GI-CO-FT confers microgravity response through Potential Regulatory Modules of Arabidopsis plants grown in Space

doi:10.21203/rs.3.rs-3803812/v1

Our previous studies suggested that FLOWERING LOCUS (FT) can serve as a central hub, integrating microgravity signals into the photoperiodic flowering pathway in Arabidopsis. However, the exact mechanisms remain unclear. In this study, we present findings from our recent experiments on the Chinese Space Station. Transcriptomic changes of space-grown plants, including wild-type (WT), gigantea (gi) mutant, CONSTANS overexpressing plants (35S:CO), and transgenic plants expressing FT with the green fluorescent protein gene controlled by a heat shock-inducible promoter (pHSP:GFP;pHSP:FT, FG), were compared with their ground-based controls. Analysis revealed a conserved transcriptional response signature primarily across the genotypes investigated. We pinpointed a new set of microgravity-responsive genes with a significant enrichment in flowering controlling pathways. In addition, this study highlighted bZIP, bHLH, BES1, and SBP family transcription factors, which are known integrators of hormone, light, and photoperiodic signals, as key mediators of the microgravity response in space.

Biological sciences/Plant sciences/Plant stress responses/Abiotic

Biological sciences/Molecular biology/Transcriptomics

Microgravity

GI-CO-FT pathway

flowering

Arabidopsis thaliana

Transcriptomics

RNA seq

Chinese Space Station

For extended human space exploration, a tightly integrated biological life support system (BLSS) is imperative, with plants playing a pivotal role in air regeneration, food production, and water recycling. Given the constrained volume and energy supply aboard space missions designated for plant cultivation, there's a pressing need not just to successfully grow edible plants in space, but also to optimize the harvest index while minimizing production time and energy consumption. However, spaceflight presents a special environment with complex stressors, including microgravity and radiation, greatly affecting plant survival and adaption. For example, many plant seedlings grown in space have been observed to be smaller than their ground controls ^1–3, while some exhibited accelerated growth ^4–5. Challenges such as reduced shoot fresh weight, compromised photosynthetic functions⁶, alterations in seed reserves^7–8, and delays in the reproductive phase⁹ are significant hurdles to utilizing plants in long-term manned space missions^5,10–11.

Spaceflight microgravity leads to physiological, morphological, and molecular alterations in plants, including the changed transcription patterns of many genes^2,12. Gene Ontology (GO) analyses of spaceflight-affected genes reported in previous studies include categories like stress responses, photosynthesis, cell wall reorganization and secondary metabolism remodeling. A growing body of evidence underscores the influence of microgravity on the flowering processes of space-grown plants^11,13. Many plants in space matured more slowly, with reproductive phases often compromised^13–14. This challenge was attributed to pollination or fertilization hindrances in Arabidopsis due to high humidity in the closed chamber in space. Advancements in cultivation chambers have now facilitated successful seed production in both dicotyledonous and monocotyledonous plants during space missions^15–18. Despite these advancements, space plants still confront abiotic stressors induced directly or indirectly by microgravity, influencing their developmental processes. For instance, they produced fewer flowers, altered inflorescence structures, and smaller seeds with changed storage reserves compared to their controls on ground.

In earlier studies, we have studied the interplay between FLOWERING LOCUS T (FT) induction and photoperiodic signaling in space-grown Arabidopsis plants^9,11. Floral initiation in space-grown Arabidopsis under long-day (LD) conditions was observed to be delayed about 20 days compared to ground controls. Using transgenic plants with heat shock-inducible FT expression, we showed that plant transcriptional responses to microgravity are related to day-length conditions, and inducing FT expression during the reproductive phase could mitigate spaceflight stresses, thereby influencing reproductive development¹¹. However, the specific mechanisms by which microgravity influences plant development and flowering remain unknown.

Arabidopsis flowering was quickly triggered by LD. The LD pathway is characterized by three pivotal components: GIGANTEA (GI), CONSTANS (CO), and FT (GI-CO-FT pathway¹⁹). Both GI and CO serve as nexus points connecting various stress pathways with flowering. The gi knockout mutants were observed to flower late and exhibited a higher tolerance to abiotic stresses^20–22. In contrast, overexpressing CO led to early flowering and diminished stress tolerance^23–24. To know the regulatory mechanism of the GI-CO-FT pathway in spaceflight adaptation, we present findings from our most recent experiments aboard the Chinese Space Station (CSS), where Arabidopsis wild-type (WT), gi, CO overexpression plants (35S:CO), and transgenic plants expressing FT with the green fluorescent protein gene controlled by a heat shock-inducible promoter (pHSP:GFP; pHSP:FT, named “FG”)¹¹ were cultivated and their transcriptomic adjustments in space were analyzed by comparing with those of their controls on ground.

Global Transcriptome Analysis Reveals the Role of the GI-CO-FT Pathway in Response to microgravity

Arabidopsis plants were cultivated in the General Biological Modular Culture System (GBMCS) aboard the CSS (Fig. 1a, b). The GBMCS is equipped with subsystems for atmospheric, temperature, and hydration regulation and a lighting system (Fig. 1c). Additionally, the GBMCS is fitted with four video cameras to facilitate image capture and to visually monitor the ongoing experiment. Seeds were germinated and grown in two culture chambers (CCs), CC1 and CC2, which were installed in GBMCS under microgravity onboard the CSS and under normal gravity on the ground, respectively (Fig. 1d, e). The images of Arabidopsis were taken by inflight cameras once every two hours to record the processes of growth and development (Fig. 2a-c; Supplementary Fig. 1). Ground control experiments were carried out in parallel with CCs in an engineering reproduction of the spaceflight GBMCS.

Compared with the ground controls, the time of bolting of WT plants grown in space appeared to be significantly delayed (Supplementary Fig. 1) as our previous reported⁹. To determine whether the microgravity response observed in WT was mediated by the key flowering time genes, the different flowering time plants that are representative of the photoperiodic floral pathways were exposed to microgravity in space. Mutant gi flowers late, while 35S:CO transgenic plants flower early relative to the WT. FG transgenic plants, which had been constructed in our previous study, flower early under a heat shock condition¹¹. Downloaded images revealed that seedlings of WT, gi, 35S:CO and FG in CCs under microgravity in space and on ground all germinated well at day 3 after sowing. After germination, WT, gi, 35S:CO and FG plants developed rosette leaves under either in space or on ground conditions (Fig. 2c). Plants of all genotypes in space grew slowly with smaller cotyledon. Seedlings of gi mutant grown in space exhibited apparently elongated hypocotyls in comparison with their controls on ground, indicating a positive role of GI in inhibiting elongation of hypocotyls under microgravity condition. Mutant gi both grown in space and on ground didn’t flower during 25 days after sowing (Fig. 2d). Because there was not enough growth time on orbit, we couldn’t rule out the effect of microgravity on the flowering of gi mutant in space. 35S:CO transgenic plants were consistently early flowering both under microgravity in space and on the ground condition, while FG plants displayed a slight earlier in flowering under microgravity in space than those on ground (Supplementary Fig. 1 and Fig. 2). These results revealed a role of CO and FT to activate microgravity response in regulating Arabidopsis flowering in space.

