Functional analysis of proton-coupled sucrose transport

It has long been known that plants, as photoautotrophic, multicellular organisms, transport sugars and amino acids from sites of primary assimilation to heterotrophic tissues that must import organic nutrients to support growth and development (Gifford & Evans, 1981; Thorne, 1985 ). For many years, plant biologists viewed the distribution of organic nutrients as a fairly simple and straightforward process involving a small number of transporters that loaded the phloem with sucrose and amide amino acids, and complementary carriers that moved these metabolites across the plasma membrane of cells importing the compounds. Underlying assumptions were that a relatively small number of substrates move systemically in the plant, and that most metabolites were synthesized in place as needed. While this model is a good approximation for the bulk of assimilate partitioning, we now recognize that transport of metabolites and other small molecules is much more complex at the cellular level. For example, sucrose partitioning in Arabidopsis relies on transporters that are part of a closely related gene family with nine members ( Kuhn et al ., 1999 ); for amino acids there are many more transporters that are represented by several gene families ( Fischer et al ., 1998 ; Ortiz-Lopez et al ., 2000 ). In both cases, these transporters are differentiated from one another by differences in their kinetic properties, expression patterns and responses to developmental and environmental cues. Other transporters that facilitate the movement of multiple classes of sugars, peptides, metabolites, osmoprotectants and nucleic acid bases have also been identified ( Schwacke et al ., 1999 ; Buttner & Sauer, 2000 ; Gillissen et al ., 2000 ; Noiraud et al ., 2001 ; Stacey, 2002). Thus, there is a complex and rich exchange of organic molecules between adjacent cells, tissues and organs that not only satisfies nutritional needs, but also conveys molecules that contribute to physiological responses and genetic programs regulating plant growth and development ( Chiou & Bush, 1998 ; Bush, 1999 ; Fletcher, 2002 ; Forde, 2002 ; Kessler & Baldwin, 2002 ; Lalonde et al ., 2003 ). These new insights regarding the complexity of transport processes in the physiology of plant growth has placed considerable importance on defining the transport properties, expression patterns and functional contributions of each transporter. This chapter outlines some of the key methods and approaches used to define the kinetics, expression patterns and functions of transporters that move organic molecules across the plasma membrane. To simplify the presentation, I focus on proton-sucrose symporters and some of the seminal methods that have been used to define the functional contributions of this important family of plant transport proteins.