A cost-effective dual reporter system in Nicotiana benthamiana

Frontiers in Plant Science · 2026-02-03

Dual reporter assay has been widely used for quantification of gene expression in various model systems. However, the assay is dependent on commercial kits, which may be inhibitory for many low-budget academic laboratories or high throughput screening. Here, we systematically characterized seven commonly used or potential reporters, and identified NanoLuc-GFP pair as the optimal kit-independent and accurate dual reporter in Nicotiana benthamiana . The NanoLuc-GFP system exhibited high stability, which can largely reduce the interference caused by biological variability and dramatic environmental fluctuations. Furthermore, it exhibits greater sensitivity compared to the conventional FLuc/RLuc system commercially available.

A New UV‐B Protectant in Plant: Insights Into THCAS and THCA's Role in UV‐B Tolerance

Plant Cell & Environment · 2025-07-14 · 2 citations

Understanding how plants respond to environmental stressors such as ultraviolet B (UV-B) radiation is critical for agricultural resilience and crop improvement. Investigation of the UV-B response in 17 cannabis accessions revealed distinct differences in morphological changes between two groups, A and B. Whole-genome resequencing and population-level analyses indicate that Δ9-tetrahydrocannabinolic acid synthase (THCAS) is subject to positive selection or a selective sweep. Sequence analysis and functional characterization of THCAS revealed that two single nucleotide variations, resulting in two amino acids change at Pro333 and Ser355, which are associated with THCAS activity and potentially linked to differential plant response to UV-B radiation. The role of THCAS in enhancing UV-B tolerance was further demonstrated by overexpressing functional THCAS in different plant species under UV-B stress environment. This study suggests that THCA, produced by the enzyme THCAS, acts as a shield against UV-B radiation. Plants with higher levels of THCA exhibited greater tolerance to UV-B exposure. Exogenous application of THCA alleviated UV-B damage in various plant species. Our findings highlight the importance of THCAS-mediated THCA synthesis in conferring UV-B tolerance and provide insights into the genomic and molecular mechanisms that govern plant adaptation to UV-B radiation.

HyperTRIBE mapping of the RNA m6A demethylase ALKBH9 binding sites in bamboo reveals its role in plant defense

PLANT PHYSIOLOGY · 2025-09-29 · 2 citations

RNA demethylation plays an important role in diverse biological processes. Intriguingly, RNA demethylation has not been reported in bamboo, which is known for its rapid growth. PheALKBH9, an m6A demethylase in bamboo, was stably transformed into rice and increased its susceptibility to rice blast disease. Heterologous expression of PheALKBH9 reduced the overall m6A modification levels in rice. Using HyperTRIBE (Targets of RNA-binding proteins Identified By Editing), we identified evolutionarily conserved PheALKBH9 target RNAs in both rice and Moso bamboo. Overexpression of PheALKBH9 led to higher protein expression and shorter poly(A) tails. Notably, PheALKBH9 directly bound to CCR4-associated factor1 (CAF1G) and poly(A)-binding genes (PABPC1 and PABPC2), potentially modulating poly(A) tail lengths. In addition, PheALKBH9 also bound to and removed m6A modifications from Perox4, JAZ7, and METS2, key players in plant immunity, suggesting that PheALKBH9 plays a role in plant disease resistance. In summary, our study unveils a previously unknown role of PheALKBH9-mediated m6A demethylation in response to blast disease and provides insights into its mechanisms in monocotyledonous plants.

Advances in bamboo genomics: Growth and development, stress tolerance, and genetic engineering

Journal of Integrative Plant Biology · 2025-05-02 · 20 citations

Bamboo is a fast-growing and ecologically significant plant with immense economic value due to its applications in construction, textiles, and bioenergy. However, research on bamboo has been hindered by its long vegetative period, unpredictable flowering cycles, and challenges in genetic transformation. Recent developments in advanced sequencing and genetic engineering technologies have provided new insights into bamboo's evolutionary history, developmental biology, and stress resilience, paving the way for improved conservation and sustainable utilization. This review synthesizes the latest findings on bamboo's genomics, biotechnology, and the molecular mechanisms governing its growth, development, and stress response. Key genes and regulatory pathways controlling its rapid growth, internode elongation, rhizome development, culm lignification, flowering, and abiotic stress responses have been identified through multi-omics and functional studies. Complex interactions among transcription factors, epigenetic regulators, and functionally important genes shape bamboo's unique growth characteristics. Moreover, progress in genetic engineering techniques, including clustered regularly interspaced short palindromic repeats-based genome editing, has opened new avenues for targeted genetic improvements. However, technical challenges, particularly the complexity of polyploid bamboo genomes and inefficient regeneration systems, remain significant barriers to functional studies and large-scale breeding efforts. By integrating recent genomic discoveries with advancements in biotechnology, this review proposes potential strategies to overcome existing technological limitations and to accelerate the development of improved bamboo varieties. Continued efforts in multi-omics research, gene-editing applications, and sustainable cultivation practices will be essential for harnessing bamboo as a resilient and renewable resource for the future. The review presented here not only deepens our understanding of bamboo's genetic architecture but also provides a foundation for future research aimed at optimizing its ecological and industrial potential.