To capture the global transcriptional impacts of microgravity under spaceflight on Arabidopsis growth, we undertook RNA sequencing (RNA seq) of leaves from plants grown in space and compared them to their ground controls. RNA was extracted from leaves of plants at day 25 after sowing in both CCs-space (sp) and CCs-ground (gr). The mean expression levels of genes in the space-grown samples were significantly different from those of the ground controls when we applied a false discovery rate (FDR) threshold of 0.01; these genes were selected as potential differentially expressed genes (DEGs) (Supplementary Table 1). Among these genes, we further refined our list by considering those with expression levels that varied more than five-fold (fold change [FC] ≥ 5) when comparing space samples to their ground controls. These analyses revealed that 4901 genes in WT, 4952 genes in gi mutant, 3871 genes in 35S:CO, and 3832 genes in FG transgenic plants were differentially expressed (Fig. 3a; Supplementary Table 2). To validate these RNA-seq data, we designed sequence-specific primers and performed real-time quantitative PCR (qRT-PCR). qRT-PCR for differential expression levels of selected genes, NUDT 21 (AT1G73540), DIC2 (AT4G24570) and glycine-rich protein (AT1G07135), match with the results analyzed by RNA-seq (Supplementary Fig. 3 and Table 2). In all genotypes, the proportion of up-regulated genes in response to spaceflight conditions consistently higher than that of down-regulated genes (Fig. 3a, b; Supplementary Table 2). Principal Component Analysis (PCA) highlighted marked differences between the transcriptomes of space-grown samples and their ground controls (Fig. 3c), suggesting the regulatory roles of GI, CO, and FT in microgravity responses of Arabidopsis plants.

To understand the biological roles of these DEGs in response to microgravity, we undertook GO analysis for WT, gi, 35S:CO, and FG. We emphasized the top ten GO enrichment terms for both up- and down-regulated genes. Predominantly, up-regulated genes were associated with oxidant detoxification, while down-regulated genes were linked to the photosynthetic process, and these trends were consistent across gi, 35S: CO, FG, and WT (Fig. 4; Supplementary Table 4).For a deeper understanding of the GI-CO-FT module's impact on plant microgravity response, we compared the DEGs involved in flowering time regulation from gi, 35S:CO, and FG with those from WT. Expression of 1517 genes in gi, 798 genes in 35S:CO, and 873 genes in FG showed similarities with WT in response to microgravity in space (termed the "common-sp" group, Fig. 5a; Supplementary Table 5), while a larger number of DEGs in gi, 35S:CO, and FG exhibited different behaviors from what they did in WT. Specifically, 4844 genes in gi, 5427 genes in 35S:CO, and 5224 genes in FG demonstrated different expression responses to microgravity in space compared to those in WT (named 'df-sp' group, Fig. 5a; Supplementary Table 5). A significant overlap, constituting about 40% of the total, was observed among the "df-sp" DEGs from gi, 35S:CO, and FG (Fig. 5b).

The ten most significantly enriched GO terms for 'df-sp' genes in gi, 35S:CO, and FG primarily involved the photosynthetic process and responses to light stimuli for up-regulated genes, while down-regulated genes were associated with oxidant detoxification processes (Fig. 5c). Furthermore, we also identified genotype-specific GO term enrichments. For instance, gi-associated terms involved secondary metabolic processes and transmembrane transport, 35S:CO was linked to carbohydrate and auxin metabolic processes, and FG showed enrichment in UV response and chlorophyll metabolism. Importantly, subcellular localization of all DEGs across gi, 35S:CO, and FG revealed a pronounced association with chloroplast-related genes (Supplementary Table 6).

The response of flowering pathway genes to microgravity in space

Our previous studies demonstrated that the spaceflight environment notably decreased the expression of FT^9,11. This suggests that the observed flowering delay in Arabidopsis plants in space might be attributed to modifications in the photoperiodic flowering network, where microgravity stress signals converge on FT expression. To gain a comprehensive view of the changed expression of flowering time genes among the whole genome transcriptional response to microgravity in space, DEGs that were involved in different flowering control pathways were identified. The number of spaceflight-responsive flowering genes identified in WT, gi, 35S:CO, and FG were 134, 132, 127, and 143 respectively (Supplementary Table 7). We categorized these transcripts based on their established or predicted functions in the flowering regulation pathway²⁵. Most of these flowering genes regulated by microgravity in space were functionally categorized to be related to the photoperiodic response (44 genes, accounting for 28% of all DEGs in the flowering pathway). Subsequent classifications highlighted genes functioning in general (34 genes, 22%), hormone regulation (19 genes, 12%), the circadian clock (15 genes, 9%), vernalization (12 genes, 8%), flower development and meristem identity (13 genes, 8%), aging (12 genes, 8%), sugar metabolism (5 genes, 3%), and ambient temperature response (4 genes, 3%). This indicates that microgravity response signaling pathways concurrently integrate into various flowering regulation pathways (Fig. 6).

The circadian clock, an endogenous timekeeping mechanism, enables plants not only to respond to environmental changes but to predict and adapt to upcoming changes. Our previous space experiments suggested that the circadian oscillator might integrate spaceflight responses with photoperiodic signals in Arabidopsis grown in space¹¹. In this study, clock transcriptional repressors (CIRCADIANCLOCKASSOCIATED1(CCA1) and LATE ELONGATED HYPOCOTYL (LHY)) were upregulated, while transcriptional activators (Arabidopsis pseudo-response regulator 3 (APRR3) and Arabidopsis pseudo-response regulator 7 (APRR7)) were downregulated in all four genotypes in space, compared to their ground controls. The expression of TIMING OF CAB EXPRESSION 1 (TOC1) was downregulated in WT but remained unchanged in gi, 35S:CO, and FG. In contrast, Arabidopsis pseudo-response regulator 9 (APRR9) expression was consistent in WT but upregulated in gi, 35S:CO, and FG. These findings suggest that the circadian oscillator participates in regulating the Arabidopsis microgravity response in both GI-CO-FT dependent and independent manners.

