The Cell Wall PAC (Proline-Rich, Arabinogalactan Proteins, Conserved Cysteines) Domain-Proteins Are Conserved in the Green Lineage.

Plant cell wall proteins play major roles during plant development and in response to environmental cues. A bioinformatic search for functional domains has allowed identifying the PAC domain (Proline-rich, Arabinogalactan proteins, conserved Cysteines) in several proteins (PDPs) identified in cell wall proteomes. This domain is assumed to interact with pectic polysaccharides and O-glycans and to contribute to non-covalent molecular scaffolds facilitating the remodeling of polysaccharidic networks during rapid cell expansion. In this work, the characteristics of the PAC domain are described in detail, including six conserved Cys residues, their spacing, and the predicted secondary structures. Modeling has been performed based on the crystal structure of a Plantago lanceolata PAC domain. The presence of ß-sheets is assumed to ensure the correct folding of the PAC domain as a ß-barrel with loop regions. We show that PDPs are present in early divergent organisms from the green lineage and in all land plants. PAC domains are associated with other types of domains: Histidine-rich, extensin, Proline-rich, or yet uncharacterized. The earliest divergent organisms having PDPs are Bryophytes. Like the complexity of the cell walls, the number and complexity of PDPs steadily increase during the evolution of the green lineage. The association of PAC domains with other domains suggests a neo-functionalization and different types of interactions with cell wall polymers.

Arabinogalactan protein 31 (AGP31), a putative network-forming protein in Arabidopsis thaliana cell walls?

BACKGROUND AND AIMS: Arabinogalactan protein 31 (AGP31) is a remarkable plant cell-wall protein displaying a multi-domain organization unique in Arabidopsis thaliana: it comprises a predicted signal peptide (SP), a short AGP domain of seven amino acids, a His-stretch, a Pro-rich domain and a PAC (PRP-AGP containing Cys) domain. AGP31 displays different O-glycosylation patterns with arabinogalactans on the AGP domain and Hyp-O-Gal/Ara-rich motifs on the Pro-rich domain. AGP31 has been identified as an abundant protein in cell walls of etiolated hypocotyls, but its function has not been investigated thus far. Literature data suggest that AGP31 may interact with cell-wall components. The purpose of the present study was to identify AGP31 partners to gain new insight into its function in cell walls. METHODS: Nitrocellulose membranes were prepared by spotting different polysaccharides, which were either obtained commercially or extracted from cell walls of Arabidopsis thaliana and Brachypodium distachyon. After validation of the arrays, in vitro interaction assays were carried out by probing the membranes with purified native AGP31 or recombinant PAC-V5-6xHis. In addition, dynamic light scattering (DLS) analyses were carried out on an AGP31 purified fraction. KEY RESULTS: It was demonstrated that AGP31 interacts through its PAC domain with galactans that are branches of rhamnogalacturonan I. This is the first experimental evidence that a PAC domain, also found as an entire protein or a domain of AGP31 homologues, can bind carbohydrates. AGP31 was also found to bind methylesterified polygalacturonic acid, possibly through its His-stretch. Finally, AGP31 was able to interact with itself in vitro through its PAC domain. DLS data showed that AGP31 forms aggregates in solution, corroborating the hypothesis of an auto-assembly. CONCLUSIONS: These results allow the proposal of a model of interactions of AGP31 with different cell-wall components, in which AGP31 participates in complex supra-molecular scaffolds. Such scaffolds could contribute to the strengthening of cell walls of quickly growing organs such as etiolated hypocotyls.

Large-scale analysis of the PAC domain structure of arogenate dehydratases reveals their evolutionary patterns in angiosperms.

Arogenate dehydratase (ADT) is the key limiting enzyme of plant phenylalanine biosynthesis, but some ADTs display a prephenate decarboxylase/dehydratase activity-conferring (PAC) domain. The genome resources of 70 species were employed to identify genes and outline their characteristics, especially the number and type of PAC domain structures. We obtained 522 ADTs, and their size, exon number, amino acid number and putative protein isoelectric point greatly varied from 306 to 2520 bp, 1 to 15, 101 to 839 and 4.37 to 11.18, respectively. We classified the ADTs into Class α (without a PAC domain) (115, 22.0 %), ß (with a type I PAC domain) (244, 46.7 %) and γ (with a type II PAC domain) (163, 31.2 %), and their distribution frequencies exhibited large differences among various branches of angiosperms. We found that Class γ members are more conserved than Class ß members, although they commonly experienced multiple duplication events and strong purifying selection, which resulted in a small number, and the putative origin order was from Class α to ß and then to γ. In addition, the co-occurrence of both Class ß and γ members could ensure the survival of angiosperms, while their optimized composition and strategically intertwined regulation may facilitate core eudicot success.