cues that indicate time of year into developmental processes that are essential for plant survival. This review summarizes knowledge of HOS1's form and function, and contextualizes this information so that it is relevant for better understanding the processes of cold signalling, flowering time, and nuclear pore complex function more broadly.

The HOS1-PIF4/5 module controls callus formation in Arabidopsis leaf explants.

A two-step plant regeneration has been widely exploited to genetic manipulation and genome engineering in plants. Despite technical importance, understanding of molecular mechanism underlying in vitro plant regeneration remains to be fully elucidated. Here, we found that the HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES 1 (HOS1)-PHYTOCHROME INTERACTING FACTOR 4/5 (PIF4/5) module participates in callus formation. Consistent with the repressive role of HOS1 in PIF transcriptional activation activity, hos1-3 mutant leaf explants exhibited enhanced callus formation, whereas pif4-101 pif5-3 mutant leaf explants showed reduced callus size. The HOS1-PIF4/5 function would be largely dependent on auxin biosynthesis and signaling, which are essential for callus initiation and proliferation. Our findings suggest that the HOS1-PIF4/5 module plays a pivotal role in auxin-dependent callus formation in Arabidopsis.

Suppression of the HOS1 Gene Affects the Level of ROS Depending on Light and Cold.

The E3 ubiquitin-protein ligase HOS1 is an important integrator of temperature information and developmental processes. HOS1 is a negative regulator of plant cold tolerance, and silencing HOS1 leads to increased cold tolerance. In the present work, we studied ROS levels in hos1Cas9Arabidopsis thaliana plants, in which the HOS1 gene was silenced by disruption of the open reading frame via CRISPR/Cas9 technology. Confocal imaging of intracellular reactive oxygen species (ROS) showed that the hos1 mutation moderately increased levels of ROS under both low and high light (HL) conditions, but wild-type (WT) and hos1Cas9 plants exhibited similar ROS levels in the dark. Visualization of single cells did not reveal differences in the intracellular distribution of ROS between WT and hos1Cas9 plants. The hos1Cas9 plants contained a high basal level of ascorbic acid, maintained a normal balance between reduced and oxidized glutathione (GSH and GSSG), and generated a strong antioxidant defense response against paraquat under HL conditions. Under cold exposure, the hos1 mutation decreased the ROS level and substantially increased the expression of the ascorbate peroxidase genes Apx1 and Apx2. When plants were pre-exposed to cold and further exposed to HL, the expression of the NADPH oxidase genes RbohD and RbohF was increased in the hos1Cas9 plants but not in WT plants. hos1-mediated changes in the level of ROS are cold-dependent and cold-independent, which implies different levels of regulation. Our data indicate that HOS1 is required to maintain ROS homeostasis not only under cold conditions, but also under conditions of both low and high light intensity. It is likely that HOS1 prevents the overinduction of defense mechanisms to balance growth.

Characterization of the key region and putative phosphorylation sites of EcaICE1 in its molecular interaction with the EcaHOS1 protein in Eucalyptus camaldulensis.

Inducer of CBF expression 1 (ICE1), a MYC-like bHLH transcriptional activator, plays an important role in plants under cold stress. The ubiquitination-proteasome pathway mediated by high expression of osmotically responsive gene1 (HOS1) can effectively induce the degradation of ICE1 and decrease the expression of CBFs and their downstream genes under cold stress response in Arabidopsis, but knowledge of ubiquitination regulation of ICE1 by HOS1 is still limited in woody plants. In this study, a E3 ubiquitin ligase gene EcaHOS1 were amplified from Eucalyptus camaldulensis and the protein interactions between EcaICE1 and EcaHOS1 were analysed. Yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assay results showed that EcaICE1 can interact with the EcaHOS1 protein in the nucleus and, further, the Y2H assay demonstrated that the 126-185 amino acid region at the N-terminus of the EcaICE1 protein was indispensable for its interaction with EcaHOS1 protein. Moreover, we found that the amino acids at positions 145, 158 and 184 within the key interaction region were the putative phosphorylation sites of EcaICE1, based on bioinformatics analysis, and only the substitution of serine (Ser) 158 by alanine (Ala) blocked the protein-protein interactions between EcaICE1 and EcaHOS1 based on Y2H and ß-galactosidase activity assays using site-directed mutagenesis. We identified Ser 158 of EcaICE1 as the key putative phosphorylation site for its interaction with the EcaHOS1 protein.

