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Research Focus

Fast moves in arbuscular mycorrhizal symbioticsignalling

Sally E. Smith1, Susan J. Barker2 and Yong-Guan Zhu1,3

1 Soil and Land Systems, School of Earth and Environmental Sciences, Waite Campus, DP636, The University of Adelaide,

Adelaide, SA 5005, Australia2 School of Plant Biology M084, University of Western Australia, 35 Stirling Highway, Crawley, WA 6009 Australia3 Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, China

Update TRENDS in Plant Science Vol.11 No.8

Exciting research looking at early events in arbuscularmycorrhizal symbioses has shown how the fungus andplant get together. Kohki Akiyama et al. have demon-strated that strigolactones in root exudates are fungalgerm tube branching factors, and Arnaud Besserer et al.found that these compounds rapidly induce fungalmitochondrial activity. Andrea Genre et al. have shownthat subsequent development of appressoria on hostroots induces construction of a transient prepenetrationapparatus inside epidermal cells that is reminiscent ofnodulation infection.

Arbuscular mycorrhiza: a beneficial symbiosis gives upsecretsOne intriguing question about arbuscular mycorrhizalsymbioses (AMS, see Box 1) is how the partnershipsbetween the fungal symbionts and the host roots areestablished without triggering rejection. Research hasbeen inhibited by some key features of AMS: lack of partnerspecificity, obligate status of the fungi, highly compatibleinteractions and non-synchronous development. Here wehighlight three articles that have made significant contri-butions towards our understanding of the early interac-tions between symbionts.

Plant signals stimulate fungal activity and transition toa symbiosis-ready stateKohki Akiyama et al. [1] used a neat and simple bioassay ofAM fungal germ tube branching [2] to identify and hencechemically characterize active compounds in root exudatesthat lead to morphogenetic changes in advance of rootcolonization. These changes appear crucial in convertinggerm tubeswith limited growth potential into presymbioticmycelium that has the capacity to initiate colonization ofroots, which is a crucial step in the life of an obligatesymbiont. The changes (shown earlier with partially pur-ified root exudates and now with purified compounds)involve rapid alterations in gene expression, an increasein mitochondrial activity (shown in vivo with Mitotrackergreen and also immunologically) as well as respiration rateand, �5 h after stimulation, increased branching [3–5].The active compounds are strigolactones and are effectiveat extremely low concentrations. This suggests that theyprobably act through a signalling pathway that can lead to

Corresponding author: Smith, S.E. (sally.smith@adelaide.edu.au)Available online 12 July 2006.

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fully effective catabolism of lipids, which are the majorcarbon currency of AM fungi. Strigolactones havepreviously been identified because of their importance instimulating germination of the economically damagingroot parasites Striga and Orobanche. The new findingslead the way to controlled investigation of changes infungal metabolism that occur as AM fungal germ tubesare stimulated, and should allow mycorrhizal researchersto capitalize on knowledge of the molecular interactionsinvolved in the initiation of root parasite infections [6] andthe tools that are being developed to combat them, such aslabelled molecules to identify strigolactone receptors inStriga itself [7]. What is fascinating is the possibility thatthe parasitic members of the advanced angiosperm familyScrophulariaceae might have coopted a recognition anddevelopmental pathway that evolved when the earliestland plants interacted with AM fungi. There is alreadyevidence for such cooption by nitrogen-fixing symbioses(NFS).

Formation of a special plant structure guiding fungalinvasion of rootsAndrea Genre et al. [8] targeted plant events during theinitial colonization step as symbiosis-ready fungal hyphaecontact roots. They showed how epidermal cells assemble aspecial intracellular structure before any cell penetrationoccurs. Using Medicago truncatula root clones expressingGFP-labelledmarkers for plant cytoskeleton and endoplas-mic reticulum (ER), they followed, by elegant confocalmicroscopy, epidermal cell responses to the formationof fungal appressoria and hyphal penetration in livingcells over many hours. First the epidermal cell nucleuswas repositioned immediately under the appressorium(Figure 1a,b). How that event is triggered remainsunknown. Highly regulated events followed, directed byfurther movements of the cell nucleus, leading to theformation of a special prepenetration apparatus (PPA)within �4–5 h of appressorium formation (Figure 1c).The PPA is formed within a cytoplasmic column; labellingshows a high-density array of microtubules and microfila-ment bundles running parallel to the column, associatedwith a region of dense ER cisternae. Once the PPA isformed, fungal entry and growth of hyphae across the cellfollow precisely the track defined by the cytoskeletal andER structures within the column (Figure 1d). Before andduring assembly of the PPA, ENOD11 expression is

