Pathogens of Arabidopsis

Like other plant species, Arabidopsis thaliana is susceptible to only a limited number of pathogens including viruses, bacteria, fungi, nematodes and insect pests. Diseases resulting from these pathogens have been reported in the wild (Holub et al. 1994, 1995; Tsuji and Somerville 1992) suggesting both the pathogen and the host share an ecological niche and when the appropriate environmental conditions are present disease can occur. Diseases have also been observed in a laboratory setting where the host is deliberately exposed to the pathogen. Regardless of the setting, nature or the laboratory, Arabidopsis responds in a similar fashion as other higher plants when exposed to viral, prokaryotic, or eukaryotic pathogens.

Perhaps the more facile class of pathogen to work with in a laboratory setting are the bacterial pathogens. Bacteria have several advantages over the other classes of pathogens for pathological studies in that they can be cultured in vitro and have relatively rapid generation times (minutes not days). In addition, most bacterial plant pathogens elicit rapid host responses (hours to days) and have pathogenicity and avirulence factors that have been documented in other plant species thereby providing a foundation to begin work in Arabidopsis. Only a small number of bacterial species are pathogenic on Arabidopsis. The predominant bacterial pathogen utilized in Arabidopsis studies is Pseudomonas syringae and the reader is referred to other chapters in this book describing Arabidopsis responses to this pathogen. Additional bacterial pathogens utilized in Arabidopsis research include Erwinia species which are causal agents of soft-rots, Ralstonia species which are causal agents of vascular wilts, and Xanthomonas campestris pathovars which are causal agents of blights and rots and are the focus of this chapter.

The Genus Xanthomonas

The genus Xanthomonas has been well described in several definitive publications (Starr 1981; Leyns et al., 1984; Swings et al., 1993) and the reader is referred to these for more details regarding this genus. In brief, Xanthomonas species are gram-negative rod-shaped aerobic bacteria with polar flagella and are found primarily in association with plants (diseased lesions, soil, plant debris) (Starr 1981). Xanthomonas species produce diagnostic pigments (termed xanthomonadins) and exude large amounts of xanthan gum, an extracellular polysaccharide which is used as a food additive (Williams 1980; Starr 1981; Swings et al., 1993). Xanthomonas species, similar to other plant pathogens, can cause symptoms such as chlorosis, necrosis, cankers, and vascular wilts (Leyns et al., 1984). Collectively, Xanthomonas isolates have been reported to cause disease on ∼400 different monocotyledonous and dicotyledonous species (Leyns et al., 1984), making Xanthomonas a significant plant pathogen. As will be discussed in more detail below, current mechanisms for control of Xanthomonas diseases rely extensively on breeding for resistance and/or cultural practices such as crop rotation and sanitation. An example of the extreme costs that can arise from lack of control of a Xanthomonas species, either through lack of breeding for resistance or ineffective control through cultural practices, is the recent epidemic of citrus canker in the United States (Brown 2001). Attempts to control citrus canker in Florida through an aggressive eradication program have cost over $200 million to date, with no end in sight to control of the disease (Brown 2001).

Reports of Arabidopsis-infecting Isolates of Xanthomonas

With respect to infection of Arabidopsis, only isolates of X. campestris have been reported to cause disease. The species, X. campestris, is a large collection of isolates that share similar biochemical and physiological phenotypes yet exhibit different host specificity and are separated into pathovars to reflect host specificity or host origin (Leyns et al., 1984; Van den Mooter and Swings 1990). Table 1 lists the X. campestris pathovars that have been examined on Arabidopsis accessions. Both resistance and disease, as demonstrated by chlorosis and necrosis on inoculated leaves as well as systemic infections, have been reported on Arabidopsis accessions, suggesting that a full range of host responses occur in Arabidopsis (Tsuji and Somerville 1988; Simpson and Johnson 1990; Davis et al., 1991; Tsuji et al., 1991; Lummerzheim et al., 1993; Parker et al., 1993; Aufsatz and Grimm 1994).

Although a number of these pathovars can incite a response on Arabidopsis, the most prevalent pathovar utilized in Xanthomonas-Arabidopsis studies is the pathovar campestris. Various strains of this pathovar have demonstrated typical race-cultivar specificity indicative of the presence of a gene-for-gene relationship between the host and the pathogen. Thus, due to the abundance of literature with X. campestris pv campestris (Xcc), a majority of this chapter will be devoted to the interactions of Arabidopsis with various strains of Xcc.

Xcc and Black Rot of Crucifers

Xcc is the causal agent of black rot of crucifers (for review see Williams 1980). Xcc has a broad host range that includes a majority of members of the Cruciferae family. Numerous agronomically important species of crucifers including broccoli, brussel sprouts, cabbage, cauliflower, radish, and turnip are susceptible to Xcc. Xcc can also infect weed species within the Cruciferae (Schaad and Dianese 1981) making weed reservoirs of Xcc a factor in the control of black rot in crucifer production fields.