CRISPR/Cas9-Mediated Knockout of HOS1 Reveals Its Role in the Regulation of Secondary Metabolism in Arabidopsis thaliana.

In Arabidopsis, the RING finger-containing E3 ubiquitin ligase HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES 1 (HOS1) functions as a main regulator of the cold signaling. In this study, CRISPR/Cas9-mediated targeted mutagenesis of the HOS1 gene in the first exon was performed. DNA sequencing showed that frameshift indels introduced by genome editing of HOS1 resulted in the appearance of premature stop codons, disrupting the open reading frame. Obtained hos1 Cas9 mutant plants were compared with the SALK T-DNA insertion mutant, line hos1-3, in terms of their tolerance to abiotic stresses, accumulation of secondary metabolites and expression levels of genes participating in these processes. Upon exposure to cold stress, enhanced tolerance and expression of cold-responsive genes were observed in both hos1-3 and hos1 Cas9 plants. The hos1 mutation caused changes in the synthesis of phytoalexins in transformed cells. The content of glucosinolates (GSLs) was down-regulated by 1.5-times, while flavonol glycosides were up-regulated by 1.2 to 4.2 times in transgenic plants. The transcript abundance of the corresponding MYB and bHLH transcription factors, which are responsible for the regulation of secondary metabolism in Arabidopsis, were also altered. Our data suggest a relationship between HOS1-regulated downstream signaling and phytoalexin biosynthesis.

Nucleoporin 160 Regulates Flowering through Anchoring HOS1 for Destabilizing CO in Arabidopsis.

Nuclear pore complexes (NPCs), which comprise multiple copies of nucleoporins (Nups), are large protein assemblies embedded in the nuclear envelope connecting the nucleus and cytoplasm. Although it has been known that Nups affect flowering in Arabidopsis, the underlying mechanisms are poorly understood. Here, we show that loss of function of Nucleoporin 160 (Nup160) leads to increased abundance of CONSTANS (CO) protein and the resulting upregulation of FLOWERING LOCUS T (FT) specifically in the morning. We demonstrate that Nup160 regulates CO protein stability through affecting NPC localization of an E3-ubiquitin ligase, HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES1 (HOS1), which destabilizes CO protein in the morning period. Taken together, these results provide a mechanistic understanding of Nup function in the transition from vegetative to reproductive growth, suggesting that deposition of HOS1 at NPCs by Nup160 is essential for preventing precocious flowering in response to photoperiod in Arabidopsis.

HOS1 acts as a key modulator of hypocotyl photomorphogenesis.

Plants recognize light as an environmental signal to determine the proper timing of growth and development. In Arabidopsis seedlings, hypocotyl growth is promoted in the dark but suppressed in the light. It is known that the red/far-red light-sensing receptor phytochrome B (phyB) suppresses the function of PHYTOCHROME INTERACTING FACTOR (PIF) transcription factors, which act as photomorphogenic repressors. However, molecular mechanisms underlying the phyB-mediated inhibition of PIF functioning remain unclear. We recently demonstrated that HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES 1 (HOS1) facilitates the phyB-mediated suppression of PIF4 during the light period to achieve hypocotyl photomorphogenesis. HOS1 inhibits the transcriptional activation activity of PIF4 by forming protein complexes. Notably, phyB-mediated light signals induce HOS1 activity, thus promoting hypocotyl photomorphogenesis. While HOS1 is known to act as an E3 ubiquitin ligase or a chromatin remodeling factor, our data illustrate a novel role of HOS1: it acts as a component of phyB-mediated light signaling in hypocotyl photomorphogenesis.