370 Update TRENDS in Plant Science Vol.11 No.8

Box 1. Arbuscular mycorrhizal symbioses

Arbuscular mycorrhizas are thought to be the most ancient

terrestrial symbioses, and plant roots and AM fungi (Glomeromy-

cota) have been coevolving for >450 million years. Arbuscular

mycorrhizal symbioses (AMS) pre-date other mycorrhizal types, as

well as nitrogen-fixing symbioses (NFS) in legumes. About 80% of

present-day terrestrial plants form AMS, distributed in a wide range

of habitats, suggesting that these biotrophic and mutualistic

associations have selective advantages for both plant and fungal

symbionts. The main basis of mutualism is bidirectional nutrient

transfer: sugars transfer from plant to fungus, in exchange for

mineral nutrients, particularly phosphorus, from the fungus. This

exchange occurs across specialized interfaces between the sym-

bionts developed deep in the root cortex [15]. The process of

colonization requires a complex signalling network and morphoge-

netic changes in both symbionts (Figure I). Our understanding of

these processes has been greatly enhanced by detailed develop-

mental and molecular studies of symbiotic mutants [10,16].

Figure I. Steps in the colonization of roots by AM fungi. (a) Hyphae (h) on the

surface of a root produce a swollen appressorium (ap) from which a penetra-

ting hypha (arrow) traverses the epidermal cell to reach the cortex where int-

racellular coils (C) develop in Paris-type AM. Abbreviation: V, vesicle. (b)

Cortical Arum-type colonization involves the spread of intercellular hyphae (ih)

and the development of highly branched intracellular arbuscules (arb). Image

(a) S.E. Smith; image (b) reproduced, with permission, from Ref. [17].

initiated in epidermal cells. This gene encodes a proteinthat is believed to be part of the plant extracellular matrix.Genre et al. [8] suggest that the transient PPA is respon-sible for the formation of an intracellular symbioticinterface, including a new membrane structure thatcreates an apoplastic plant compartment separating theinvading fungus from the plant.

It is tempting to speculate, as the authors do, that theinvasion process parallels the way that infection threads‘lead’ rhizobium through legume root hairs into the rootcortex during the establishment of NFS nodules. Plantgenetic overlaps between AMS and NFS are well estab-lished [9,10] and are strengthened by the demonstration ofinduction of ENOD11, which is also involved in NFS

Figure 1. Formation of the prepenetration apparatus (PPA) that facilitates passage of AM

host cell (a) results in initial nuclear movement toward the surface appressorium (b). Th

created during the subsequent transcellular nuclear migration (c). Finally, the AM i

constructed within the cytoplasmic column (d). Colour coding is as follows: cell nucleus

red; ER, white. Reproduced, with permission, from Ref. [8].

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establishment, and by the finding that no PPAs are formedin dmi2 or dmi3 mutants of M. truncatula, which do notbecome infected by AM fungi or rhizobium. The findingsimplicate common symbiosis genes in PPA formation,but also indicate that these are not involved in the primarypre-penetration signalling because nuclear movement inthe epidermal cells occurs normally in the mutants. Theway in which putative ‘myc factors’ are perceived andtransduced to facilitate AM penetration via the PPA willbe a fascinating line of future research.

Coordination and timing of sweet talk between thesymbiontsAM fungal spores can germinate in the soil in the absenceof any plants. The root signals now identified [1,5] alert thefungi to the presence of a potential host and hence a vitalsource of organic carbon. The papers show that AM-activestrigolactones are produced by Lotus and Sorghum andthat they induce responses in phylogenetically distinct anddistant AM fungi. It seems reasonable to predict thatthey are extremely widespread plant products. However,Arnaud Besserer et al. [5] caution that other classes ofactive molecule must not be ruled out. The increasedbranching of fungal germ tubes probably increases thechances of root contact, and changes in gene expressionand respiratory activity convert the fungus to aninfection-ready state. When contact is made, fungalmorphogenesis changes once more, with the formation ofslightly swollen appressoria on the root epidermis (Box 1).Now fungal signals alert the plant, which prepares the wayfor intracellular penetration and forms the PPA andinterfacial structures that lead the fungus inward towardsthe root cortex [8]. The initial stages of colonization arecompleted in 10 h. Elucidation of the next stages ofdevelopment will be more challenging because theintracellular interfaces involved in nutrient transferbetween symbionts develop within root cortical cells. Weanticipate that formation of intracellular arbuscules andcoils will involve highly coordinated cellular processes,similar to those described in epidermal cells, but probablymore complicated. The development of cortical interfacesprobably takes a further 1–2 days after penetration andinvolves nuclear repositioning and changes in expressionand localization of the membrane and matrix proteinsthat facilitate nutrient transfers [11]. ENOD11 expressionis also induced in these cells, independently of epidermal

fungi through epidermal cells. Appressorium formation on the outer surface of the

is is followed by the assembly of the transient PPA within the cytoplasmic column

nfection hypha crosses the epidermal cell through the apoplastic compartment

, dark brown; plasma membrane, light brown; microtubules, green; actin bundles,

Update TRENDS in Plant Science Vol.11 No.8 371

expression [12]. Intriguing new data also implicatea reactive oxygen species-inactivating system in signaltransduction between the symbionts [13] andhaemoglobin-encoding gene in suppression of NO-baseddefence processes during arbuscule formation [14].Advances in our understanding of these late stages wouldbe greatly facilitated by an extended range of mutants.

