Laser microdissection reveals that transcripts for five plant and one fungal phosphate transporter genes are contemporaneously present in arbusculated cells.

The establishment of a symbiotic interaction between plant roots and arbuscular mycorrhizal (AM) fungi requires both partners to undergo significant morphological and physiological modifications which eventually lead to reciprocal beneficial effects. Extensive changes in gene expression profiles recently have been described in transcriptomic studies that have analyzed the whole mycorrhizal root. However, because root colonization by AM fungi involves different cell types, a cell-specific gene expression pattern is likely to occur. We have applied the laser microdissection (LMD) technology to investigate expression profiles of both plant and fungal genes in Lycopersicon esculentum roots colonized by Glomus mosseae. A protocol to harvest arbuscule-containing cells from paraffin sections of mycorrhizal roots has been developed using a Leica AS LMD system. RNA of satisfactory quantity and quality has been extracted for molecular analysis. Transcripts for plant phosphate transporters (LePTs), selected as molecular markers for a functional symbiosis, have been detected by reverse-transcriptase polymerase chain reaction assays and associated to distinct cell types, leading to novel insights into the distribution of LePT mRNAs. In fact, the transcripts of the five phosphate transporters (PTs) have been detected contemporaneously in the same arbusculated cell population, unlike from the neighboring noncolonized cells. In addition, fungal H(+)ATPase (GmHA5) and phosphate transporter (GmosPT) mRNAs were found exclusively in arbusculated cells. The discovery that five plant and one fungal PT genes are consistently expressed inside the arbusculated cells provides a new scenario for plant-fungus nutrient exchanges.

[1]  K. Izui,et al.  Knockdown of an arbuscular mycorrhiza-inducible phosphate transporter gene of Lotus japonicus suppresses mutualistic symbiosis. , 2006, Plant & cell physiology.

[2]  A. Osbourn,et al.  Comparative transcriptomics of rice reveals an ancient pattern of response to microbial colonization , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[3]  M. Bucher Functional biology of plant phosphate uptake at root and mycorrhiza interfaces. , 2007, The New phytologist.

[4]  N. Requena,et al.  Symbiotic Status, Phosphate, and Sucrose Regulate the Expression of Two Plasma Membrane H+-ATPase Genes from the Mycorrhizal Fungus Glomus mosseae1 , 2003, Plant Physiology.

[5]  R. Macknight,et al.  Transcript analysis of laser microdissected plant cells , 2006 .

[6]  G. Bécard,et al.  A Diffusible Factor from Arbuscular Mycorrhizal Fungi Induces Symbiosis-Specific MtENOD11 Expression in Roots ofMedicago truncatula 1 , 2003, Plant Physiology.

[7]  M. J. Harrison,et al.  A Medicago truncatula phosphate transporter indispensable for the arbuscular mycorrhizal symbiosis , 2007, Proceedings of the National Academy of Sciences.

[8]  D. Schachtman,et al.  A Lycopersicon esculentum phosphate transporter (LePT1) involved in phosphorus uptake from a vesicular-arbuscular mycorrhizal fungus. , 1999, The New phytologist.

[9]  F. W. Smith,et al.  Cereal phosphate transporters associated with the mycorrhizal pathway of phosphate uptake into roots , 2005, Planta.

[10]  P. Bonfante At the Interface Between Mycorrhizal Fungi and Plants: the Structural Organization of Cell Wall, Plasma Membrane and Cytoskeleton , 2001 .

[11]  I. Jakobsen,et al.  Mycorrhizal Fungi Can Dominate Phosphate Supply to Plants Irrespective of Growth Responses1 , 2003, Plant Physiology.

[12]  P. Bonfante,et al.  Arbuscular Mycorrhizal Fungi Elicit a Novel Intracellular Apparatus in Medicago truncatula Root Epidermal Cells before Infection[W] , 2005, The Plant Cell Online.

[13]  Haiyan Huang,et al.  Transcription Profile Analyses Identify Genes and Pathways Central to Root Cap Functions in Maize , 2006, Plant Molecular Biology.

[14]  R. Balestrini,et al.  The interface compartment in arbuscular mycorrhizae: A special type of plant cell wall? , 2005 .

[15]  M. Bucher,et al.  Symbiotic phosphate transport in arbuscular mycorrhizas. , 2005, Trends in plant science.

[16]  F. A. Smith,et al.  Structure and function of the interfaces in biotrophic symbioses as they relate to nutrient transport. , 1990, The New phytologist.