Microgravity resulted in altered expression levels of photoperiod-responsive genes, with 22 upregulated and 12 downregulated in WT (Fig. 6). For example, the expression of basic helix-loop-helix (bHLH63) in spaceflight samples was downregulated in WT but upregulated only in 35S:CO, with no changes observed in gi mutants and FG plants in comparison with their controls on ground. Notably, FLAVIN-BINDING, KELCH REPEAT 1 (FKF1) remained unaltered in 35S:CO but was upregulated in WT, gi, and FG. Both bHLH63 and FKF1 have been reported to interact with CO to regulate flowering^26–27. Thus, spaceflight-induced alterations in bHLH63 and FKF1 expression levels, especially in 35S:CO, suggest a potential co-regulation between bHLH63-FKF1 and CO in the regulation of photoperiodic flowering in response to microgravity in space.

In the Gibberellin (GA) pathway, spaceflight caused variations in the transcription levels of GA "activating enzymes", GIBBERELLIN 3-OXIDASE 1 (GA3OX1), GA20OX3, and DELLA proteins RGA-LIKE PROTEIN 3 (RGL3) in WT, while no effect was observed in the gi mutant. GA2OX3 and RGL3 were upregulated, whereas GA INSENSITIVE DWARF1A (GID1A) and GA20OX1 were downregulated specifically in 35S:CO plants. In contrast, three GA "activating enzymes" GA20OX1/2/3 notably changed transcription levels in FG in response to microgravity in space (Fig. 6 and Fig. 7).

Regarding the vernalization pathway, the expression of genes like WRKY34, FRIGIDA LIKE 2 (FRL2), SUPPRESSOR of FRI 4 (SUF4), and VERNALIZATION INSENSITIVE 3 (VIN3) displayed unconsistent variations in gi, 35S:CO, and FG in response to spaceflight compared to those in WT. WRKY34 contains two WRKY domains found in the promoters of numerous stress-related plant genes²⁸. All of these genes are essential for regulating flowering in response to vernalization (Fig. 7).

In the aging pathway, spaceflight caused notable changes in the expression levels of 9 genes in WT. Among these genes, miR156 a/c, TARGET OF EARLY ACTIVATION TAGGED (EAT) 2 (TOE2) and TOE3, and SCHLAFMUTZE (SMZ) were upregulated, while SQUAMOSA PROMOTER BINDING PROTEIN-LIKE 3 (SPL3), SPL4, and SPL5 were downregulated (Fig. 6). MiR156 is known to mediate the repression of SPLs and inhibit Arabidopsis floral inductive signals²⁹. The expression of SPL5 in gi, 35S:CO and FG plants grown in space remained unchanged in comparison with their controls on ground, but SCHNARCHZAPFEN (SNZ) was apparently downregulated specifically in gi compared to WT. Conversely, SPL5 was upregulated, and SPL9 was downregulated in 35S:CO in response to microgravity in space (Fig. 6 and Fig. 7).

In summary, spaceflight profoundly influences the expression of key genes in floral pathways. The GI-CO-FT module plays a significant role in the response of Arabidopsis to microgravity in space.

Patterns of cis-element enrichment unveil potential regulatory modules in the transcriptional control of GI-CO-FT pathway in response to microgravity in space

To determine if variations in the expression of GI, CO, and FT influence plant responses to microgravity in space, we employed plant regulomics analyses (bioinfo.sibs.ac.cn, P_adjust<0.01) to find overrepresented motifs in the 2-kb upstream sequence (-2000 to + 100) of spaceflight DEGs in WT, gi, 35S:CO, and FG. A total of 214 potential cis-elements correspond to 22 transcription factor (TF) families were found. These corresponded closely with known elements linked to stress-responsive and developmental TFs, such as ethylene responsive factor (ERF), Basic (region) leucine zippers (bZIP), bHLH, MYB, Squamosa promoter Binding Protein (SBP), and WRKY (Table 1, Fig. 8, Supplementary Table 8). Of these cis-elements, 79% (169 members) were observed across all four genotypes (Fig. 8). Motifs highly enriched (Enrichment score > 150% and P_adjust <0.01) among spaceflight-regulated genes prominently corresponded to ERF, bZIP, bHLH, and BES1 transcription factors (Table 2).

Based on enrichment scores across the four genotypes, these potential spaceflight-responsive TFs can be divided into two groups. Group I encompass 26 identified TFs all from the ERF family, exhibiting high enrichment scores across genotypes. This suggests a universal microgravity-response regulatory module among the four genotypes. Group II contains specific subsets of genes (e.g., BRI1-EMS-SUPPRESSOR1 (BES1), bHLH, bZIP, ERF, and SBP) that showed high enrichment scores in one or more of the genotypes (Table 2). Notably, five ERF members (ERF027, COOPERATIVELY REGULATED BY ETHYLENE AND JASMONATE 1 (CEJ1), RELATED TO AP2 1 (RAP2.1), DEHYDRATION RESPONSE ELEMENT-BINDING PROTEIN 26 (DREB26) and ERF015) and four bZIP members (ABA-RESPONSIVE ELEMENT BINDING PROTEIN 3 (AREB3), ABSCISIC ACID RESPONSIVE ELEMENT-BINDING FACTOR 1 (ABF1), bZIP28 and Abscisic acid-insensitive (ABI5)) showed enrichment in gi, 35S:CO, and FG, but not in WT. This implies a GI-CO-FT associated regulatory module in response to microgravity in space. Remarkably, 35S:CO showed a greater number of specific enriched TFs in microgravity response compared to gi and FG (Table 2).

The interaction network between microgravity regulated cis-element TFs and regulators of flowering induction

The blueprints for development and response to environment in plants are encoded by cis-regulatory elements (CREs)³⁰, which are often provides an indirection for co-regulation among particular sets of genes. To explore the potential co-regulation of the microgravity response and the flowering induction in space, TFs corresponding to a set of CREs highly enriched among microgravity responsive genes in gi, 35S:CO, and FG, respectively were identified (Group II in Table 2). Within the clock pathway, 15 clock pathway genes were potentially regulated by 12 interconnected TFs in Group II (Fig. 9a). The phytochrome-interacting factor PHYTOCHROME-INTERACTING FACTOR7 (PIF7) emerged as a central interactor of microgravity-responsive clock pathway proteins. Additionally, interactions encompassing the Abscisic Acid (ABA) signaling pathway (namely ABI5 and ABF1 in Group II) were identified. These findings suggest that light regulation and ABA signaling might influence the clock pathway in response to microgravity in space.

