Knock‐down of the MEP pathway isogene 1‐deoxy‐d‐xylulose 5‐phosphate synthase 2 inhibits formation of arbuscular mycorrhiza‐induced apocarotenoids, and abolishes normal expression of mycorrhiza‐specific plant marker genes

Abstract

SummaryThe first step of the plastidial methylerythritol phosphate (MEP) pathway is catalyzed by two isoforms of 1-deoxy-D-xylulose 5-phosphate synthase (DXS1 and DXS2). In Medicago truncatula, MtDXS1 and MtDXS2 genes exhibit completely different expression patterns. Most prominently, colonization by arbuscular mycorrhizal (AM) fungi induces the accumulation of certain apocarotenoids (cyclohexenone and mycorradicin derivatives) correlated with the expression of MtDXS2 but not of MtDXS1. To prove a distinct function of DXS2, a selective RNAi approach on MtDXS2 expression was performed in transgenic hairy roots of M. truncatula. Repression of MtDXS2 consistently led to reduced transcript levels in mycorrhizal roots, and to a concomitant reduction of AM-induced apocarotenoid accumulation. The transcript levels of MtDXS1 remained unaltered in RNAi plants, and no phenotypical changes in non-AM plants were observed. Late stages of the AM symbiosis were adversely affected, but only upon strong repression with residual MtDXS2-1 transcript levels remaining below approximately 10%. This condition resulted in a strong decrease in the transcript levels of MtPT4, an AM-specific plant phosphate transporter gene, and in a multitude of other AMinduced plant marker genes, as shown by transcriptome analysis. This was accompanied by an increased proportion of degenerating and dead arbuscules at the expense of mature ones. The data reveal a requirement for DXS2-dependent MEP pathway-based isoprenoid products to sustain mycorrhizal functionality at later stages of the symbiosis. They further validate the concept of a distinct role for DXS2 in secondary metabolism, and offer a novel tool to selectively manipulate the levels of secondary isoprenoids by targeting their precursor supply.

Knockdown of the Symbiotic Sucrose Synthase MtSucS1 Affects Arbuscule Maturation and Maintenance in Mycorrhizal Roots of Medicago truncatula

“…Here, the generally highly branched intraradical hyphal network is less differentiated, accompanied by alterations in fungal root occupancy, the timing of infection, fungal development, and, most strikingly, in the longevity of the asynchronous AM symbiosis. This arbuscule phenotype matches observations for M. truncatula plants depleted in the AM marker genes MtPT4 (Javot et al, 2007) and MtDsx2 (Floß et al, 2008). Since during nodulation, MtSucS1-as plants also exhibited early symbiosome senescence (Baier et al, 2007), MtSucS1 is likely involved in the C supply to microsymbionts and in the sufficient development of endosymbiotic interfaces in general.…”

“…4, C-K). In mycorrhizal controls, the distribution of the stages young, mature, degenerating, and dead arbuscules (Floß et al, 2008) was 1:2.3:1.4:1.1 (Fig. 4L), depicting a successive colonization process with balanced rates of arbuscule decay and reemergence.…”

“…Hyphal, arbuscular, and vesicular colonization were determined in colonized regions by the magnified (2003) line intersect method (McGonigle et al, 1990). Arbuscule developmental stages (Floß et al, 2008) were studied on eight samples per transgenic line on WGA-AlexaFluor488-stained root segments (Javot et al, 2007) by a Leica SP2 laser-scanning microscope (Leica Microsystems) recording more than 190 arbuscules per biological replicate (c, 2,160; as12, 1,800; as19, 1,900 arbuscules). Image processing was carried out with Adobe Photoshop 7.0 (Adobe Systems) and Leica LCSlight.…”

Transcription Reprogramming during Root Nodule Development in Medicago truncatula

“…Data mining revealed that only six regulators (MT000013, MT003698, MT006389, MT007392, MT009598, MT015410) were also up-regulated during the arbuscular mycorrhizal symbiosis [29], [66], [67]. These were distributed between six TF families and four regulation classes (“bacA”, “diff2”, “fix+” and “NN”) and all of them were also expressed in non symbiotic conditions.…”

Communication and Signaling in the Plant–Fungus Symbiosis: The Mycorrhiza

“…In the case of AM, these have been named mycorradicins and identified as acyclic C14 apocarotenoid polyenes (Bothe et al 1994 ;Klinger et al 1995a ;Schliemann et al 2006) . Although Fester et al (2002a) demonstrated that some highly mycorrhizal roots may completely lack mycorradicin, apocarotenoid synthesis seems to be important for AM establishment because mutants deficient of (Fester et al 2002a) or with reduced carotenoid biosynthesis capacity (Fester 2008) show a reduced development of functional symbiotic structures (Floß et al 2008) . In addition to mycorradicins, esterified mycorradicins and glycosylated C13 cyclohexenone apocarotenoid derivatives can accumulate in mycorrhizal tissues (Maier et al 1995 ;Strack and Fester 2006 ;Schliemann et al 2008) .…”