Unveiling the phenology and associated floral regulatory pathways of Humulus lupulus L. in subtropical conditions.

MAIN CONCLUSION: The hop phenological cycle was described in subtropical condition of Brazil showing that flowering can happen at any time of year and this was related to developmental molecular pathways. Hops are traditionally produced in temperate regions, as it was believed that vernalization was necessary for flowering. Nevertheless, recent studies have revealed the potential for hops to flower in tropical and subtropical climates. In this work, we observed that hops in the subtropical climate of Minas Gerais, Brazil grow and flower multiple times throughout the year, independently of the season, contrasting with what happens in temperate regions. This could be due to the photoperiod consistently being inductive, with daylight hours below the described threshold (16.5 h critical). We observed that when the plants reached 7-9 nodes, the leaves began to transition from heart-shaped to trilobed-shaped, which could be indicative of the juvenile to adult transition. This could be related to the fact that the 5th node (in plants with 10 nodes) had the highest expression of miR156, while two miR172s increased in the 20th node (in plants with 25 nodes). Hop flowers appeared later, in the 25th or 28th nodes, and the expression of HlFT3 and HlFT5 was upregulated in plants between 15 and 20 nodes, while the expression of HlTFL3 was upregulated in plants with 20 nodes. These results indicate the role of axillary meristem age in regulating this process and suggest that the florigenic signal should be maintained until the hop plants bloom. In addition, it is possible that the expression of TFL is not sufficient to inhibit flowering in these conditions and promote branching. These findings suggest that the reproductive transition in hop under inductive photoperiodic conditions could occur in plants between 15 and 20 nodes. Our study sheds light on the intricate molecular mechanisms underlying hop floral development, paving the way for potential advancements in hop production on a global scale.

The floral development of the allotetraploid Coffea arabica L. correlates with a small RNA dynamic reprogramming.

Noncoding and coding RNAs are key regulators of plant growth, development, and stress responses. To investigate the types of transcripts accumulated during the vegetative to reproductive transition and floral development in the Coffea arabica L., we sequenced small RNA libraries from eight developmental stages, up to anthesis. We combined these data with messenger RNA and PARE sequencing of two important development stages that marks the transition of an apparent latent to a rapid growth stage. In addition, we took advantage of multiple in silico tools to characterize genomic loci producing small RNAs such as phasiRNAs, miRNAs, and tRFs. Our differential and co-expression analysis showed that some types of small RNAs such as tRNAs, snoRNAs, snRNAs, and phasiRNAs preferentially accumulate in a stage-specific manner. Members of the miR482/miR2118 superfamily and their 21-nucleotide phasiRNAs originating from resistance genes show a robust co-expression pattern that is maintained across all the evaluated developmental stages. Finally, the majority of miRNAs accumulate in a family stage-specific manner, related to modulated hormonal responses and transcription factor expression.

Small RNAs: Promising Molecules to Tackle Climate Change Impacts in Coffee Production.

Over the centuries, human society has evolved based on the ability to select and use more adapted species for food supply, which means making plant species tastier and more productive in particular environmental conditions. However, nowadays, this scenario is highly threatened by climate change, especially by the changes in temperature and greenhouse gasses that directly affect photosynthesis, which highlights the need for strategic studies aiming at crop breeding and guaranteeing food security. This is especially worrying for crops with complex phenology, genomes with low variability, and the ones that support a large production chain, such as Coffea sp. L. In this context, recent advances shed some light on the genome function and transcriptional control, revealing small RNAs (sRNAs) that are responsible for environmental cues and could provide variability through gene expression regulation. Basically, sRNAs are responsive to environmental changes and act on the transcriptional and post-transcriptional gene silencing pathways that regulate gene expression and, consequently, biological processes. Here, we first discuss the predicted impact of climate changes on coffee plants and coffee chain production and then the role of sRNAs in response to environmental changes, especially temperature, in different species, together with their potential as tools for genetic improvement. Very few studies in coffee explored the relationship between sRNAs and environmental cues; thus, this review contributes to understanding coffee development in the face of climate change and towards new strategies of crop breeding.

Metabolic Pathway Reconstruction Indicates the Presence of Important Medicinal Compounds in Coffea Such as L-DOPA.

The use of transcriptomic data to make inferences about plant metabolomes is a useful tool to help the discovery of important compounds in the available biodiversity. To unveil previously undiscovered metabolites of Coffea, of phytotherapeutic and economic value, we employed 24 RNAseq libraries. These libraries were sequenced from leaves exposed to a diverse range of environmental conditions. Subsequently, the data were meticulously processed to create models of putative metabolic networks, which shed light on the production of potential natural compounds of significant interest. Then, we selected one of the predicted compounds, the L-3,4-dihydroxyphenylalanine (L-DOPA), to be analyzed by LC-MS/MS using three biological replicates of flowers, leaves, and fruits from Coffea arabica and Coffea canephora. We were able to identify metabolic pathways responsible for producing several compounds of economic importance. One of the identified pathways involved in isoquinoline alkaloid biosynthesis was found to be active and producing L-DOPA, which is a common product of POLYPHENOL OXIDASES (PPOs, EC 1.14.18.1 and EC 1.10.3.1). We show that coffee plants are a natural source of L-DOPA, a widely used medicine for treatment of the human neurodegenerative condition called Parkinson's disease. In addition, dozens of other compounds with medicinal significance were predicted as potential natural coffee products. By further refining analytical chemistry techniques, it will be possible to enhance the characterization of coffee metabolites, enabling a deeper understanding of their properties and potential applications in medicine.