For the photoperiod pathway, 13 cis-element TFs in Group II emerged as interactors (Fig. 9b). Here, PIF1, PIF7, and BZR2 showed interactions with phytochromes (PHYTOCHROME A (PhyA), PhyB, PhyC, and PhyD) and cryptochromes (CRYPTOCHROME 1 (CRY1) and CRY2). Meanwhile, ABI5 and ABF1 engaged with nuclear factor Y (NF-YA1, NF-YC3, and NF-YC4) TFs. This pattern indicates a microgravity-induced interplay between light, ABA, and BR signals in modulating the photoperiod pathway.

In the GA pathway, PIF1, BZR2, ABI5, and ABF2 in Group II (Table 2) showed interactions with GA hormone signaling pathway DELLA proteins (GIBBERELLIC ACID INSENSITIVE (GAI), RGL1, RGL2, RGL3, RGA) and with GA enzymes (GA20OXs, GA2OXs, and GA3OXs) (Fig. 9c). These findings hint a possible crosstalk among GA, ABA, BR and light signaling pathways in response to microgravity in Arabidopsis during flowering in space.

In aging pathway, SPL11, which was highly enriched specifically in 35:CO in response to microgravity (Table 2), showed interactions with TARGET OF EAT 2 (TOE2), TOE3, SMZ, and SNZ (Fig. 9d). TOE2/TOE3 were reported to be controlled by the miR156-SPL regulation module of developmental timing^31–32, while SMZ and SNZ are important regulators of flowering time in response to nitrate³³. This result indicates that SPL11 could play as an important role in regulating expression of TOE2/TOE3 and SMZ/SNZ to control flowering of Arabidopsis plants in space.

Plants exhibit a pronounced developmental plasticity in response to environmental signals, including microgravity, which could trigger multiple stressors⁹. In a previous study, we discovered that FT can serve as a hub, integrating spaceflight stress signals into the photoperiodic flowering pathway in space-grown Arabidopsis. However, the precise mechanism by which FT regulates the microgravity response remained elusive. In our current investigation, we conducted transcriptional analyses comparing microgravity responses in various Arabidopsis flowering time genotypes, namely gi mutant (late flowering), 35S:CO (constitutionally early flowering), and FG (stress inducible early flowering), against the WT (Supplementary Fig. 1). Our data reveal the modifications in the specific microgravity response and common microgravity response (Fig. 10). Moreover, several genes tied to specific flowering induction processes primarily or solely responded to microgravity in gi, 35S:CO, and FG, relative to WT. Furthermore, potential regulators associated with ERF, bZIP, bHLH, and BES1 TFs, along with interaction networks between these microgravity response cis-element TFs and pivotal flowering induction regulators, also play roles in modulating the GI-CO-FT module in space.

Microgravity poses a unique stress type that impacts the absorption, transport, and distribution of water and nutrients in plants grown in space, as well as their photosynthesis^3,34–35. Existing databases are available for plant response to various spaceflight conditions^36–37. They showed that the removal gravity by spaceflight evokes notable stress responses, altering plant growth and initiating vast changes in gene expression linked to primary metabolism, detoxification of reactive oxygen species, heat-shock stress, and photosynthesis in both seedlings^38–40 and mature plants during flowering^9,11. Yet, our understanding of how microgravity influences the flowering time control pathway is limited. In our study, GO group analyses highlighted responses common across WT, gi, 35S:CO, and FG, involving metabolic processes (e.g., phenylpropanoid, flavonoid, glutathione) tied to detoxification and phosphate starvation in up-regulated groups, with photosynthesis being dominant in down-regulated groups. Notably, the most overrepresented GO terms for differentially expressed genes in gi, 35S:CO, or FG versus WT were photosynthetic processes, light stimuli (e.g., light intensity), and oxidant detoxification processes (e.g., H₂O₂ catabolic and metabolic processes), revealing a pronounced enrichment in chloroplast-related genes (Fig. 4c).

The chloroplast, essential for photosynthesis and sugar production, communicates with the nucleus via retrograde signaling (RS), which manages the expression of photosynthesis-associated nuclear genes in response to various environmental factors^41–42. One primary chloroplast RS pathway is associated with reactive oxygen species produced by the photosynthetic electron transport chain⁴³. Recent research showed that mutants with defects in chlorophyll biosynthesis and chloroplast development experienced premature flowering in Arabidopsis under long-day conditions. This effect was attributed to the impaired ability of CO to activate FT⁴⁴. Overexpressing certain photoperiod-sensitive genes in rice, such as OsDREB1 and OsNF-YB2, not only hastened flowering but also boosted crop yields by enhancing nitrogen utilization and photosynthesis⁴⁵. Chloroplasts serve as pivotal light intensity sensors influencing flowering⁴⁴ and as gravity signal sensors in gravity-sensing cells⁴⁶. Past space studies reported that genes related to light perception and photosynthesis were significantly downregulated in Arabidopsis under microgravity conditions^44,47. Our findings present an intriguing link between the regulation of the GI-CO-FT pathway and chloroplast gene expression under microgravity. Adjustments in the GI-CO-FT flowering pathway could potentially impact chloroplast function, thereby balancing plant growth and development in space.

This study identified 22 TF families implicated in the GI-CO-FT controlled flowering response to microgravity. A set of genes influenced by microgravity across WT, gi, 35S:CO, and FG was prominently enriched only with motifs from the ERF family, specifically ERF groups A2, A4, A5, and B1, B2, B3, B4, and B6 (Table 2). Previous investigations have shown that the concentration of ethylene in growth chambers increased markedly due to disrupted convective air movement, which hindered plant reproduction in space¹⁶. Technological advancements in growth chamber environments have addressed the ethylene issue, but the enclosed gaseous environment surrounding plant tissues in microgravity could still stress the plants^7,48. In our research setting, ethylene was eliminated from the growth chamber. The pronounced enrichment of ERF family motifs, common across spaceflight responses, might be also associated with the enclosed gaseous microenvironment surrounding plant tissues under microgravity in space.