HOS1 Facilitates the Phytochrome B-Mediated Inhibition of PIF4 Function during Hypocotyl Growth in Arabidopsis.

Upon exposure to light, developing seedlings undergo photomorphogenesis, as illustrated by inhibition of hypocotyl elongation, cotyledon opening, and leaf greening. During hypocotyl photomorphogenesis, light signals are sensed by multiple photoreceptors, among which the red/far-red light-sensing phytochromes have been extensively studied. However, it is not fully understood how the phytochromes modulate hypocotyl growth. Here, we demonstrated that HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES 1 (HOS1), which is known to either act as E3 ubiquitin ligase or affect chromatin organization, inhibits the transcriptional activation activity of PHYTOCHROME INTERACTING FACTOR 4 (PIF4), a key transcription factor that promotes hypocotyl growth. Consistent with the negative regulatory role of HOS1 in hypocotyl growth, HOS1-defective mutants exhibited elongated hypocotyls in the light. Notably, phyB induces HOS1 activity in inhibiting PIF4 function. Taken together, these observations provide a molecular basis for the phyB-mediated suppression of hypocotyl growth in Arabidopsis.

Overexpression of an Orchid (Dendrobium nobile) SOC1/TM3-Like Ortholog, DnAGL19, in Arabidopsis Regulates HOS1-FT Expression.

Flowering in the appropriate season is critical for successful reproduction in angiosperms. The orchid species, Dendrobium nobile, requires vernalization to achieve flowering in the spring, but the underlying regulatory network has not been identified to date. The MADS-box transcription factor DnAGL19 was previously identified in a study of low-temperature treated D. nobile buds and was suggested to regulate vernalization-induced flowering. In this study, phylogenetic analysis of DnAGL9 and the MADS-box containing proteins showed that DnAGL19 is phylogenetically closely related to the SOC1-like protein from orchid Dendrobium Chao Parya Smile, DOSOC1. The orchid clade closed to but is not included into the SOC1-1/TM3 clades associated with either eudicots or monocots, suggesting that DnAGL19 is an SOC1-1/TM3-like ortholog. DnAGL19 was found to be highly expressed in pseudobulbs, leaves, roots, and axillary buds but rarely in flowers, and to be substantially upregulated in axillary buds by prolonged low-temperature treatments. Overexpression of DnAGL19 in Arabidopsis thaliana resulted in a small but significantly reduced time to bolting, suggesting that flowering time was slightly accelerated under normal growth conditions. Consistent with this, the A. thaliana APETELA1 (AP1) gene was expressed at an earlier stage in transgenic lines than in wild type plants, while the FLOWERING LOCUS T (FT) gene was suppressed, suggesting that altered regulations on these transcription factors caused the weak promotion of flowering. HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENE 1 (HOS1) was slightly activated under the same conditions, suggesting that the HOS1-FT module may be involved in the DnAGL19-related network. Under vernalization conditions, FT expression was significantly upregulated, whereas HOS1 expression in the transgenic A. thaliana has a level similar to that in wild type. Taken together, these results suggest that DnAGL19 controls the action of the HOS1-FT module depending on temperature cues, which could contribute to regulation of D. nobile flowering time. These data provide insights into how flowering is fine-tuned in D. nobile to acclimate the plant to seasonal changes in temperature.

The E3 ubiquitin ligase HOS1 is involved in ethylene regulation of leaf expansion in Arabidopsis.

Ethylene regulates a variety of physiological processes, such as flowering, senescence, abscission, and fruit ripening. In particular, leaf expansion is also controlled by ethylene in Arabidopsis. Exogenous treatment with ethylene inhibits leaf expansion, and consistently, ethylene insensitive mutants show increased leaf area. Here, we report that the RING finger-containing E3 ubiquitin ligase HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES 1 (HOS1) regulates leaf expansion in an ethylene signaling pathway. The HOS1-deficient mutant showed reduced leaf area and was insensitive to ethylene perception inhibitor, silver thiosulfate (STS). Accordingly, genes encoding ethylene signaling components were significantly up-regulated in hos1-3. This study demonstrates that the HOS1 protein is involved in ethylene signal transduction for the proper regulation of leaf expansion possibly under environmentally stressful conditions.