Xcc typically infects host plants through natural openings such as hydathodes, stomates, or wounds (for review see Williams 1980). Once the bacterium is in the vascular system, it can become systemic resulting in blackened veins and thus the name “black rot”. Under favorable environmental conditions, the pathogen can become seed-borne (for review see Williams 1980). For commercial crucifers, black rot is a significant issue. The fact that the pathogen can be seed-borne has led to seed certification programs in developed countries. The certification process adds extensive cost and effort for commercial seed production.

Several lines of evidence have firmly established Arabidopsis as a natural host of Xcc. First, classic black rot symptoms have been reported following infection of Arabidopsis with specific Xcc strains (Simpson and Johnson 1990; Tsuji et al., 1991). Second, systemic infection occurs in highly susceptible accessions following challenge with Xcc (Figure 1; Buell and Somerville 1997). Third, a preference for hydathode infection, rather than stomatal infection, was observed in Arabidopsis leaves which is consistent with the entry route favored by Xcc in other cruciferous hosts (Hugouvieux et al., 1998). Fourth, infections of Arabidopsis by Xcc do occur in nature as reported by Tsuji and Somerville (1992).

Pathogenicity Mechanisms of Xanthomonas species

There have been several excellent reviews on pathogenicity and virulence mechanisms in gram-negative plant pathogenic bacteria and the reader is referred to these articles for more in-depth discussions (Collmer 1998; Staskawicz et al., 2001). With respect to Xcc pathogenicity on non-Arabidopsis crucifers, there are a number of publications by M. J. Daniels and colleagues and the reader is referred to these for a summary of Xcc virulence mechanisms (Daniels et al., 1993; Dow and Daniels 1994). In brief, Xcc employs a vast array of degradative and regulatory mechanisms to parasitize its host. Extracellular polysaccharide production, cell wall degrading enzymes, proteases, as well as genes that encode for secretion of these products are involved in virulence on cruciferous hosts. In addition to these structural components, regulatory components of pathogenicity and virulence have been identified in Xanthomonas (for review see Daniels et al., 1993; Dow and Daniels 1994).

Although there is a substantial amount of information currently available on pathogenicity mechanisms in Xanthomonas, the level of information is about to increase exponentially. The first genome of a plant pathogen completely sequenced was Xylella fastidosa (Simpson et al., 2000) which is the causal agent of citrus variegated chlorosis and is closely related to Xanthomonas. Not surprisingly, several components of virulence and pathogenicity in Xcc have orthologs in X. fastidosa, including genes involved in xanthan gum (extracellular polysaccharide) production, regulation of pathogenicity factors, and the type II secretion system that is necessary for export of degradative enzymes (Dow and Daniels 2000; Simpson et al., 2000).

Even more exciting is the new genome sequencing projects focused on Xanthomonas species. X. axononpodis pv citri, the causal agent of citrus canker, is being sequenced by the Organization for Nucleotide Sequencing and Analysis (ONSA), a consortium of laboratories in Brazil (Kamoun and Hogenhout 2001;  http://genoma4.iq.usp.br/Xanthomonas/). This pathogen is closely related to Xcc and major insights into the virulence mechanisms of Xanthomonas will be revealed from this project. The other Xanthomonas genome project is focused on Xcc (Kamoun and Hogenhout 2001;  http://genoma.fcav.unesp.br/xc-campestris/home/xc_menu.html). In addition, the genome of Ralstonia solanacearum (Kamoun and Hogenhout 2001;  http://www.genoscope.cns.fr/externe/English/Projets/Projet_Y/Y.html) and Pseudomonas syringae pv tomato ( http://www.tigr.org;  http://ppi.cornell.edu/), both pathogens of Arabidopsis, are the focus of genome sequencing efforts and these genomic sequences will provide additional resources for the identification of pathogenicity mechanisms in Xanthomonas.

Genetic Basis for Resistance to Xanthomonas

Responses in Arabidopsis to Xanthomonas infections are dependent on genetic factors present in the host and in the pathogen. Several types of resistance responses have been documented in Arabidopsis-Xanthomonas interactions. These include tolerance, resistance mediated through a hypersensitive response (HR) and non-HR-mediated resistance. Genes involved in the resistance response can be classified into three classes: (1) those involved in the recognition of the pathogen (also known as R genes), (2) genes involved in signal transduction events, and (3) genes involved in the suppression of pathogen growth and development (defense response genes). Currently, five genes have been identified genetically that are hypothesized to be involved in recognition of the pathogen (Table 2; Tsuji et al., 1991; Buell and Somerville 1997; Godard et al., 2000). These have been termed RXC for reaction to Xanthomonas campestris and are named RXC1-5. Additional genes have been identified in both the signal transduction pathway and in the suppression of pathogen growth/development and are discussed in more detail below (see also Table 3).