Expression of coffee florigen CaFT1 reveals a sustained floral induction window associated with asynchronous flowering in tropical perennials.

The behavior of florigen(s) and environment-influenced regulatory pathways that control floral initiation in tropical perennials species with complex phenological cycles is poorly understood. Understanding the mechanisms underlying this process is important for food production in the face of climate change, thus, we used Coffea sp. L. (Rubiaceae) as a model to explore this issue. Homologs of FLOWERING LOCUS T (CaFT1) and environment-related regulators CONSTANS (CaCO), PHYTOCHROME INTERACTING FACTOR 4 (CaPIF4) and FLOWERING LOCUS C (CaFLC) were retrieved from coffee genomes and identified through phylogenetic analysis. Overexpression of CaFT1 in Arabidopsis caused early-flowering phenotype and yeast two hybrid studies indicated CaFT1 binding to bZIP floral regulator FD, which suggests that CaFT1 is a coffee florigen. Expression of CaFT1 and other floral regulators, together with carbohydrate analysis, were evaluated over one year using three contrasting genotypes, two C. arabica cultivars and C. canephora. All genotypes showed active and variable CaFT1 transcription from February until October, indicating the potential window for floral induction that reached a maximum in the cold period of June. CaCO expression, as expected, varied over a 24-hour day period and monthly with day length, whereas expression of temperature-responsive homologs, CaFLC and CaPIF4, did not correlate with temperature changes nor CaFT1 expression, suggesting alternative FT regulatory pathways in coffee. Based on our results, we suggest a continuum of floral induction that allows different starting points for floral activation, which explains developmental asynchronicity and prolonged anthesis events in tropical perennial species.

Genome-Wide Analyses of MADS-Box Genes in Humulus lupulus L. Reveal Potential Participation in Plant Development, Floral Architecture, and Lupulin Gland Metabolism.

MADS-box transcription factors (TFs) are involved in multiple plant development processes and are most known during the reproductive transition and floral organ development. Very few genes have been characterized in the genome of Humulus lupulus L. (Cannabaceae), an important crop for the pharmaceutical and beverage industries. The MADS-box family has not been studied in this species yet. We identified 65 MADS-box genes in the hop genome, of which 29 encode type-II TFs (27 of subgroup MIKCC and 2 MIKC*) and 36 type-I proteins (26 α, 9 ß, and 1 γ). Type-II MADS-box genes evolved more complex architectures than type-I genes. Interestingly, we did not find FLOWERING LOCUS C (FLC) homologs, a transcription factor that acts as a floral repressor and is negatively regulated by cold. This result provides a molecular explanation for a previous work showing that vernalization is not a requirement for hop flowering, which has implications for its cultivation in the tropics. Analysis of gene ontology and expression profiling revealed genes potentially involved in the development of male and female floral structures based on the differential expression of ABC homeotic genes in each whorl of the flower. We identified a gene exclusively expressed in lupulin glands, suggesting a role in specialized metabolism in these structures. In toto, this work contributes to understanding the evolutionary history of MADS-box genes in hop, and provides perspectives on functional genetic studies, biotechnology, and crop breeding.

Dose-response effect of prebiotic ingestion (ß-glucans isolated from Saccharomyces cerevisiae) in diabetic rats with periodontal disease.

