Transcriptome Analyses of Leaves Reveal That Hexanoic Acid Priming Differentially Regulate Gene Expression in Contrasting Coffea arabica Cultivars

Citation: Budzinski IGF, Camargo PO, Rosa RS, Calzado NF, Ivamoto-Suzuki ST and Domingues DS (2021) Transcriptome Analyses of Leaves Reveal That Hexanoic Acid Priming Differentially Regulate Gene Expression in Contrasting Coffea arabica Cultivars. Front. Sustain. Food Syst. 5:735893. doi: 10.3389/fsufs.2021.735893 Transcriptome Analyses of Leaves Reveal That Hexanoic Acid Priming Differentially Regulate Gene Expression in Contrasting Coffea arabica Cultivars


INTRODUCTION
Coffee (Coffea spp.) is one of the most important traded commodities in the international market [International Coffee Organization (ICO), 2018]. C. arabica, the only allotetraploid species in the genus, is the most planted one. Despite its economic importance, evaluation of physiological changes taking account molecular responses to biostimulants are still scarce. When plants recognize potential biotic/abiotic challenges, they often switch to a primed state of enhanced defense. This mechanism enables plants to respond robustly after exposure to stress (Aranega-Bou et al., 2014). The response of C. arabica plants to priming, in terms of transcriptional profiles, is a big gap in this area. Hexanoic acid (Hx) is a natural priming agent with proven efficiency in a wide range of host plants and pathogens (Llorens et al., 2016), including coffee pathogens. In this study we aimed to investigate the effect of Hx priming in C. arabica leaves transcriptome. We hypothesize if Hx application could modulate genes related to defense responses, being a potential eliciting agent in C. arabica. To test this effect, we applied Hx in roots of two Brazilian C. arabica cultivars with distinct breeding histories and contrasting resistance to the major disease in Arabica coffee, coffee leaf rust. While Catuai Vermelho is among the most used cultivars in Brazil, but it is susceptible to leaf rust, Obatã is a moderately resistant cultivar (Del Grossi et al., 2013). We performed transcriptome analysis of leaves. Reads were mapped to the C. arabica public genome and up to 94% of reads were mapped. Transcript expression level was quantified and differentially expressed genes (DEGs) were identified based on FPKM ratio and statistical analyses. A total of 57 and 63 DEGs were found in Catuai Vermelho and Obatã, respectively. Most DEGs correspond to upregulated genes in response to Hx, in both cultivars (86% Catuai Vermelho and 73% Obatã). Eight DEGs were found modulated in both cultivars, including ferredoxin-NADP reductase and phenylalanine ammonia-lyase. DEGs were functionally annotated through Blast2GO. Biological process and molecular function categorizations revealed that DEGs related to cellular, oxidationreduction, organic substance and primary metabolic process, as well as transferase and ion binding activity might play a dominant role in the leaf response to priming. These data contribute to identify key genes differentially expressed in response to Hx as well as indicate pathways modulated by this eliciting agent.

VALUE OF DATA
• The species C. arabica is responsible for 60% of the world coffee production due to its fine flavor and aroma. Brazil is the largest producer and exporter of this commodity. • Hexanoic acid (Hx) is a priming agent which leads plant to a physiological state that enables them to respond more rapidly and/or robustly to biotic or abiotic stress scenario. • To date, there is no transcriptome analysis for C. arabica cultivars Catuai Vermlho and Obatã in response to the priming agent hexanoic acid (Hx) application. • These data will contribute to identify key genes differentially expressed in response to Hx application and it can also indicates which pathways are modulated by Hx.

Plant Material
Five-month-old plants of C. arabica cv. Catuaí Vermelho IAC 144 and Obatã IAC 1669-20 (4-5 leaf pairs) were used in this study. Plants were selected based on size uniformity and were transferred to pots containing 3 L of aerated nutrient solution (ANS), adapted from Clark (1975) by de Carvalho et al. (2013). The hexanoic acid experiment was carried out as described in Silva et al. (2020), under controlled temperature (23 ± 2 • C) and light/dark cycle (12/12 h, photosynthetically active photon flux density of ∼400 µmol.m −2 .s −1 ). The following treatments were assessed: (a) ANS (control); (b) ANS + hexanoic acid (Merck, final concentration 0.55 mM) for 48 h. Plants were grown in 3 to 6 plastic pots in which three pots received each treatment. The experiments were repeated 3 times to obtain biological replicates. The potted plants were grouped in "pools" (made of 9-18 plants), which were considered a biological replicate. Three biological replicates were used. The mature leaves of the middle third of the plants were collected within the 3rd hour of the light period and stored at −80 • C to further analyses.
Total RNA Extraction and Quality Control, Library Preparation, and RNA-Seq

DEGs Data Annotation and Gene Ontology (GO) Analysis
Sequence annotation and functional analysis were done for the DEGs by using Blast2GO (Conesa et al., 2005), at the OmicsBox (Götz et al., 2008) platform. Sequences were annotated by blasting nucleotide sequences against the NCBI NR database (BLASTX, evalue ≤ 1.10 −5 ). A total of 57 and 63 DEGs were found in Catuai Vermelho and Obatã, respectively. Most DEGs correspond to upregulated genes in response to Hx, in both cultivars (86% Catuai Vermelho and 73% Obatã). Eight DEGs were commonly found in the Catuai Vermelho and Obatã cultivars (AAA-ATPase, auxininduced protein (two transcripts), ferredoxin-NADP reductase, premnaspirodiene oxygenase-like, glutathionyl-hydroquinone reductase, flavonol 3-O-glucosyltransferase and phenylalanine ammonia-lyase). DEGs were analyzed by Gene Ontology and were functionally assigned to the relevant terms. Biological process and molecular function categorizations revealed that DEGs related to cellular, oxidation-reduction, organic substance, and primary metabolic process, as well as transferase and ion binding activity might play a dominant role in the leaf response to priming. Most of the DEGs have a role in plant defense, corroborating to our hypothesis. These data contribute to identify key genes differentially expressed in response to Hx as well as indicate pathways modulated by this eliciting agent.

DATA AVAILABILITY STATEMENT
The datasets presented in this study can be found in online repositories.