Beyond ubiquitination: proteolytic and nonproteolytic roles of HOS1.

The E3 ubiquitin ligase HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENES 1 (HOS1) functions as a cold signaling attenuator by degrading the INDUCER OF CBF EXPRESSION 1 transcription factor, which is a key regulator of the cold-induced transcriptome and freezing tolerance in plants. Recent studies demonstrate that HOS1 also plays nonproteolytic roles in gene expression regulation. HOS1 acts as a chromatin remodeling factor that modulates FLOWERING LOCUS C chromatin in cold regulation of flowering time. It associates with the nuclear pore complex to facilitate nucleocytoplasmic mRNA export to maintain circadian periodicity over a range of light and temperature conditions. In this review, we summarize recent advances in molecular mechanisms underlying HOS1 function during plant development in response to fluctuating environmental conditions.

Controlled turnover of CONSTANS protein by the HOS1 E3 ligase regulates floral transition at low temperatures.

The timing of flowering is coordinately regulated by complex gene regulatory networks that integrate developmental and environmental cues. Light and temperature are major environmental determinants in flowering time control. Temperature signals include two major categories: ambient temperature signals and cold nonfreezing temperature signals. Notably, the effects of cold temperatures on flowering timing are profoundly differentiated, depending on the duration of cold exposure. Whereas long-term exposure to cold temperatures, designated vernalization, promotes flowering, short-term cold exposure delays flowering. Genes constituting the vernalization pathway and underlying molecular mechanisms have been extensively studied. However, how cold stress signals delay flowering is largely unknown. We have recently reported that the HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENE 1 (HOS1)-CONSTANS (CO) module is at least partly responsible for the daily sensing of cold stress signals in flowering time control. Intermittent cold stress triggers the degradation of CO, a central activator of photoperiodic flowering, via a ubiquitination pathway that involves the HOS1 E3 ubiquitin ligase, leading to suppression of FLOWERING LOCUS T (FT) gene and delayed flowering. It is proposed that CO serves as a molecular knot that integrates photoperiod and temperature signals into the flowering pathways, fine-tuning photoperiodic flowering under short-term temperature fluctuations.

HOS1-mediated activation of FLC via chromatin remodeling under cold stress.

The Arabidopsis E3 ubiquitin ligase HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENE 1 (HOS1) has been shown to act as a negative regulator of cold responses by degrading the INDUCER OF CBF EXPRESSION 1 (ICE1) transcription factor through the ubiquitin/proteasome pathway. Notably, loss-of-function hos1 mutants exhibit early flowering, and the transcript level of the floral repressor FLOWERING LOCUS C (FLC) is downregulated in the mutants. However, it is largely unknown how HOS1 regulates FLC transcription. We found that HOS1 activates FLC transcription by inhibiting the activity of histone deacetylase 6 (HDA6) under cold stress. Cold temperatures induce the binding of HOS1 to FLC chromatin in an FVE-dependent manner. Cold-activated HOS1 promotes the dissociation of HDA6 from FLC chromatin, and the cold effects disappear in both hos1 and fve mutants. It is therefore clear that HOS1 regulates FLC transcription via chromatin remodeling, providing new insights into the signaling crosstalks between cold response and flowering time control.

Suppression of Arabidopsis RING E3 ubiquitin ligase AtATL78 increases tolerance to cold stress and decreases tolerance to drought stress.

AtATL78 is an Arabidopsis RING E3 ubiquitin ligase. RT-PCR and promoter-GUS assays revealed that AtATL78 was up-regulated by cold stress and down-regulated by drought. AtATL78 was localized at the plasma-membrane. Suppression of AtATL78 increased tolerance to cold stress but decreased tolerance to drought. Our data suggests that AtATL78 is a negative regulator of cold stress response and a positive regulator of drought stress response in Arabidopsis. These results further suggest that AtATL78 plays opposing roles in cold and drought stress responses.