Isolation and characterization of 1-palmitoyl-2-linoleoyl-sn-glycerol as a hormogonium-inducing factor (HIF) from the coralloid roots of Cycas revoluta (Cycadaceae)

Yasuyuki Hashidoko,Manato Tanaka,Kanako Murata,Hiroaki Nishizuka,Makoto Hashimoto,Yuta Murai

doi:10.1038/s41598-019-39647-8

Abstract

Coralloid roots are specialized tissues of cycads (Cycas revoluta) that are involved in symbioses with nitrogen-fixing Nostoc cyanobacteria. We found that a crude methanolic extract of coralloid roots induced differentiation of the filamentous cell aggregates of Nostoc species into motile hormogonia. Hence, the hormogonium-inducing factor (HIF) was chased using bioassay-based isolation, and the active principle was characterized as a mixture of diacylglycerols (DAGs), mainly composed of 1-palmitoyl-2-linoleoyl-sn-glycerol (1), 1-palmitoyl-2-oleoyl-sn-glycerol (2), 1-stearoyl-2-linolenoyl-sn-glycerol (3), and 1-stearoyl-2-linoleoyl-sn-glycerol (4). Enantioselectively synthesised compound 1 showed a clear HIF activity at 1 nmol (0.6 µg) disc−1 for the filamentous cells, whereas synthesised 2-linoleoyl-3-palmitoyl-sn-glycerol (1′) and 1-palmitoyl-2-linoleoyl-rac-glycerol (1/1′) were less active than 1. Conversely, synthesised 1-linoleoyl-2-palmitoyl-rac-glycerol (8/8′) which is an acyl positional isomer of compound 1 was inactive. In addition, neither 1-monoacylglycerols nor phospholipids structurally related to 1 showed HIF-like activities. As DAGs are protein kinase C (PKC) activators, 12-O-tetradecanoylphorbol-13-acetate (12), urushiol C15:3-Δ10,13,16 (13), and a skin irritant anacardic acid C15:1-Δ8 (14) were also examined for HIF-like activities toward the Nostoc cells. Neither 12 nor 13 showed HIF-like activities, whereas 14 showed an HIF-like activity at 1 nmol/disc. These findings appear to indicate that some DAGs act as hormogonium-inducing signal molecules for filamentous Nostoc cyanobacteria.

Highlights

Under certain environmental conditions, Nostoc species form motile filaments of cells referred to as hormogonia[7,8]
When dense filamentous colonies in the stationary phase were transferred into fresh but nitrogen-poor medium, frequent induction of hormogonium differentiation has often been observed in some Nostocean cyanobacteria, suggesting that nitrogen starvation triggers induction of hormogonia
Similar induction of hormogonia was sometimes observed even in a fresh medium containing sufficient nitrogen sources. This observation suggests the presence of hormogonia repressing factors in the spent medium, and removal of the factors should administer an indirect induction of hormogonium differentiation. It has for a long time been believed that hormogonium induction in symbiotic Nostocean cyanobacteria is promoted by the host plants[8]

Summary

Introduction

Nostoc species form motile filaments of cells referred to as hormogonia[7,8]. This observation suggests the presence of hormogonia repressing factors in the spent medium, and removal of the factors should administer an indirect induction of hormogonium differentiation It has for a long time been believed that hormogonium induction in symbiotic Nostocean cyanobacteria is promoted by the host plants[8]. Given the nature of such symbiotic interactions, host cycad and Nostocean symbiont would exchange chemical signals during symbiotic events (Supplementary Fig. S1)[8,14,15], similar to those characterizing the symbiosis between nodulation bacteria and leguminous plants[16,17,18,19] In this regard, host plants are known to produce exudates known as a hormogonium-inducing factors (HIFs)[14,20,21]. In incubation at 30 °C for 5 weeks, the fraction eluted with 100% EtOAc (equivalent to 2 g of fresh coralloid roots/10 mL medium) showed pronounced cyanobacterial cell growth, suggesting the presence of certain signaling factor(s) for growth stimulation of the Nostocean cyanobacteria

Methods

Results

Conclusion