Functions of the XTH Gene Family of Arabidopsis

Work on the XTH gene family is supported by the NSF 2010 Project.

The cell wall is a major contributor to plant structural properties, but it is not simply a static framework. The cell wall is a dynamic compartment that responds to the changing needs of the plant during growth and development. In addition, wall components have also been shown to have cell-signaling capabilities. Wall modification can affect properties of the cell and, as a consequence, the overall physiology of the plant. Despite these critical roles of the plant cell wall, the molecular bases of wall modifications and how they are regulated are largely unknown. However, a variety of proteins capable of modifying wall constituents have been identified and are being characterized; here, we propose to elucidate the functions of the XTHs.

Xyloglucan endotransglucosylases/hydrolases. Xyloglucan is a major component of the dicot cell wall. This polysaccharide can form hydrogen bonds with cellulose microfibrils and may act as a tether between adjacent microfibrils, restraining cell expansion and enabling the generation of turgor. In vitro, XTHs catalyze cleavage of xyloglucan polymers and transfer newly generated reducing ends to other xyloglucan chains; some XTHs also have the ability to hydrolyze xyloglucan polymers without subsequent transglycosylation. Although these enzymes have the powerful potential to modify a major structural component of the cell wall, the physiological consequences of XTH activity remain unknown.

The discovery (Xu et al., 1995 PDF ) that one of the mechanostimulus-responsive TCH genes of Arabidopsis encodes an XTH revealed the possible physiological relevance of the unusual regulation of expression of the TCH genes. That is, modification of the cell wall by TCH4 may be a mechanism for either enhancing mechanical properties or altering cell expansion processes. Regulation of genes en coding cell wall-modifying enzymes, such as TCH4, may underlie plant morphogenetic responses to the environment.(Figure reproduced with permission from Xu et al., 1995 PDF, Plant Cell.)

The XTH family. Efforts to elucidate the biochemical, cellular and physiological functions of TCH4 have led us to the finding of a gene family encoding related proteins in Arabidopsis (Xu et al. 1996 PDF; Campbell & Braam 1999a PDF; Campbell & Braam 1999b PDF ); with the completion of the genomic sequencing of Arabidopsis at least 33 XTH-related sequences have been identified (Rose et al., 2002 PDF).(Link to XTH gene family table) (Link to phylogenetic tree) (Link to chromosomal locations)

Analyses of the expression behaviors of the XTH genes leads to evidence of where and when gene products may function and provides insights to guide rational approaches to investigate function through reverse genetics. Current information are at the following links (i) touch-induction and darkness-induction (Lee et al., 2005 PDF); Genevestigator (website link, Zimmerman et al., 2004 PDF)-derived (ii) developmental-stage expression, (iii) organ expression, (iv) biotic stimulus-induced expression; (v) abiotic stimulus-induced expression; (vi) root tissue expression (website link; Birnbaum et al., 2003 PDF ), (vi) MPSS (website link; Meyers et al., 2004 PDF ), (vii) ESTs (www.arabidopsis.org), and (viii) XTH:GUS expression patterns.

XTH:GUS transgenic and homozygous xth mutant seeds are available upon request and are available from the Arabidopsis Biological Resource Center.

These expression data reveal that the Arabidopsis XTHs are likely expressed in every developmental stage from seed germination through flowering. All organs show XTH::GUS expression and most, if not all, are found to express multiple XTH::GUS genes. These data sugest that XTHs may contribute to morphogenesis at every developmental stage and in every plant organ. Different XTHs have remarkably diverse and distinct expression patterns indicating that paralogous gene have evolved differential expression reguation perhaps contributing to the maintenance of the large gene family. Extensive overlap in XTH expression patterns is evident; thus, XTHs may act combinatorially in determining wall properties of specific tissues or organs.

We are investigating the functions of the XTH family. XTHs are defined by having greater than 30% sequence identity over the coding region. We aim to reveal the physiological roles of XTHs in morphogenesis and response to diverse environments. Biochemical properties of the proteins are being assessed (Campbell & Braam 1998 PDF ; Campbell & Braam 1999a PDF ; Steele et al. 2001 PDF ); in addition, we investigate the controls and mechanisms of XTH regulation and elucidate how wall modifications may contribute to responses to environmental stimuli and how feedback pathways are involved in wall homeostasis (Iliev et al., 2002 PDF ). Identification and characterization of mutants is ongoing. For example, XTH15 mutants fail to properly elongate hypocotyls at elevated temperature, indicating a role for XTH15 in cell wall integrity in seedling hypocotyls. DNA sequencing and RNA analyses indicate that the two mutations are allelic and unlikely to result in gene product. The hypocotyl defect of xth15-2 is rescued by addition of an additional copy of the XTH15 locus.  XET activity detected in extracts from wild type and mutant hypocotyls indicate that the xth15 mutants have reduced XET activity.  Structural analyses that indicate that internal hypocotyl cell layers of the xth15 mutants are collapsed under these growth conditions. The xth15 mutants also fail to show a prominent feature of thigmomorphogenesis, that is, a repetitive touch-induced delay in the transition to flowering.  Whereas wild type are delayed in inflorescence stem development if subjected to repetitive touch stimulation throughout their lives, timing of xth15 -1 and xth15-2 plants inflorescence generation is unaffected by repetitive touch stimulation.  XTH24, in contrast, is nonessential for the touch-induced delay of flowering since the xth24-1 mutant flowering delay is comparable to wild type (Col-0). We have ongoing collaborations with Ewa Mellerowiz, Umea, Sweden, to characterize xylem cell size and wood composition and ultrastructure in a subset of XTH mutants. To aid in the characterization of wall changes attributable to XTHs, we are conducting NMR analyses to evaluate carbon distribution among lignin, carbohydrates, lipids and proteins in mutants versus wild type. Fourier Transform Infrared Spectroscopy (FTIR) analysis in collaboration with Maureen McCann and Nick Carpita (Purdue University) provides evidence that the xth mutants have altered wall architectures (in progress). This work, supported by the NSF, will also lead to insights into the relevance of the large XTH family by identifying the specific and the shared regulatory and functional characteristics of the diverse family members.

(2010 Abstract)
This material is based upon work supported by the National Science Foundation under Grant No. 0313432. Any opinions, findings, and conclusions or recommendations expressed in this material are those of the author and do not necessarily reflect the views of the National Science Foundation.