Moreover, cis-elements linked with ERF, bZIP, bHLH, and BES1 TFs displayed specific high enrichment in gi, 35S:CO, and/or FG, but not in WT (Table 2). Elements connected to bZIP, bHLH, BES1, and SBP transcription factors appear to play central roles in regulating the GI-CO-FT module's specific microgravity response. Meanwhile, several ERF TFs (e.g., ERF4, ERF11, ERF13, ERF 19) might be interacting with bZIP TFs in regulating the flowering process under microgravity (Fig. 8). The prevalence of elements linked to crucial TFs underscores the conservation of a key subset of TFs that respond to microgravity, contingent upon the GI-CO-FT module during the flowering process. The enrichment of elements like ABF1, ABF2, ABI5, BES1, SPL11, PIF1, and PIF7, specific to gi, 35S:CO, and FG, indicates the significance of ABA, BR, and light signaling in modulating the GI-CO-FT flowering pathway under microgravity stress in space. This offers the hypothesis that optimizing hormone and light signaling will control microgravity response of and enhance crop productivity in space.

It's worth noting that for our transcription analysis, plant samples were collected in orbit and frozen at -80°C before examination. Thus, the DEGs we identified above could represent those microgravity responsive genes, without being affected by altered gravity during landing. Our knowledge of the subject about flowering in space begin to increase in recent years due to the increase of long-term space experiments from seed-to-seed^9,11,18. However, effective models for effects of microgravity on flowering in space have not been established yet. This study contributed to identify, bZIP, bHLH, BES1 and SBP family transcription factors as integrators of microgravity response and the control of flowering signaling pathways.

Plant materials and spaceflight experiments

The following Arabidopsis thaliana samples were used: the wild-type (WT, ecotype Columbia-0, Col-0), the GI (At1g22770) mutant gi-2 ⁴⁹ ( gi ), the CO overexpression line (35S:CO in a Col-0 background⁵⁰), and the transgenic plants pHSP:FT; pHSP:GFP (FG), which were developed by Wang et al¹¹. Seed preparation and planting in two culture chambers (CCs) as described by Wu et al⁵¹ and Xie et al⁹. In short, seeds underwent a sterilization process which began with a 1-minute exposure to 75% (v/v) ethanol, followed by a 20-minute soak in a 2% (v/v) NaClO bleach solution containing 0.01% (v/v) Triton X-100 detergent. Subsequent to five sterile water rinses, seeds were attached to cheesecloth pieces (5×10 mm, ten seeds per piece) with sterile 0.1% (v/v) methyl cellulose solution. These seeds were then dried in a flow bench and stored for the spaceflight experiments.

The General Biological Modular Culture System (GBMCS) was used for the spaceflight experiments aboard the Chinese Space Station (CSS) (Fig. 1a and b) and is equipped with subsystems for atmospheric, temperature, and hydration regulation and a lighting system (Fig. 1c). Additionally, it is fitted with four video cameras to facilitate image capture and to visually monitor the ongoing experiment. Arabidopsis plants were cultured in CC1 and CC2. Each CC measured 60 × 300 × 260 mm externally. The internal space measured 60 × 200 × 210 mm, while the root module was 60 × 200 × 50 mm (Fig. 1d). The cheesecloth seed carriers were set in holes on a panel of root modules, which filled with commercially available vermiculite (Fig. 1e). Prior to the launch of the CSS's Wentian experiment module on 24 July, 2022, the seeds in the CCs were stored in a dry state for approximately 45 days. The initiation of the space experiment occurred on 29 July, 2022, with the hydration of the seeds, controlled remotely from Earth. This date was designated as Day 0. At the start of the experiment, 280 ml of sterile MS medium (PhytoTech, Beijing XMJ Scientific Co. Ltd) was supplied via a pump system (Fig. 1c). The germinated plants were grown at 22 ± 1℃, under the light from white/red (2:1) LEDs with an intensity of 120 µmol m^− 2 s^− 1 on the surface plane of the root module for a LD photoperiod (16h light/8h darkness) within the GBMCS. Inside the CCs, relative humidity was kept between 88–98%. Readings for temperature, humidity, and light levels were taken every minute throughout the flight. The GBMCS environment was maintained by a slow air exchange system (20 mL min^− 1) that directed filtered air from an external system into the CCs. The system's atmospheric pressure was stabilized at one atmosphere. The growth chamber's ethylene concentration was removed following the methods in Zheng et al⁵². For ground control experiments, a GBMCS facility mirroring the one used in space was set up. The commencement of ground experiments was on 6 September, 2022 in Shanghai. The ground control experiment parameters were set to match the data collected from the space experiment.

For RNA-Seq analysis, the rosette leaves from plants were harvested 25 days post-imbibition (Fig. 2) and subsequently frozen within designated sample vials. These vials were stored in the Experiment Refrigerator (-80℃) onboard the CSS. Prior to their return to Earth, these samples were transferred to a low-temperature freezer (-20℃) aboard the Shenzhou − 14 spacecraft on 4 Dec. 2022. Once the frozen seedlings reached Earth, they were immediately transported on dry ice and promptly preserved using RNALater solution (ambion, life technologies, CA, USA) for subsequent RNA-seq analyses.

RNA extraction and sequencing

RNA was individually extracted from each sample vial (six samples were obtained to collect approximately 8–12 seedlings each extraction). Following the extraction, total RNA from the combined samples was isolated. Subsequent to DNA removal, the quantity and quality of RNA were determined using the Nanodrop 2000. mRNA isolation was achieved through the use of Oligo(dT) beads. Once fragmented mRNA (300bp) was prepared, an RNA-seq library was constructed. Sequencing was executed using the Illumina Novaseq 6000 system. The RNA-seq generated 165.75 Gb of clean data with an average quality score (Q30) exceeding 94% and an average yield of 6.03 Gb of clean data per sample (Supplementary Table 1). The quality of the raw data was checked to ensure the reliability of the high-throughput sequence data. Approximately 98% of the cleaned reads were mapped to the Arabidopsis TAIR10 genomes using either TopHat2 (available at http://tophat.cbcb.umd.edu/) or HISAT2 (available at http://ccb.jhu.edu/software/hisat2/index.shtml). About 90% of these reads were uniquely mapped, counted, and subsequently used for gene-level differential expression analysis with the R software for statistical computations. For further analysis, only those features with a read count exceeding one per million in at least three samples were retained. This resulted in a total of 28,080 genes expressed across all conditions (Supplementary Table 1). Differentially expressed (DE) genes were identified by controlling the false discovery rate (FDR) at 1% and filtering for log₂ fold change (FC) values either below or exceeding 2.3. Gene Ontology (GO) enrichment analyses were conducted using the GOC website (http://geneontology.org).