Developmental diversity?The research discussed here understandably used a lim-ited range of fungi and plants that best suited the technicalchallenges. But we know that there is considerable devel-opmental and functional diversity and even limited speci-ficity among the symbioses that are formed between thehuge array of potential host plants and the 120 or so fungiin the Glomeromycota. Do the root and fungal signals andtheir receptors carry the necessary information to encom-pass this diversity in the developmental programmes? Anddoes the limited cortical colonization of some mutantsnormally blocked at the surface result from bypassingepidermal cell penetration and thus avoiding the require-ment for triggered formation of a PPA? This again is for thefuture.

AcknowledgementsOur collaborations are funded by the Australian Research Council andthe Natural Science Foundation of China (40225002).

References1 Akiyama, K. et al. (2005) Plant sesquiterpenes induce hyphal

branching in arbuscular mycorrhizal fungi. Nature 435, 824–8272 Nagahashi, G. and Douds, D.D. (1999) Rapid and sensitive bioassay to

study signals between root exudates and arbuscular mycorrhizal fungi.Biotechnol. Tech. 13, 893–897

3 Buee, M. et al. (2000) The pre-symbiotic growth of arbuscularmycorrhizal fungi is induced by a branching factor partially purifiedfrom plant root exudates. Mol. Plant Microbe Interact. 13, 693–698

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4 Tamasloukht, M. et al. (2003) Root factors induce mitochondrial-related gene expression and fungal respiration during thedevelopmental switch from asymbiosis to presymbiosis in thearbuscular mycorrhizal fungus Gigaspora rosea. Plant Physiol. 131,1468–1478

5 Besserer, A. et al. (2006) Strigolactones stimulate arbuscularmycorrhizal fungi by activating mitochondria. PLoS Biol. 4 (7), e226

6 Yoder, J.I. (2001) Host–plant recognition by parasiticScrophulariaceae. Curr. Opin. Plant Biol. 4, 359–365

7 Reizelman, A. et al. (2003) Synthesis and bioactivity of labelledgermination stimulants for the isolation and identification of thestrigolactone receptor. Org. Biomol. Chem. 1, 950–959

8 Genre, A. et al. (2005) Arbuscular mycorrhizal fungi elicit a novelintracellular apparatus in Medicago truncatula root epidermal cellsbefore infection. Plant Cell 17, 3489–3499

9 Kistner, C. et al. (2005) Seven Lotus japonicus genes required fortranscriptional reprogramming of the root during fungal andbacterial symbiosis. Plant Cell 17, 2217–2229

10 Harrison, M.J. (2005) Signaling in the arbuscular mycorrhizalsymbiosis. Annu. Rev. Microbiol. 59, 19–42

11 Karandashov, V. and Bucher, M. (2005) Symbiotic phosphate transportin arbuscular mycorrhizas. Trends Plant Sci. 10, 22–29

12 Chabaud, M. et al. (2002) Targeted inoculation ofMedicago truncatulain vitro root cultures reveals MtENOD11 expression during earlystages of infection by arbuscular mycorrhizal fungi. New Phytol.156, 265–273

13 Lanfranco, L. et al. (2005) Themycorrhizal fungusGigasporamargaritapossesses a CuZn superoxide dismutase that is up-regulated duringsymbiosis with legume hosts. Plant Physiol. 137, 1319–1330

14 Vieweg, M.F. et al. (2005) Two genes encoding different truncatedhemoglobins are regulated during root nodule and arbuscularmycorrhiza symbioses of Medicago truncatula. Planta 220, 757–766

15 Smith, S.E. and Read, D.J. (1997) Mycorrhizal Symbiosis, (Ed 2).Academic Press

16 Parniske, M. (2004) Molecular genetics of the arbuscular mycorrhizalsymbiosis. Curr. Opin. Plant Biol. 7, 414–421

17 Smith, S.E. et al. (2004) Functional diversity in arbuscular mycorrhizal(AM) symbioses: the contribution of the mycorrhizal P uptake pathwayis not correlated with mycorrhizal responses in growth or total Puptake. New Phytol. 162, 511–524

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