Investigating the phenotypic plasticity of two red grapevine varieties: an integrated study of transcriptome and environmentome by WGCNA

Grapevine (Vitis spp., family Vitaceae) is characterized by a marked phenotypic plasticity and its ability to withstand various environmental conditions depends on the activation of highly coordinated responses, resulting from the interaction among genotypes (G) and environmental factors (E). Understanding genotype by environment (GxE) interplays is highly complex and challenging, and often the outcome of the genetic responses to field performance is not straightforward also due to the difficulties to reach a precise definition of the E component in field studies. In this work, the transcriptome of commercially ripe berries of Cabernet sauvignon and Aglianico genotypes grown in open-field at three different sites of central-southern Italy (Campania, Molise and Sicily) was analyzed by RNAseq. These transcriptomic data were integrated with a comprehensive set of climatic indices, forming an “environmentome”, through Weighted Gene Co-expression Network Analysis (WGCNA). A total of 11,887 differentially expressed genes (DEGs) have been retrieved, most of them found in the Aglianico genotype, suggesting that Aglianico transcriptome is largely influenced by the environment. Furthermore, the comparisons involving the Sicilian site exhibited the highest number of DEGs for both genotypes. WGCNA suggested that most of the climatic data (such as daily maximum air temperature, relative humidity, air pressure, dew point, hours of sun radiation) significantly correlated with “lightcyan1” module. Among these eingengenes, the low expression of ACA10 cation transporter is suggested to be indirectly related to the low anthocyanin content in Cabernet sauvignon in Campania, thus confirming WGCNA as powerful approach to identify gene of high biological interest. Similarly, transcriptome was also correlated with quality trait, such as total soluble solids and polyphenol content, leading to the discovery of hub genes that might function as markers of wine quality. Overall design: In order to explored the gene-environment (GxE) interactions of two red grape varieties, samples of fully ripe berries of Aglianico and Cabernet Sauvignon varieties grown in open-field conditions at three different sites in southern Italy (Sicily, Molise and Campania) were collected. We then performed RNAseq and gene expression analysis considering three biological replicates for each sample above described Differential expression analysis of each variety in the trhee sites was performed WGCNA was applied to correlate the RNAseq results with several climatic parameters, as well as and with quality traits including total soluble solid, anthocyanin and polyphenol content.

葡萄属（Vitis spp.，葡萄科Vitaceae）具有显著的表型可塑性，其耐受多种环境条件的能力源于基因型（G）与环境因子（E）之间的互作所介导的高度协调的响应激活。解析基因型-环境（G×E）互作机制极具复杂性与挑战性；且由于田间研究中难以精准界定环境组分，遗传响应与田间表现之间的关联往往并非直接明确。
本研究通过RNA测序（RNAseq）分析了在意大利中南部3个不同试验点（坎帕尼亚、莫利塞和西西里）露天种植的赤霞珠（Cabernet sauvignon）与阿利亚尼科（Aglianico）两个基因型葡萄的商业成熟浆果转录组。随后将该转录组数据与一套完整的气候指标数据集整合，构建“环境组（environmentome）”，并通过加权基因共表达网络分析（Weighted Gene Co-expression Network Analysis, WGCNA）开展关联分析。
本研究共检索到11887个差异表达基因（differentially expressed genes, DEGs），其中多数在阿利亚尼科基因型中被检测到，表明阿利亚尼科的转录组受环境影响更为显著。此外，相较于其他试点，西西里试验点的两个基因型均检测到最多的差异表达基因。WGCNA分析显示，多数气候参数（如每日最高气温、相对湿度、气压、露点温度、日照时长）与“lightcyan1”共表达模块显著相关。在该模块的特征基因（eigengenes）中，ACA10阳离子转运蛋白的低表达被认为与坎帕尼亚产区赤霞珠的低花青素含量间接相关，这证实了WGCNA是筛选具有重要生物学意义基因的有效手段。
类似地，本研究还将转录组数据与果实品质性状（如总可溶性固形物、多酚含量）进行关联分析，鉴定出了可作为葡萄酒品质标记的枢纽基因。
总体实验设计：为探究两个红葡萄品种的基因-环境（G×E）互作机制，本研究采集了在意大利南部3个不同试点（西西里、莫利塞、坎帕尼亚）露天种植的阿利亚尼科与赤霞珠两个品种的完全成熟浆果样本。对每个上述样本设置3次生物学重复，随后开展RNA测序与基因表达分析。针对每个品种在3个试点的样本，分别进行差异表达分析，并通过WGCNA将转录组测序结果与多项气候参数，以及总可溶性固形物、花青素、多酚含量等品质性状进行关联分析。