Real-Time qRT-PCR

Total RNA was extracted from the leaves of both space and ground samples as previously detailed. The genes under study, along with their respective qRT-PCR primers, are listed in Supplementary Table 3. The Arabidopsis ACTIN gene served as the loading control for all real-time qRT-PCR experiments. Each biological replicate was represented by at least four technical replicates in the real-time qRT-PCR analyses.

Competing interests

The authors declare no competing interests.

Author contributions

HZ: Conceptualization, funding acquisition, project administration, resources, supervision, validation, writing- original draft, review and editing. JX and JZ: Data curation, formal analysis, and investigation. LW, YJ, YD, ZZ, BL and MH: carried out space experiment of plant culture, data curation and investigated gene expression. All authors have read and agreed to the published version of the manuscript.

Acknowledgements

This work was supported by the China Manned Space Flight Technology Project Chinese Space Station (YYWT-0801-EXP-02), the Strategic Pioneer of CAS (XDB37020104), the National Natural Science Foundation of China (32271277), Shanghai Technology Innovation Action Plan (22JC1401300).The authors are indebted to Prof. Tao Zhang’s group in Shanghai Institute of Technique Physics for culture chamber construction, and Technology and Engineering Center for Space Utilization for supporting the space experiment.

Kiss, J.Z., Brinckmann, E., Brillouet, C.: Development and Growth of Several Strains of Arabidopsis Seedlings in Microgravity. Int. J. Plant. Sci. 161, 55–62 (2000)
Paul, A.L., et al.: Spaceflight Transcriptomes: Unique Responses to a Novel Environment. Astrobiology. 12, 40–56 (2012)
Wang, L., Han, F., Zheng, H.: Photoperiod-controlling Guttation and Growth of rice Seedlings under Microgravity on Board Chinese Spacelab TG-2. Microgravity Sci. Tech. 30, 834–847 (2018)
Matía, I., et al.: Plant Cell Proliferation and Growth Are Altered by Microgravity Conditions in Spaceflight. J. Plant. Physiol. 167, 184–193 (2010)
Hoson, T., et al.: Growth Stimulation in Inflorescences of anArabidopsistubulin Mutant under Microgravity Conditions in Space. J. Plant. Biol. 16, 91–96 (2014)
Tripathy, B.C., Brown, C.S., Levine, H.G., Krikorian, A.D.: Growth and Photosynthetic Responses of Wheat Plants Grown in Space. Plant. Physiol. 110, 801–806 (1996)
Musgrave, M.E., Kuang, A., Tuominen, L.K., Levine, L.H., Morrow, R.C.: Seed storage reserves and glucosinolates in Brassica rapa L. grown on the International Space Station. J. Am. Soc. Hortic. Sci. 130, 848–856 (2005)
Link, B.M., Busse, J.S., Stankovic, B.: Seed-to-seed-to-seed Growth and Development of Arabidopsis in Microgravity. Astrobiology. 14, 866–875 (2014)
Xie, J., Wang, L., Zheng, H.: Molecular Basis to integrate microgravity signals into the photoperiodic flowering pathway in Arabidopsis thaliana under Spaceflight Condition. Int. J. Mol. Sci. 23, 63 (2022)
Zheng, H.: Flowering in space. Microgravity Sci. Technol. 30, 783–791 (2018)
Wang, L., Xie, J., Mou, C., Jiao, Y., Dou, Y., Zheng, H.: Transcriptomic analysis of the interaction between FLOWERING LOCUS T induction and photoperiodic signaling in response to spacefligh. Front. Cell. Dev. Biol. 9, 813246 (2022)
Johnson, C.M., Subramanian, A., Pattathil, S., Correll, M.J., Kiss, J.Z.: Comparative transcriptomics indicate changes in cell wall organization and stress response in seedlings during spaceflight. Am. J. Bot. 104, 1219–1231 (2017)
Halstead, T.W., Dutcher, F.R.: Plants in space. Annu. Rev. Plant. Physiol. 38, 317–345 (1987)
Nechitailo, G.S., Maskinsky, A.L.: Space Biology: Studies at Orbital Station; Mir Publishers: Moscow, Russia, 293–330 (1993)
Kuang, A., Xiao, Y., McClure, G., Musgrave, M.E.: Influence of microgravity on ultrastructure and storage reserves in seeds of Brassica rapa L. Ann. Bot. 85, 851–859 (2000)
Campbell, F., et al.: Comparative floral development of Mir-grown and ethylene-treated, Earth-grown Super Dwarf wheat. J. Plant. Physiol. 158, 1051–1060 (2001)
Link, B.M., Durst, S.J., Zhou, W., Stankovic, B.: Seed-to-seed growth of Arabidopsis thaliana on the international space station. Space Life Sciences: Gravity-Related Processes in Plants. Adv. Space Res. 31, 2237–2243 (2003)
Karahara, I., et al.: Vegetative and reproductive growth of Arabidopsis under microgravity conditions in space. J. Plant. Res. 133, 571–585 (2020)
Andre & Coupland: The genetic basis of flowering responses to seasonal cues. Nat. Rev. Genet. 13, 627–639 (2012)
Fornara, F., et al.: The GI-CDF module of Arabidopsis affects freezing tolerance and growth as well as flowering. Plant. J. 81, 695–706 (2015)
Cao, S., Jiang, S., Zhang, R.: The role of GIGANTEA gene in mediating the oxidative stress response and in Arabidopsis. Plant. Growth Regul. 48, 261–270 (2006)
Qin, F., et al.: SPINDLY, a negative regulator of gibber-ellic acid signaling, is involved in the plant abiotic stress response. Plant. Physiol. 157, 1900–1913 (2011)
Liu, H., et al.: CONSTANS-like 9 (COL9) delays the flowering time in Oryza sativa by repressing the Ehd1 pathway. Biochem. Biophys. Res. Commun. 479, 173–178 (2016)
Du, J., et al.: CONSTANS interacts with and antagonizes ABF transcription factors during salt stress under long-day conditions. Plant. Physiol. 193, 1675–1694 (2023)
Bouché, F., Lobert, G., Tocquin, P., Périlleux, C.: FLOR-ID: An interactive database of flowering- time gene networks in Arabidopsis thaliana. Nucleic. Acids Res. 44, D1167–1171 (2016)
