Evidence for Variation in the Optimal Translation Initiation Complex: Plant eIF4B,eIF4F,and eIF(iso)4F Differentially Promote Translation of mRNAs

Expansins are cell wall proteins associated with the process of plant growth. However, investigations in which expansin gene expression has been manipulated throughout the plant have often led to inconclusive results. In this article, we report on a series of experiments in which overexpression of expansin was targeted to specific phases of leaf growth using an inducible promoter system. The data indicate that there is a restricted window of sensitivity when increased expansin gene expression leads to increased endogenous expansin activity and an increase in leaf growth. This phase of maximum expansin efficacy corresponds to the mid phase of leaf growth. We propose that the effectiveness of expansin action depends on the presence of other modulating factors in the leaf and we suggest that it is the control of expression of these factors (in conjunction with expansin gene expression) that defines the extent of leaf growth. These data help to explain some of the previously observed variation in growth response following manipulation of expansin gene expression and highlight a potential linkage of the expression of modifiers of expansin activity with the process of exit from cell division.Expansins were initially identified as cell wall proteins that had the ability to promote the extension of plant tissue in vitro (McQueen-Mason et al., 1992). Further work on these proteins and the genes encoding them has revealed a picture in which, although a general correlation with growth has often been substantiated, it is clear that control of growth is a much more complex process than the control of expression of a single protein type (for review, see Cosgrove, 2000; Lee et al., 2001; Li et al., 2003). In addition, although it is clear that expansins play a role in many growth processes, there are a number of open questions about exactly how expansins contribute to these processes. First, we still have a very limited understanding of the molecular mechanism of expansin action. Efforts to identify classical enzymatic activities associated with expansins have proven fruitless (McQueen-Mason and Cosgrove, 1995; Li and Cosgrove, 2001) and the remaining, somewhat speculative, interpretation is that expansins intercalate within carbohydrate matrices in the cell wall, leading to transient loosening of noncovalent interactions and, thus, the ability of these matrices to move relative to each other (McQueen-Mason and Cosgrove., 1994). In addition, by unlocking aspects of the molecular architecture of the cell wall, expansins may allow access of other cell wall proteins/enzymes to particular substrates. Depending on the nature of these other proteins/enzymes, expansin activity could thus be associated not only with growth processes, but also with cell wall modifications linked with differentiation. Such a mechanism would help to explain observations (described below) that the effectiveness of expansin action appears to be context dependent and is not only associated with changes in plant growth but also with differentiation.Various analyses have revealed that expansins are present in a wide range of plants, including bryophytes, ferns, angiosperms, and conifers (Hutchison et al., 1999; Kim et al., 2000; Schipper et al., 2002). Moreover, they are generally encoded by relatively large gene families whose members often show distinct patterns of gene expression (Kende et al., 2004). Some of these expression patterns correlate with growth processes, such as root growth (Wu et al., 1996), internode growth (Cho and Kende, 1997), leaf growth (Muller et al., 2007), and cotton (Gossypium hirsutum) fiber growth (Ruan et al., 2001), whereas others correlate with events of differentiation, such as fruit ripening (Rose et al., 1997; Brummell et al., 1999b), grass tiller formation (Reidy et al., 2001), and endosperm breakdown (Chen and Bradford, 2000). In addition, some novel nonplant expansin activities have been identified that suggest that pathogens may induce altered cell wall structure via an expansin-mediated mechanism (Qin et al., 2004). Since in vitro assays have suggested that the activities of expansins extracted from different sources tend to be similar (Cosgrove, 2000), it has been proposed that this tissue, organ, and environmental specificity of expression pattern reflects a specialized role for expansins in specific contexts rather than any major difference in activity of the protein. As stated above, this specific function may depend on the presence (or absence) of tissue-specific cofactors, the nature of which is as yet unclear.In addition to biochemical approaches to understanding expansin function, numerous groups have undertaken transgenic experiments to alter expansin gene expression in plants to observe the outcome on plant phenotype. Although some successes with antisense strategies have been reported (Brummell et al., 1999a; Cho and Cosgrove, 2000), the encoding of expansin by large gene families means that genetic redundancy poses a significant problem for such approaches (e.g. Schipper et al., 2002). Simple overexpression strategies to alter expansin activity may also be difficult to interpret. For example, when expansins were constitutively overexpressed throughout Arabidopsis (Arabidopsis thaliana), tomato (Solanum lycopersicum), and rice (Oryza sativa) plants, the outcomes tended to be pleiotropic, including a decrease in overall plant growth (Cho and Cosgrove, 2000; Rochange et al., 2001; Choi et al., 2003). However, when altered expansin expression was targeted more specifically to a particular tissue or organ, then more easily interpretable results were obtained. For example, when altered expansin expression was directed to the developing leaf petiole in Arabidopsis, altered leaf growth was observed (Cho and Cosgrove, 2000), consistent with the idea that expansins promote growth, and when inducible expression of expansin was targeted throughout rice plants, quantitative changes in growth were observed (Choi et al., 2003). The results of these experiments indicate that expansin gene expression can be used as a tool to modulate growth, but that the timing and spatial extent of expression can have a significant influence on the phenotype observed. Again, these data support the hypothesis that the effectiveness of expansin in promoting specific growth or differentiation events is dependent on the presence of particular tissue- or developmental-specific cofactors. So far, little progress has been made on the identification and characterization of these cofactors.In previous work, we reported on the characterization of transgenic lines of tobacco (Nicotiana tabacum) in which a cucumber (Cucumis sativus) expansin (CsEXP1) could be induced by application of a chemical inducer (anhydrotetracycline Ahtet]). In these experiments, we targeted expansin overexpression to localized regions of either the shoot apical meristem or very young leaf primordia, which led to localized promotion of growth (Pien et al., 2001), consistent with the idea that expansins play a role in the endogenous mechanism of leaf initiation (Reinhardt et al., 1998). However, when inductions were performed throughout the plant the resulting phenotypes were variable and difficult to interpret (S. Pien and A. Fleming, unpublished data), in line with other reports (Rochange et al., 2001). To investigate the possibility that this variable response reflected a differential sensitivity to expansin in different tissues at different stages of development, we performed a series of experiments (reported here) in which overexpression of expansin was targeted to specific stages of leaf growth. Our data indicate that the efficacy of expansin action depends on the presence of other factors that are present in a developmentally controlled fashion, so that increased expansin gene expression is only effective in promoting leaf growth during a specific developmental period of leaf growth. This period corresponds to the inflection point of relative growth rate (RGR) and, thus, to the phase of maximum leaf growth rate. An intriguing article by Cookson et al. (2005) reported on potential correlations between various parameters of leaf growth and final leaf size. They found that the best predictor of final leaf size was the maximum value of absolute leaf growth rate. Thus, the experiments reported here identify a novel, developmental control of expansin efficacy in the regulation of leaf growth, investigate the reported correlation between maximal leaf expansion rate and leaf size, and provide an insight into potential means of controlling leaf growth.