**Genome-Wide Identification and Expression Profiling of the Polygalacturonase (***PG***) and Pectin Methylesterase (***PME***) Genes in Grapevine (***Vitis vinifera* **L.)**

**Nadeem Khan 1,**† **, Fizza Fatima 2,**† **, Muhammad Salman Haider 3,**† **, Hamna Shazadee <sup>4</sup> , Zhongjie Liu <sup>3</sup> , Ting Zheng <sup>3</sup> and Jinggui Fang 3,\***


Received: 19 May 2019; Accepted: 27 June 2019; Published: 28 June 2019

**Abstract:** In pectin regulation, polygalacturonases (PGs) and pectin methylesterases (PMEs) are critical components in the transformation, disassembly network, and remodeling of plant primary cell walls. In the current study, we identified 36 *PG* and 47 *PME* genes using the available genomic resources of grapevine. Herein, we provide a comprehensive overview of PGs and PMEs, including phylogenetic and collinearity relationships, motif and gene structure compositions, gene duplications, principal component analysis, and expression profiling during developmental stages. Phylogenetic analysis of PGs and PMEs revealed similar domain composition patterns with *Arabidopsis*. The collinearity analysis showed high conservation and gene duplications with purifying selection. The type of duplications also varied in terms of gene numbers in PGs (10 dispersed, 1 proximal, 12 tandem, and 13 segmental, respectively) and PMEs (23 dispersed, 1 proximal, 16 tandem, and 7 segmental, respectively). The tissue-specific response of *PG* and *PME* genes based on the reported transcriptomic data exhibited diverged expression patterns in various organs during different developmental stages. Among PGs, *VvPG8, VvPG10, VvPG13, VvPG17, VvPG18, VvPG19, VvPG20, VvPG22*, and *VvPG23* showed tissue- or organ-specific expression in majority of the tissues during development. Similarly, in PMEs, *VvPME3, VvPME4, VvPME5, VvPME6, VvPME19, VvPME21, VvPME23, VvPME29, VvPME31*, and *VvPME32* suggested high tissue-specific response. The gene ontology (GO), Kyoto Encyclopedia of Genes and Genomics (KEGG) enrichment, and cis-elements prediction analysis also suggested the putative functions of PGs and PMEs in plant development, such as pectin and carbohydrate metabolism, and stress activities. Moreover, qRT-PCR validation of 32 *PG* and *PME* genes revealed their role in various organs of grapevines (i.e., root, stem, tendril, inflorescence, flesh, skins, and leaves). Therefore, these findings will lead to novel insights and encourage cutting-edge research on functional characterization of PGs and PMEs in fruit crop species.

**Keywords:** polygalacturonase (PGs), pectin methylesterase (PMEs), collinearity analysis; gene duplications; expression profiling; grapevine

## **1. Introduction**

Plant cell walls are mainly composed of various interacting networks of carbohydrate polymers, such as polymers of cellulose, hemicellulose, and pectins [1]. These are produced by the plant cells before being systemically delivered to the apoplast tissues [2], providing support and mechanical strength to the plants. However, the disruption of the cell wall structure exposes fruit crops to disorders, such as softening and susceptibility to diseases [3]. Until now, several studies have shown multiple associated roles linked to pectin degradation enzymes in plants. In particular, polygalacturonase (PGs; EC 3.2.1.15) hydrolytic enzymes are involved in plant organ cell separation events, reproductive developments, leaf morphology, and organ shedding [4–7]. Furthermore, expression profiling, both at a temporal and spatial level, has revealed that transcript accumulations are also responsible for cell wall softening during abscission, ripening, and dehiscence [8]. Moreover, these correlation analyses are useful for identifying cell wall sites during disassembly and studying the transcriptional patterns of key genes in cell-wall degrading enzymes [5]. In *Arabidopsis,* during different developmental stages, several members of PGs can be detected by real-time PCR (RT-PCR) in various organs, such as roots, leaves, pollen tubes, flowers, and siliques [9,10]. In addition, few members of PGs have been functionally characterized in some fruit crop species, such as grapevine (*VvPG1* and *VvPG2*), apple (*MdPG36*), banana (*MAPG1* to *MAPG4*), and pear (*PcPGl* and *PcPG3*) [11–14]. However, the silencing of *FaPG1* gene in strawberries declines the halt of the middle lamella and reduces fruit softening [15]. In pears and bananas, the softening of fruits is regulated by *Pc-PG1* and *Pc-PG2* as well as *MaPG3* and *MaPG4*, in an ethylene-dependent pattern [16,17].

Pectin methylesterases (PMEs; EC 3.1.1.11) are also hydrolytic enzymes present in plants, which play a pivotal role in the firmness and softening of the cell walls, specifically through remodeling and disassembly of pectin [18,19]. The PMEs also play a crucial role in plant development, including fruit maturity, pollen development, pollen tube growth, and other growth-related factors [20,21]. However, the activity of both PGs and PMEs during fruit softening is largely unknown and various studies have shown their contradictory actions [22,23]. PGs in plants contain three groups, namely class A, B, and C, whereas the PMEs are categorized into two main types, namely type I and type II [19,24]. PGs and PMEs are a major gene family and have extensively been studied in various crops, including *Brassica rapa, Gossypium* species, *Arabidopsis thaliana*, *Solanum lycopersicum*, *Malus domestica*, *Cucumis sativus*, and *Citrullus lanatus* [5,11,25–28].

Grapevine (*Vitis vinifera* L.) is one of the most important woody fruit crop resource in the world [29]. They have become widely popular due to their nutritional value and health benefits. Improving the grapevine berry quality is a crucial issue, and the availability of the grapevine genome (Version 2.1) provides an excellent opportunity for its genetic study, momentously facilitating research in grapevine biology. In grapevine, the members of PGs and PMEs have not yet been comprehensively identified by genome-wide analysis and their elucidation is largely unknown. In this study, we utilized genomic resources to characterize PGs and PMEs in grapevine by various bioinformatic tools. In total, 36 *PG* and 47 *PME* genes were identified in grapevine and compared with *Arabidopsis* in order to test their domain composition. The expression patterns of genes provide valuable clues for understanding their functions. We also tested 32 *PG* and *PME* genes for various organs of grapevine, and their correlation factors were further compared. Overall, our study provides a brief understanding of the pectin-related genes in grapevine, and their substantial role in regulating organs during different developmental stages.

## **2. Results**