Plant physiology: Rethinking CRY photoreceptors

Current Biology · 2025-01-01 · 3 citations

The two action mechanisms of plant cryptochromes

Trends in Plant Science · 2025-01-28 · 21 citations

Plant cryptochromes (CRYs) are photolyase-like blue-light receptors that contain a flavin adenine dinucleotide (FAD) chromophore. In plants grown in darkness, CRYs are present as monomers. Photoexcited CRYs oligomerize to form homo-tetramers. CRYs physically interact with non-constitutive or constitutive CRY-interacting proteins to form the non-constitutive or constitutive CRY complexes, respectively. The non-constitutive CRY complexes exhibit a different affinity for CRYs in response to light, and act by a light-induced fit (lock-and-key) mechanism. The constitutive CRY complexes have a similar affinity for CRYs regardless of light, and act via a light-induced liquid-liquid phase separation (LLPS) mechanism. These CRY complexes mediate blue-light regulation of transcription, mRNA methylation, mRNA splicing, protein modification, and proteolysis to modulate plant growth and development.

Light signaling in plants—a selective history

PLANT PHYSIOLOGY · 2024-03-02 · 48 citations

In addition to providing the radiant energy that drives photosynthesis, sunlight carries signals that enable plants to grow, develop and adapt optimally to the prevailing environment. Here we trace the path of research that has led to our current understanding of the cellular and molecular mechanisms underlying the plant's capacity to perceive and transduce these signals into appropriate growth and developmental responses. Because a fully comprehensive review was not possible, we have restricted our coverage to the phytochrome and cryptochrome classes of photosensory receptors, while recognizing that the phototropin and UV classes also contribute importantly to the full scope of light-signal monitoring by the plant.

The eukaryotic translation initiation factor <scp>eIF4E</scp> regulates flowering and circadian rhythm in Arabidopsis

The Plant Journal · 2024-08-15 · 1 citations

Translation initiation is a critical, rate-limiting step in protein synthesis. The eukaryotic translation initiation factor 4E (eIF4E) plays an essential role in this process. However, the mechanisms by which eIF4E-dependent translation initiation regulates plant growth and development remain not fully understood. In this study, we found that Arabidopsis eIF4E proteins are distributed in both the nucleus and cytoplasm, with only the cytoplasmic eIF4E being involved in the control of photoperiodic flowering. Genome-wide translation profiling using Ribo-tag sequencing reveals that eIF4E may regulate plant flowering by maintaining the homeostatic translation of components in the photoperiodic flowering pathway. eIF4E not only regulates the translation of flowering genes such as FLOWERING LOCUS T (FT) and FLOWERING LOCUS D (FLD) but also influences the translation of circadian genes like CIRCADIAN CLOCK ASSOCIATED 1 (CCA1) and PSEUDO-RESPONSE REGULATOR 9 (PRR9). Consistently, our results show that the eIF4E modulates the rhythmic oscillation of the circadian clock. Together, our study provides mechanistic insights into how the protein translation regulates multiple developmental processes in Arabidopsis, including the circadian clock and photoperiodic flowering.

The dark activity of Arabidopsis blue-light receptor CRY2

Science China Life Sciences · 2024-11-29 · 1 citations

Co-condensation with photoexcited cryptochromes facilitates MAC3A to positively control hypocotyl growth in <i>Arabidopsis</i>

Science Advances · 2023-08-09 · 22 citations

Cryptochromes (CRYs) are blue light receptors that mediate plant photoresponses through regulating gene expressions. We recently reported that Arabidopsis CRY2 could form light-elicited liquid condensates to control RNA methylation. However, whether CRY2 condensation is involved in other gene expression–regulatory processes remains unclear. Here, we show that MOS4-associated complex subunits 3A and 3B (MAC3A/3B) are CRY-interacting proteins and assembled into nuclear CRY condensates. mac3a3b double mutants exhibit hypersensitive photoinhibition of hypocotyl elongation, suggesting that MAC3A/3B positively control hypocotyl growth. We demonstrate the noncanonical activity of MAC3A as a DNA binding protein that modulates transcription. Genome-wide mapping of MAC3A-binding sites reveals that blue light enhances the association of MAC3A with its DNA targets, which requires CRYs. Further evidence indicates that MAC3A and ELONGATED HYPOCOTYL 5 (HY5) occupy overlapping genomic regions and compete for the same targets. These results argue that photocondensation of CRYs fine-tunes light-responsive hypocotyl growth by balancing the opposed effects of HY5 and MAC3A.