BACKGROUND: Periodontal disease is one of the most frequent comorbidities in diabetic patients and can contribute to poor blood glucose control. OBJECTIVE: To evaluate the effects of ingesting different doses of beta-glucans (BG) isolated from Saccharomyces cerevisiae on alveolar bone loss (ABL) and inflammatory/metabolic parameters in normal and diabetic rats with ligature-induced periodontal disease (PD). DESIGN: Sixty male rats were assigned into two groups: non-diabetic or diabetic (i.p. 70 mg/kg streptozotocin) with PD. Then, groups were subdivided into five subgroups according BG doses: 0 mg/Kg; 10 mg/Kg; 20 mg/Kg; 40 mg/Kg or 80 mg/Kg. Animals received BG for 28 days and ligatures were placed on lower first molars during the last 14 days. RESULTS: ABL of diabetic and non-diabetic animals receiving BG 40 mg/kg (1.33 ± 0.03 mm and 0.77 ± 0.07 mm, respectively) and 80 mg/kg (1.26 ± 0.07 mm and 0.78 ± 0.05 mm, respectively) doses was lower (p < 0.05) in comparison to respective controls (1.59 ± 0.11 mm and 0.90 mm ±0.08). COX-2 (Control: 1.66 ± 0.12; 40 mg/kg: 1.13 ± 0.07; 80 mg/kg: 0.92 ± 0.18) and RANKL expressions (Control: 1.74 ± 0.34; 40 mg/kg: 1.03 ± 0.29 ;80 mg/kg: 0.75 ± 0.21), together with the RANKL/OPG ratio (Control: 1.17 ± 0.08; 40 mg/kg: 0.67 ± 0.09; 80 mg/kg: 0.63 ± 0.28) were attenuated above the same dose (p < 0.05). BG did not influence (p > 0.05) metabolic parameters in non-diabetic rats. In diabetic animals, doses above 40 mg/kg reduced IL-1ß (Control: 387 ± 66; 40 mg/kg: 309 ± 27; 80 mg/kg: 300 ± 14) and TNF-α (Control: 229 ± 19; 40 mg/kg: 128 ± 53; 80 mg/kg: 71 ± 25), blood glucose levels (Control: 402 ± 49; 40 mg/kg: 334 ± 32; 80 mg/kg: 287 ± 56), total cholesterol (Control: 124 ± 8; 40 mg/kg: 120 ± 10; 80 mg/kg: 108 ± 9), LDL-c + VLDL-c (Control: 106 ± 8; 40 mg/kg: 103 ± 10; 80 mg/kg: 87 ± 10) and triacylglycerols (Control: 508 ± 90; 40 mg/kg: 301 ± 40; 80 mg/kg: 208 ± 61), whereas increased HDL-c (Control: 18 ± 0.5; 40 mg/kg: 19 ± 1; 80 mg/kg: 21 ± 1) (p < 0.05). Optimal dose needed to reduce ABL was higher in diabetic animals with PD. CONCLUSIONS: BG ingestion reduced ABL and improved inflammatory profile in a dose-dependent manner. Best effects were achieved with doses above 40 mg/kg.

Elevated Temperatures Impose Transcriptional Constraints and Elicit Intraspecific Differences Between Coffee Genotypes.

The projected impact of global warming on coffee production may require the heat-adapted genotypes in the next decades. To identify cellular strategies in response to warmer temperatures, we compared the effect of elevated temperature on two commercial Coffea arabica L. genotypes exploring leaf physiology, transcriptome, and carbohydrate/protein composition. Growth temperatures were 23/19°C (day/night), as optimal condition (OpT), and 30/26°C (day/night) as a possible warmer scenario (WaT). The cv. Acauã showed lower levels of leaf temperature (Tleaf) under both conditions compared to cv. Catuaí, whereas slightly or no differences for other leaf physiological parameters. Therefore, to explore temperature responsive pathways the leaf transcriptome was examined using RNAseq. Genotypes showed a marked number of differentially-expressed genes (DEGs) under OpT, however DEGs strongly decrease in both at WaT condition indicating a transcriptional constraint. DEGs responsive to WaT revealed shared and genotype-specific genes mostly related to carbohydrate metabolism. Under OpT, leaf starch content was greater in cv. Acauã and, as WaT temperature was imposed, the leaf soluble sugar did not change in contrast to cv. Catuaí, although the levels of leaf starch, sucrose, and leaf protein decreased in both genotypes. These findings revealed intraspecific differences in the underlying transcriptional and metabolic interconnected pathways responsive to warmer temperatures, which is potentially linked to thermotolerance, and thus may be useful as biomarkers in breeding for a changing climate.

Flower development in Coffea arabica L.: new insights into MADS-box genes.

Coffea arabica L. shows peculiar characteristics during reproductive development, such as flowering asynchrony, periods of floral bud dormancy, mucilage secretion and epipetalous stamens. The MADS-box transcription factors are known to control several developmental processes in plants, including flower and fruit development. Significant differences are found among plant species regarding reproductive development and little is known about the role of MADS-box genes in Coffea reproductive development. Thus, we used anatomical and comparative molecular analyses to explore the flowering process in coffee. The main morphological changes during flower development in coffee were observed by optical and scanning electron microscopy. Flowering asynchrony seems to be related to two independent processes: the asynchronous development of distinct buds before the reproductive induction and the asynchronous development of floral meristems within each bud after the reproductive induction. A total of 23 C. arabica MADS-box genes were characterized by sequence comparison with putative Arabidopsis orthologs and their expression profiles were analyzed by RT-PCR in different tissues. The expression of the ABC model orthologs in Coffea during floral development was determined by in situ hybridization. The APETALA1 (AP1) ortholog is expressed only late in the perianth, which is also observed for the APETALA3 and TM6 orthologs. Conversely, the PISTILLATA ortholog is widely expressed in early stages, but restrict to stamens and carpels in later stages of flower development, while the expression of the AGAMOUS ortholog is always restricted to fertile organs. The AP1 and PISTILLATA orthologs are also expressed at specific floral organs, such as bracts and colleters, respectively, suggesting a potential role in the development of such structures. Altogether, the results from our comprehensive expression analyses showed significant differences between the spatiotemporal expression profiles of C. arabica MADS-box genes and their orthologs, which suggests differential functionalization in coffee. Moreover, these differences might also partially explain the particular characteristics of floral development in coffee, such as mucilage secretion and formation of epipetalous stamens.