Immunolocalization of meiotic proteins in Brassicaceae: method 1.

Plant meiosis studies have enjoyed a fantastic boom in recent years with the use of Arabidopsis thaliana as an important model species for developmental studies because of its small genome, short life cycle, and large mutant collections. Unlike other eukaryotic models, plant meiosis does not display strict checkpoints and rarely commits to apoptotic processes, which makes it possible to investigate the whole meiotic process (spanning from premeiotic interphase to spore formation) in knockout mutants. In this chapter we describe a protocol for immunolabelling Arabidopsis and Brassica meiotic proteins on robustly spread chromosomes. This protocol allows the detection of a large range of proteins on well-preserved chromosomes and throughout the entire meiotic process.

Arabidopsis formin AtFH6 is a plasma membrane-associated protein upregulated in giant cells induced by parasitic nematodes.

Plant-parasitic nematodes Meloidogyne spp induce an elaborate permanent feeding site characterized by the redifferentiation of root cells into multinucleate and hypertrophied giant cells. We have isolated by a promoter trap strategy an Arabidopsis thaliana formin gene, AtFH6, which is upregulated during giant cell formation. Formins are actin-nucleating proteins that stimulate de novo polymerization of actin filaments. We show here that three type-I formins were upregulated in giant cells and that the AtFH6 protein was anchored to the plasma membrane and uniformly distributed. Suppression of the budding defect of the Saccharomyces cerevisiae bni1Delta bnr1Delta mutant showed that AtFH6 regulates polarized growth by controlling the assembly of actin cables. Our results suggest that AtFH6 might be involved in the isotropic growth of hypertrophied feeding cells via the reorganization of the actin cytoskeleton. The actin cables would serve as tracks for vesicle trafficking needed for extensive plasma membrane and cell wall biogenesis. Therefore, determining how plant parasitic nematodes modify root cells into giant cells represents an attractive system to identify genes that regulate cell growth and morphogenesis.