Recipient Organization
CORNELL UNIVERSITY
(N/A)
ITHACA,NY 14853
Performing Department
PLANT BIOLOGY
Non Technical Summary
The shoot apical meristem of flowering plants is the ultimate source of all shoot-borne organs in the plant. Hence, the elucidation of meristematic programs of gene expression is required to understand plant development. Meristems are divided into distinct tissue layers and functional zones. This structural subdivision is an integral component of meristem function; quiescent meristems fail to maintain distinctive zonation (Rhinne and Van Der Schoot, 1998; Gisel et al., 1999). Traditional genetic analyses of meristem function and leaf development, in which mutations of a single gene are characterized at the phenotypic and molecular level, can be sluggish ventures and are often hampered by genetic redundancy. Discerning the global gene expression patterns of different meristematic domains promises to provide new and abundant information vital to our understanding of meristem maintenance, and lateral organogenesis. However, the small size of the meristem and its intimate
association with tiny leaf primordia present significant technical challenges to the isolation of shoot meristems via traditional dissection. More impractical is the use of surgery to isolate and analyze distinct, meristematic subdomains. Thus, previous analyses of global gene expression patterns in plant apices have utilized samples prepared from an assortment of tissues samples, including whole meristems that are abundantly contaminated by adjoining leaf, stem, and inflorescence tissues. The advent of the use of laser microdissection in plant developmental research enables analyses of discrete microdomains within the shoot meristem and leaf primordia where key developmental pattering events occur that control plant morphology.
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Goals / Objectives
Plants are comprised of a series of repeated lateral organ segments that develop from small clumps of morphogenic tissue called shoot meristems. The leaf is a ground state lateral organ; the developmental mechanisms generating leaves are repeated and modified during development of inflorescences and flowers. Thus, an understanding of leaf development is central to our comprehension of overall plant development. Previous molecular genetic analyses in diverse plant species have generated testable models for mechanisms of leaf initiation and growth. In addition, comparative approaches are important toward understanding how diverse leaf morphology has evolved, and may be modified for agronomic advantage in the future. This research will exploit recessive mutations that affect the initiation and/or expansion of leaves. Experiments will investigate the expression patterns and functions of these genes, to understand the development and initiation of mersitems, and the
development and morphological evolution of leaves.
Project Methods
Laser dissection microscopy is a powerful technique that permits the isolation of RNA from specific cell types within fixed plant tissues. RNA collected from 1,000-10,000 cells is sufficient for use in microarray analyses of gene expression. The relatively large size of the maize vegetative meristem, approximately 250 founder cells are recruited into the incipient maize leaf, renders this plant especially tractable for this experimental system. We propose to utilize the laser-capture microdissection/microarray technique to capture cells from specific domains of the maize meristem and developing leaves for use in comparative analyses of gene expression. Selected differentially expressed genes will be verified by real time RT-PCR and in situ analyses of transcript accumulation in maize tissues. Reverse genetic analyses of insertion-mutagenized, differentially expressed genes in maize and Arabidopsis will enable functional analyses of candidate genes involved in meristem
organization and leaf initiation. These experiments will microdissect gene expression patterns in meristems and leaf primordia, and promise to provide novel insight into mechanisms of plant development The maize/Arabidopsis genes NARROW SHEATH (NS)/PRESSED FLOWER (PRS) are required for the initiation of lateral leaf domains from shoot meristems. Maize and Arabidopsis leaves are simple, whereas tomato forms compound leaves comprised of many leaflets. The NS/PRS gene will be cloned from tomato, and molecular-genetic analyses will determine the function of this gene during the development of compound tomato leaves. The developmental timing and meristematic tissue layer-specificity of PRS accumulation and function will be analyzed with fluorescently tagged proteins and specific antibodies. These experiments will investigate mechanisms of PRS non-cell autonomy, wherein PRS functions beyond those cells and tissues in which PRS is expressed. The RAGGEDSEEDLING2 gene will be identified and
analyses of RGD2 expression during maize development will generate models for how RGD2 functions during the lateral growth and expansion of leaves.