Naoki Tani

Circadian rhythms allow plants to regulate internal processes to align with daily rhythms in the environment. Comparing circadian clocks across environments is essential because latitude-dependent variation in light and temperature imposes distinct selection pressures that shape the evolution and function of circadian timing systems. However, c ircadian clock studies have largely focused on temperate and subtropical species, leaving transcriptional circadian networks in tropical plants under relatively constant environments poorly understood. We report the first comprehensive circadian transcriptome of the ecologically important dipterocarp species Rubroshorea leprosula , a dominant species in tropical rainforests in Southeast Asia. One sapling was sampled every four hours for 48 hours under constant darkness and used for transcriptomic analyses. Only 283 of 20,748 expressed genes (∼1.3%) exhibited significant circadian oscillations, with periods strongly concentrated between 23 and 25 h. Hierarchical clustering revealed four temporal clusters with alternate phases of expression and functional specialisation; morning clusters in processes related to light and chloroplast, midday clusters in hormone and signalling mechanisms, afternoon clusters in mitochondrial and peptide biosynthesis functions, and night clusters in protein quality control and autophagy. Comparative analysis identified clear orthologs for all major Arabidopsis circadian clock components ( LHY, CCA1, PRRs, TOC1, ELF4, LUX, GI, ZTL, RVE s), with conserved synteny to Parashorea chinensis , a close relative of R. leprosula . Time-lagged cross-correlation (TLCC) network reconstruction identified a characteristic circadian topological similarity with Arabidopsis, including coupled morning and evening feedback loops and paralog expansion that maintained overall structure. Peak expression timing of these core clock genes in the tropical tree was largely consistent with that observed in Arabidopsis thaliana . In contrast to this conserved phase relationship, Rubroshorea orthologs exhibited reduced amplitudes and lower coefficients of variation in their circadian oscillations, suggesting diminished robustness of rhythmic gene expression. These findings demonstrate a conserved but regulated circadian mechanism in R. leprosula , in preparation for adaptation to tropical rainforest’s stable light and temperature regimes. This study lays the molecular foundation for circadian regulation in dipterocarps and offers a system for integrating rhythmic gene expression to ecological function and forest productivity in tropical communities.

Latitudinal gradients in temperature seasonality shape the evolution of thermal tolerance and acclimation capacity in organisms. According to Janzen's climate variability hypothesis, tropical species experiencing stable temperatures evolve narrower thermal niches than temperate counterparts. To test whether this principle extends to gene expression dynamics, we compared annual transcriptome profiles of the tropical tree Rubroshorea leprosula and the temperate evergreen trees Lithocarpus edulis and Quercus glauca under natural field conditions. Time‐series RNA‐seq analyses revealed that R. leprosula exhibited sporadic transcriptional shifts triggered by slight cooling events (minimum temperatures of 21°C–22°C), whereas the temperate species showed clear annual cycles characterized by winter‐specific expression patterns. Mild temperature decline in the tropical tree triggered widespread down‐regulation of photosynthesis‐related genes and activation of stress‐response and jasmonate‐associated signaling pathways, suggesting coordinated responses even to mild temperature declines. Cross‐species comparison of 3793 single‐copy orthologs showed that the sensitivity to temperature and dynamic range of gene expression were substantially larger relative to the narrow dynamic range of temperature in the tropics, indicating amplified transcriptional responses per unit of temperature variation. Conversely, temperate species displayed broad but proportionate transcriptomic shifts that paralleled large seasonal temperature fluctuations. These results demonstrate that transcriptional sensitivity to temperature is exaggerated in tropical species and buffered in temperate ones, extending Janzen's climate variability hypothesis from physiological tolerance to its molecular basis.

Introduction: The role of syngameons in adaption to microgeographical environmental heterogeneity is important and could be one of the sources of rich species diversity in tropical forests. In addition, negative frequency- or density-dependent selection is one of the major processes contributing to the maintenance of genetic diversity.Methods: To assess genetic factors that affect the fitness of seedlings of Rubroshorea curtisii, a dominant canopy tree species in hill dipterocarp forests, the inter- and intra-population genetic structure of individuals from natural populations and individuals at two permanent plots in a hill dipterocarp forest with reproductive stage was studied. Further, a total of 460 seedlings derived from six mother trees in the plot were raised in a nursery, and their pollen donors were identified using genetic marker based paternity assignment. Seed weight, bi-parental genetic relatedness, and bi-parental genetic heterogeneity based on the clustering analysis were used to analyze their effects on seedling fitness.Results: A Bayesian based clustering analysis revealed that three genetically distinct clusters were observed in almost all populations throughout the distributional range of the species in Malay Peninsula and provided the optimum explanation for the genetic structure of 182 mature individuals in the plots. The two clusters showed larger genetic differentiation from the ancestral admixture population, but the other one was not differentiated. The bi-parental larger genetic heterogeneity was associated with a significantly higher probability of seedling survivorship, and likewise, higher performance of vertical growth of the seedlings; but the seed weight and genetic relatedness did not significantly affect those.Discussion: This evidence suggests that fitter seedlings derived from mating between parents with different genetic clusters contribute to maintaining genetic diversity through negative frequency-dependent selection and may have an important role in adaptation in the tropical forest plant community.

Climatic factors have commonly been attributed as the trigger of general flowering, a unique community-level mass flowering phenomenon involving most dipterocarp species that forms the foundation of Southeast Asian tropical rainforests. This intriguing flowering event is often succeeded by mast fruiting, which provides a temporary yet substantial burst of food resources for animals, particularly frugivores. However, the physiological mechanism that triggers general flowering, particularly in dipterocarp species, is not well understood largely due to its irregular and unpredictable occurrences in the tall and dense forests. To shed light on this mechanism, we employed ecological transcriptomic analyses on an RNA-seq dataset of a general flowering species,Shorea curtisii(Dipterocarpaceae), sequenced from leaves and buds collected at multiple vegetative and flowering phenological stages. We assembled 64,219 unigenes from the transcriptome of which 1,730 and 3,559 were differentially expressed in the leaf and the bud, respectively. Differentially expressed unigene clusters were found to be enriched with homologs ofArabidopsis thalianagenes associated with response to biotic and abiotic stresses, nutrient level, and hormonal treatments. When combined with rainfall data, our transcriptome data reveals that the trees were responding to a brief period of drought prior to the elevated expression of key floral promoters and followed by differential expression of unigenes that indicates physiological changes associated with the transition from vegetative to reproductive stages. Our study is timely for a representative general flowering dipterocarp species that occurs in forests that are under the constant threat of deforestation and climate change as it pinpoints important climate sensitive and flowering-related homologs and offers a glimpse into the cascade of gene expression before and after the onset of floral initiation.