Laser Microdissection Unravels Cell-Type-Specific Transcription in Arbuscular Mycorrhizal Roots, Including CAAT-Box Transcription Factor Gene Expression Correlating with Fungal Contact and Spread  

Abstract

Arbuscular mycorrhizae (AM) are the most widespread symbioses on Earth, promoting nutrient supply of most terrestrial plant species. To unravel gene expression in defined stages of Medicago truncatula root colonization by AM fungi, we here combined genome-wide transcriptome profiling based on whole mycorrhizal roots with real-time reverse transcription-PCR experiments that relied on characteristic cell types obtained via laser microdissection. Our genome-wide approach delivered a core set of 512 genes significantly activated by the two mycorrhizal fungi Glomus intraradices and Glomus mossae. Focusing on 62 of these genes being related to membrane transport, signaling, and transcriptional regulation, we distinguished whether they are activated in arbuscule-containing or the neighboring cortical cells harboring fungal hyphae. In addition, cortical cells from nonmycorrhizal roots served as a reference for gene expression under noncolonized conditions. Our analysis identified 25 novel arbuscule-specific genes and 37 genes expressed both in the arbuscule-containing and the adjacent cortical cells colonized by fungal hyphae. Among the AM-induced genes specifying transcriptional regulators were two members encoding CAAT-box binding transcription factors (CBFs), designated MtCbf1 and MtCbf2. Promoter analyses demonstrated that both genes were already activated by the first physical contact between the symbionts. Subsequently, and corresponding to our celltype expression patterns, they were progressively up-regulated in those cortical areas colonized by fungal hyphae, including the arbuscule-containing cells. The encoded CBFs thus represent excellent candidates for regulators that mediate a sequential reprogramming of root tissues during the establishment of an AM symbiosis.

DELLA proteins regulate arbuscule formation in arbuscular mycorrhizal symbiosis

“…Transcriptome analyses reveal substantial alterations in the expression of genes encoding enzymes of gibberellic acid (GA) biosynthesis, degradation, and signaling during AM symbiosis (28,(35)(36)(37)(38)(39). Consistent with these alterations, GA levels increase significantly in mycorrhizal roots (40).…”

“…A similar model is proposed for AM symbiosis, and GRAS factors that regulate hyphopodia development (24) and cortical colonization levels have been identified (25), but regulators specific for arbuscule formation are unknown. Transcript profiling and promoter-reporter gene analyses indicate complex changes in plant gene expression in the root cortex during arbuscule development, suggesting that multiple signaling pathways may be involved in arbuscule formation (14,(26)(27)(28)(29)(30). Complex regulation of arbuscule formation might be expected, because the symbiosis is modulated in response to the plant's phosphate status, nitrogen status, and photosynthetic capacity (31)(32)(33)(34).…”

Gene expression in mycorrhizal orchid protocorms suggests a friendly plant–fungus relationship

“…Although chitinases can be induced in pathogenic interactions (Salzer et al 2000), specific class III chitinases have been found to be up-regulated in arbuscular mycorrhizal (AM) Medicago truncatula roots, but not in interactions with pathogens (Salzer et al 2000). Predominant expression of these chitinases was in the arbuscule-containing cells (Bonanomi et al 2001;Hogekamp et al 2011), where they may play a role in suppressing plant defence by the reduction of chitin-like elicitors during the formation of functional symbiotic interfaces (Salzer et al 2000). A similar role may be envisaged in S. vomeracea mycorrhizal protocorms, but further studies are required to investigate regulation of SvChit3 by other biotic and abiotic stimuli.…”

DELLA proteins regulate expression of a subset of AM symbiosis-induced genes inMedicago truncatula

“…5,6 The gene expression changes that underlie symbiotic development have been documented through detailed transcript profiling 7,8 and some transcriptional regulators have been identified, some of which are downstream components of the symbiosis signaling pathway. 4,[9][10][11] Development of AM symbiosis is also influenced by plant hormones and treatment of roots with gibberellic acid (GA) leads either to alterations in fungal entry into the root or abolishes arbuscule development.…”

“…The precise outcome depends on the concentration of GA applied. 12 Genes encoding enzymes of GA biosynthesis and metabolism are differentially regulated during symbiosis 7,[13][14][15][16][17] and associated with the colonized regions of the cortex. 18 Furthermore, both bioactive and inactive GAs are present in mycorrhizal roots.…”