RESUMEN
In plants, the ability to produce hydrophobic substances that would provide protection from dehydration was required for the transition to land. This genome-wide investigation outlines the evolution of GDSL-type esterase/lipase (GELP) proteins in the moss Physcomitrium patens and suggests possible functions of some genes. GELP proteins play roles in the formation of hydrophobic polymers such as cutin and suberin that protect against dehydration and pathogen attack. GELP proteins are also implicated in processes such as pollen development and seed metabolism and germination. The P. patens GELP gene family comprises 48 genes and 14 pseudogenes. Phylogenetic analysis of all P. patens GELP sequences along with vascular plant GELP proteins with reported functions revealed that the P. patens genes clustered within previously identified A, B and C clades. A duplication model predicting the expansion of the GELP gene family within the P. patens lineage was constructed. Expression analysis combined with phylogenetic analysis suggested candidate genes for functions such as defence against pathogens, cutin metabolism, spore development and spore germination. The presence of relatively fewer GELP genes in P. patens may reduce the occurrence of functional redundancy that complicates the characterization of vascular plant GELP genes. Knockout lines of GELP31, which is highly expressed in sporophytes, were constructed. Gelp31 spores contained amorphous oil bodies and germinated late, suggesting (a) role(s) of GELP31 in lipid metabolism in spore development or germination. Future knockout studies of other candidate GELP genes will further elucidate the relationship between expansion of the family and the ability to withstand the harsh land environment.
Asunto(s)
Bryopsida , Lipasa , Lipasa/genética , Lipasa/metabolismo , Filogenia , Deshidratación/genética , Esterasas/genética , Esterasas/metabolismo , Bryopsida/genética , Genes de Plantas , Proteínas de Plantas/metabolismo , EsporasRESUMEN
MAIN CONCLUSION: PpORS-produced 2'-oxo-5-pentacosylresorcinol (2'-oxo-C25-RL) restored dehydration tolerance in ors-3, a knockout mutant of PpORS. Feeding experiments with [14C]-2'-oxo-C25-RL suggested the role of PpORS products in cuticular polymer that confer dehydration resistance. 2'-Oxoalkylresorcinol synthase from the moss Physcomitrium (Physcomitrella) patens (PpORS) is the earliest diverged member of plant type III polyketide synthases, and produces very-long-chain 2'-oxoalkylresorcinols in vitro. Targeted knockouts of PpORS (ors) exhibited an abnormal phenotype (increased susceptibility to dehydration), and a defective cuticle in ors was suggested (Li et al., Planta 247:527-541, 2018). In the present study, we investigated chemical rescue of the ors phenotype and also metabolic fates of the PpORS products in the moss. Using C24-CoA as substrate, 2'-oxo-5-pentacosylresorcinol (2'-oxo-C25-RL) and two minor pyrones were first enzymatically prepared as total in vitro products. When a knockout mutant (ors-3) and control strains were grown in the presence of the total in vitro products or purified 2'-oxo-C25-RL, the ability of ors-3 and the control to survive dehydration stress increased in a dose-dependent manner. Structurally analogous long-chain alkylresorcinols also rescued the ors phenotype, although less efficiently. When the moss was grown in the presence of 14C-radiolabeled 2'-oxo-C25-RL, 96% of the radioactivity was recovered only after acid hydrolysis. These findings led us to propose that 2'-oxoalkylresorcinols are the functional in planta products of PpORS and are incorporated into cuticular biopolymers that confer resistance to dehydration. In addition, the earliest diverging ORS clade in phylogenetic trees of plant type III PKSs exclusively comprises bryophyte enzymes that share similar active site substitutions with PpORS. Further studies on these bryophyte enzymes may shed light on their roles in early plant evolution and offer a novel strategy for improving dehydration tolerance in plants.