[17]  A. Pühler,et al.  Overlaps in the Transcriptional Profiles of Medicago truncatula Roots Inoculated with Two Different Glomus Fungi Provide Insights into the Genetic Program Activated during Arbuscular Mycorrhiza1[w] , 2005, Plant Physiology.

[18]  M. Beatty,et al.  The application of laser microdissection to in planta gene expression profiling of the maize anthracnose stalk rot fungus Colletotrichum graminicola. , 2006, Molecular plant-microbe interactions : MPMI.

[19]  L. Lanfranco,et al.  Fungal and plant gene expression in arbuscular mycorrhizal symbiosis , 2006, Mycorrhiza.

[20]  N. Inada,et al.  Novel tissue preparation method and cell-specific marker for laser microdissection of Arabidopsis mature leaf , 2005, Planta.

[21]  E. Hewitt Sand and Water Culture Methods Used in the Study of Plant Nutrition , 1966 .

[22]  Maria J Harrison,et al.  Phosphate in the arbuscular mycorrhizal symbiosis: transport properties and regulatory roles. , 2007, Plant, cell & environment.

[23]  Andrea Genre,et al.  A Diffusible Signal from Arbuscular Mycorrhizal Fungi Elicits a Transient Cytosolic Calcium Elevation in Host Plant Cells1[W] , 2006, Plant Physiology.

[24]  A. Trouvelot,et al.  Mesure du taux de mycorhization VA d'un systeme radiculaire. Recherche de methodes d'estimation ayant une significantion fonctionnelle , 1986 .

[25]  Guohua Xu,et al.  The characterization of novel mycorrhiza-specific phosphate transporters from Lycopersicon esculentum and Solanum tuberosum uncovers functional redundancy in symbiotic phosphate transport in solanaceous species. , 2005, The Plant journal : for cell and molecular biology.

[26]  D. van Tuinen,et al.  Characterization of root colonization profiles by a microcosm community of arbuscular mycorrhizal fungi using 25S rDNA‐targeted nested PCR , 1998, Molecular ecology.

[27]  U. Paszkowski,et al.  Rice phosphate transporters include an evolutionarily divergent gene specifically activated in arbuscular mycorrhizal symbiosis , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[28]  B. Persson,et al.  Functional analysis and cell-specific expression of a phosphate transporter from tomato , 1998, Planta.

[29]  K. Raghothama,et al.  Tomato phosphate transporter genes are differentially regulated in plant tissues by phosphorus. , 1998, Plant physiology.

[30]  Sally E. Smith,et al.  Enzymatic studies on the metabolism of vesicular-arbuscular mycorrhizas. V, Is H+-ATPase a component of ATP-hydrolysing enzyme activities in plant-fungus interfaces ? , 1991 .

[31]  P. Bonfante,et al.  Building a mycorrhizal cell: How to reach compatibility between plants and arbuscular mycorrhizal fungi , 2005 .

[32]  C. Town,et al.  Transcript Profiling Coupled with Spatial Expression Analyses Reveals Genes Involved in Distinct Developmental Stages of an Arbuscular Mycorrhizal Symbiosis Online version contains Web-only data. Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tp , 2003, The Plant Cell Online.

[33]  M. J. Harrison,et al.  A Phosphate Transporter from Medicago truncatula Involved in the Acquisition of Phosphate Released by Arbuscular Mycorrhizal Fungi Article, publication date, and citation information can be found at www.plantcell.org/cgi/doi/10.1105/tpc.004861. , 2002, The Plant Cell Online.

[34]  Timothy Nelson,et al.  Laser microdissection of plant tissue: what you see is what you get. , 2006, Annual review of plant biology.

[35]  M. J. Harrison,et al.  MOLECULAR AND CELLULAR ASPECTS OF THE ARBUSCULAR MYCORRHIZAL SYMBIOSIS. , 1999, Annual review of plant physiology and plant molecular biology.

[36]  R. Macknight,et al.  Be more specific! Laser-assisted microdissection of plant cells. , 2005, Trends in plant science.

[37]  L. Lanfranco,et al.  Expression profiles of a phosphate transporter gene (GmosPT) from the endomycorrhizal fungus Glomus mosseae , 2005, Mycorrhiza.

[38]  M. J. Harrison,et al.  Signaling in the arbuscular mycorrhizal symbiosis. , 2005, Annual review of microbiology.

[39]  K. Ramsay,et al.  Laser capture microdissection: a novel approach to microanalysis of plant-microbe interactions. , 2006, Molecular plant pathology.

[40]  J. Jansa,et al.  A phosphate transporter expressed in arbuscule-containing cells in potato , 2001, Nature.