Liu, Y., et al.: CIB1 and CO interact to mediate CRY2-dependent regulation of flowering. EMBO Rep. 19, e45762 (2018)
Song, Y.H., Smith, R.W., To, B.J., Millar, A.: J.& Imaizumi T. FKF1 conveys timing information for CONSTANS stabilization in photoperiodic flowering. Science. 336, 1045–1049 (2012)
Wani, S., Anand, S., Singh, B., Bohra, A., Joshi, R.: WRKY transcription factors and plant defense responses: latest discoveries and future prospects. Plant. Cell. Rep. 40, 1071–1085 (2021)
Jung, J.H., Lee, H.J., Ryu, J.Y., Park, C.M.: SPL3/4/5 integrate developmental aging and photoperiodic signals into the FT-FD module in Arabidopsis flowering. Mol. Plant. 9, 1647–1659 (2016)
Marand, P.A., Chen, Z.L., Gallavotti, A., Schmitz, J.R.: A cis-regulatory atlas in maize at single-cell resolution. Cell. 184, 3041–3055e21 (2021)
Zhang, B., Wang, L., Zeng, L., Zhang, C., Ma, H.: Arabidopsis TOE proteins convey a photoperiodic signal to antagonize CONSTANS and regulate flowering time. Genes Dev. 29, 975–987 (2015)
Jung, J.H., Lee, S., Yun, J., Lee, M., Park, C.M.: The miR172 target TOE3 represses AGAMOUS expression during Arabidopsis floral patterning. Plant. Sci. 215–216, 29–38 (2014)
Gras, D.E., et al.: SMZ/SNZ and gibberellin signaling are required for nitrate-elicited delay of flowering time in Arabidopsis thaliana. J. Exp. Bot. 69, 619–631 (2018)
De Micco, V., De Pascale, S., Paradiso, R., Aronne, G.: Microgravity Effects on Different Stages of Higher Plant Life Cycle and Completion of Theseed-To-Seedcycle. J. Plant. Biol. 16, 31–38 (2014)
Wolff, S.A., Coelho, L.H., Zabrodina, M., Brinckmann, E., Kittang, A.I.: Plant mineral nutrition, gas exchange and photosynthesis in space: A review. Adv. Space Res. 51, 465–475 (2013)
Berrios, D.C., Galazka, J., Grigorev, K., Gebre, S., Costes, S.V.: NASA GeneLab: interfaces for the exploration of space omics data. Nucleic. Acids Res. 49, D1515–1522 (2021)
Wang, S., et al.: Database of space life investigations and information on spaceflight plant biology. Planta. 258, 58 (2023)
Paul, A.L., Wheeler, R.M., Levine, H.G., Ferl, R.J.: Fundamental plant biology enabled by the space shuttle. J. Am. Bot. 100, 226–234 (2013)
Zupanska, A.K., et al.: ARG1 functions in the physiological adaptation of undifferentiated plant cells to spaceflight. Astrobiology. 17, 1077–1111 (2017)
Herranz, R., et al.: RNAseq analysis of the response of Arabidopsis thaliana to fractional gravity under blue-light stimulation during spaceflight. Front. In Plant. Sci. 10, 1529 (2019)
Chan, K., Phua, S.Y., Crisp, P., McQuinn: R.& Pogson B.J. Learning the Languages of the Chloroplast: Retrograde Signaling and Beyond. Annu. Rev. Plant. Biol. 67, 25–53 (2016)
Song, Y., Feng, L., Alyafei, M.A.M., Jaleel, A., Ren, M.: Function of Chloroplasts in Plant Stress Responses. Int. J. Mol. Sci. 22, 13464 (2021)
Hernández-Verdeja, T., Strand, Å.: Retrograde Signals Navigate the Path to Chloroplast Development. Plant. Physiol. 176, 967–976 (2018)
Susila, H., et al.: Chloroplasts prevent precocious flowering through a GOLDEN2-LIKE-B-BOX DOMAIN PROTEIN module. Plant. Commun. 4, 100515 (2023)
Wei, S., et al.: A transcriptional regulator that boosts grain yields and shortens the growth duration of rice. Science. 377, eabi8455 (2022)
Kamachi, H., Tamaoki, D., Karahara, I.: Plasma membrane-anchored chloroplasts are necessary for the gravisensing system of Ceratopteris richardii prothalli. J. Plant. Res. 130, 397–405 (2017)
Vandenbrink, J.P., et al.: RNA-seq analyses of Arabidopsis thaliana seedlings after exposure to blue-light phototropic stimuli in microgravity. Am. J. Bot. 106, 1466–1476 (2019)
Popova, A.F., Musgrave, M., Kuang, A.: The development of embryos in Brassica rapa L. in microgravity. Cytol. Genet. 43, 21–26 (2009)
Kim, W.Y., et al.: ZEITLUPE is a circadian photoreceptor stabilized by GIGANTEA in blue light. Nature. 449, 356–360 (2007)
Yoo, S.K., et al.: CONSTANS activates SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 through FLOWERING LOCUS T to promote flowering in Arabidopsis. Plant. Physiol. 139, 770–778 (2005)
Wu, Y., Xie, J., Wang, L., Zheng, H.: Circumnutations and growth of inflorescence stems of Arabidopsis thaliana in response to microgravity under different photoperiod conditions. Life (Basel). 10, 26 (2020)
Zheng, H., et al.: Live imaging technique for studies of growth and development of Chinese cabbage under microgravity in a recoverable satellite (SJ-8). Microgravity Sci. Technol. 20, 137–143 (2008)

Table 1

Profiles of potential transcript factor (TF) family that are significantly overrepresented in the regulators of spaceflight response genes as detected by the plant regulomics (bioinfo.sibs.ac.cn/plant-n, FDR P_value<0.01). Members of a dedicated gene family are indicated.
TF Family	No.				Family member
TF Family	WT	gi	35S:CO	FG	Family member
ERF	68	69	70	69	139
bZIP	26	27	26	26	127
bHLH	22	25	25	26	225
MYB	14	15	14	15	168
SBP	11	11	11	11	30
WRKY	2	3	10	6	90
TCP	5	9	8	4	33
BES1	5	5	5	5	14
ARF	4	4	4	4	37
G2-like	3	1	4	3	64
CAMTA	3	3	3	3	10
ZF-HD	2	2	3	2	18
Trihelix	2	2	2	2	34
FAR1	2	2	2	2	26
AP2	2	2	2	2	30
NAC	1	3	2	3	138
GATA	1	1	2	1	41
C2H2	1	1	2	1	116
ARR-B	2	1	1	0	21
LFY	1	1	1	1	1
HD-ZIP	1	1	1	1	58
E2F/DP	0	1	1	1	16

Table 2 List of microgravity stress regulated transcription factors with potential significance to the pattern of cis-element enrichment among the target genes in WT, gi, 35S:CO and/or FG.

			Total enrichment score of target motif (%)
Name	Gene_ID	Family	WT	gi	35S:CO	FG
Group I (common microgravity response)
ERF15	AT2G31230	ERF	166	170	172	165
AT4G18450	AT4G18450	ERF	166	168	169	166
ERF104	AT5G61600	ERF	166	169	166	169
ERF2	AT5G47220	ERF	164	169	165	161
ERF5	AT5G47230	ERF	163	175	165	170
ESE3	AT5G25190	ERF	163	174	170	173
PUCHI	AT5G18560	ERF	163	168	168	161
ESE1	AT3G23220	ERF	162	164	166	161
LEP	AT5G13910	ERF	162	173	165	167
AT1G28160	AT1G28160	ERF	162	167	165	161
ERF10	AT1G03800	ERF	160	166	161	158
AT1G75490	AT1G75490	ERF	159	167	158	161
ERF105	AT5G51190	ERF	159	164	165	158
AT2G33710	AT2G33710	ERF	158	168	152	166
ABR1	AT5G64750	ERF	157	170	172	159
AT1G71450	AT1G71450	ERF	157	172	166	169
AT1G44830	AT1G44830	ERF	157	164	152	160
AT4G28140	AT4G28140	ERF	157	161	155	161
RAP2.12	AT1G53910	ERF	157	163	167	157
RAP2.6	AT1G43160	ERF	155	164	166	160
ERF9	AT5G44210	ERF	155	166	161	165
RAP2.11	AT5G19790	ERF	155	157	157	162
AT1G22810	AT1G22810	ERF	154	167	152	164
AT5G67000	AT5G67000	ERF	154	155	160	157
AT5G18450	AT5G18450	ERF	153	164	154	158
AT1G77640	AT1G77640	ERF	150	160	161	164
Group II (GI-CO-FT specific microgravity response)
AREB3	AT3G56850	bZIP		154	154	150
ABF1	AT1G49720	bZIP		153	154	151
bZIP28	AT3G10800	bZIP		156	155	151
ABI5	AT2G36270	bZIP		155	160	162
CEJ1	AT3G50260	ERF		154	150	159
AT1G12630	AT1G12630	ERF		152	156	157
RAP2.1	At1g46768	ERF		151	155	158
DREB26	AT1G21910	ERF		157	153	154
AT4G31060	AT4G31060	ERF		156	155	158
ERF069	AT1G22985	ERF		152	154
AT3G57600	AT3G57600	ERF		153	150
AT4G18890	AT4G18890	BES1			153	153
PIF7	AT5G61270	bHLH			153	151
ABF2	AT1G45249	bZIP			153	156
bZIP68	AT1G32150	bZIP			151	154
AT4G16750	AT4G16750	ERF			153	158
AT1G36060	AT1G36060	ERF			152	152
ERF38	AT2G35700	ERF			151	156
RAP2.10	AT4G36900	ERF		154
ERF4	AT3G15210	ERF		150
PIF1	AT2G20180	bHLH			154
ERF109	AT4G34410	ERF			155
ERF13	AT2G44840	ERF			155
ERF11	AT1G28370	ERF			154
ERF8	AT1G53170	ERF			153
ERF008	AT2G23340	ERF			152
CRF4	AT4G27950	ERF			152
CRF2	AT4G23750	ERF			150
SPL11	AT1G27360	SBP			150
BZR2	AT1G19350	BES1				154
bHLH31	AT1G59640	bHLH				151

There is NO Competing Interest.

SupplementaryTableS1.xlsx
Supplementary Table 1 Differential expression data of all transcripts identified in WT, gi, 35S:CO and FG samples grown in space in comparison with those on ground.
SupplementaryTableS2.xlsx
Supplementary Table 2 Oligonucleotide primers for real-time qPCR in this study.
SupplementaryTableS3.xlsx
Supplementary Table 3 Differential expression data of genes in WT, gi, 35S:CO and FG samples grown in space (sp) in comparison with those on ground (gr). FC≥5, P_adjust<0.01.
SupplementaryTableS4.xlsx
Supplementary Table 4 GO term enrichment of differentially expressed genes in WT, gi, 35S: CO and FG, respectively, in space (sp) compared with those on ground(gr). P_adjust<0.05.
SupplementaryTableS5.xlsx
Supplementary Table 5 Comparison of spaceflight response DEGs identified in gi, 35S:COand FG, respectively, with those DEGs in WT. common-sp: Differential gene expression levels in gi, 35S:CO and FG, respectively, in response to spaceflight was up- or down-regulated less than 1.5-fold in comparison with those in WT. dl-sp: Differential gene expression levels in gi, 35S:COand FG, respectively, in response to spaceflight was up- or down-regulated more than 1.5-fold in comparison with those in WT. +, up-regulation; -, down-regulation.
SupplementaryTableS6.xlsx
Supplementary Table 6 Common and different GO term enrichment of spaceflight-affected genes that exhibited differentially behavior in gi, 35S:CO and FG, respectively, comparing with those in WT. P_adjust<0.05.
SupplementaryTableS7.xlsx
Supplementary Table 7 Differential expression (Log₂FC) of identified flowering time related genes of WT, gi, 35S:CO and FG, respectively, in space in comparison with their controls on ground. FC≥2 and P_adjust≤0.05.
SupplementaryTableS8.xlsx
Supplementary Table 8 Motif enrichment of differential expression genes in response to spaceflight in WT, gi, 35S:CO and FG, respectively. P_adjust<0.01.
SupplFigure.pdf
Supplemental Figures

GI-CO-FT confers microgravity response through Potential Regulatory Modules of Arabidopsis plants grown in Space

Status:

Version 1

Abstract

Figures

Introduction

Results

The response of flowering pathway genes to microgravity in space

The interaction network between microgravity regulated cis-element TFs and regulators of flowering induction

Discussion

Materials and Methods

Plant materials and spaceflight experiments

RNA extraction and sequencing

Real-Time qRT-PCR

Declarations

Competing interests

Author contributions

Acknowledgements

References

Tables

Additional Declarations

Supplementary Files

Status:

Version 1