At the present, only a single corresponding gene for avirulence has been reported from a crucifer-infecting Xanthomonas isolate. This avirulence gene, avrXca, was isolated from X. c. raphani and confers incompatibility in a large number of Arabidopsis accessions (Parker et al., 1993). avrXca encodes a large protein (∼67 kDa) that is possibly secreted from the bacterium and the upstream region of avrXca contains a putative hrp box suggesting avrXca is controlled by the hrp regulatory system. The hrp pathway is an essential component of pathogenicity and virulence in bacterial plant pathogens and the reader is referred to these reviews for more information (Alfano and Collmer 1997; Cornelis and Van Gijsegem 2000). The lack of identification of corresponding avirulence gene(s) in Xcc isolates may reflect either technical difficulties in screening for such a gene or a lack of the gene in this pathogen. Genome sequencing efforts (see above) should provide valuable insight into this unresolved issue.

Tolerance to Xcc

Using the 2D520 strain of Xcc, Tsuji et al., (1991) reported tolerance to the pathogen in the Columbia accession (Col-0). While the Pr-0 accession developed chlorosis and necrosis following infiltration with Xcc2D520, Col-0 remained asymptomatic. As the in planta bacterial levels were indistinguishable between these two accessions, the response in Col-0 was termed tolerance as limited bacterial growth was supported in Col-0 without the development of symptoms. Thus, an uncoupling of symptom formation and pathogen growth is seen in the interaction of Col-0 and Pr-0 with Xcc 2D520. Genetic mapping efforts indicated that a single dominant gene, termed RXC1 (Tsuji et al., 1991), confers tolerance. RXC1 was been mapped to the lower arm of chromosome 2 (Buell and Somerville 1997).

The biochemical and physiological mechanism by which Col-0 is able to tolerate 2D520 is not well understood. This tolerance does not involve the synthesis of the phytoalexin, camalexin (Tsuji et al., 1992). At the molecular level, examination of the mRNA accumulation levels of several genes involved in defense responses did not reveal an accumulation of mRNA unique to Col-0 and thus no genes could be correlated specifically with tolerance (Buell and Somerville 1995). Although an accumulation of pathogenesis-related protein 1 (PR-1) mRNA was observed, it was expressed at high levels in both Col-0 and Pr-0.

Non-HR Resistance to Xcc

Previous work with Xcc2D520 utilized the differential accessions Col-0 and Pr-0. A further survey of Arabidopsis accessions revealed additional susceptible accessions. Specifically, the accession, Landsberg erecta (Ler), is highly susceptible to Xcc2D520 (Tsuji and Somerville 1988; Buell and Somerville 1997). Inoculation of Ler results in severe chlorosis and necrosis which can spread systemically resulting in necrosis of the entire plant (Figure 1; Buell and Somerville 1997). A qualitative difference in the susceptible response is apparent between Pr-0 and Ler. Whereas Pr-0 lesions are primarily chlorotic in nature, Ler lesions are more necrotic with darkening of the vascular tissue that is absent from Pr-0 lesions (Figure 2). In planta bacterial levels in Ler are 10-100- fold higher than in Col-0 and in Pr-0, revealing a suppression of bacterial growth in Col-0 and Pr-0 in comparison to Ler (Buell and Somerville 1995, 1997). Thus, Col-0 exhibits two responses to Xcc2D520: tolerance to limited bacterial growth (as compared to the susceptible accession Pr-0) and resistance without a HR (when compared to the highly susceptible accession Ler).

Examination of the genetic basis for resistance to the 2D520 isolate indicated three loci, RXC2, RXC3, and RXC4, were involved in resistance (Buell and Somerville 1997). The major locus for resistance, RXC2, behaved as a single dominant gene whereas the other two loci, RXC3 and RXC4, functioned in a digenic manner. These loci have been placed on the genetic map and map to chromosome 5 (RXC2, RXC3) and chromosome 2 (RXC4) (Buell and Somerville 1997). Although RXC2 and RXC3 map to distinct locations on chromosome 5, RXC4 maps to the same region of chromosome 2 as RXC1. Due to limited recombination events, it is not clear whether RXC1 and RXC4 are the same gene or simply map to similar regions of the chromosome.

Hypersensitive Response: Interactions with Xcc750

The Xcc750 isolate has been shown to induce a HR-like response on resistant accessions of Arabidopsis such as Col-0 while causing chlorosis on susceptible accessions such as Oy-0 (Aufsatz and Grimm 1994). The HR observed on Col-0 is coupled with a lack of significant bacterial growth, consistent with the HR observed in Arabidopsis with other pathogens such as Pseudomonas. Resistance in Arabidopsis is associated with high basal expression and an induction of expression of a low molecular weight protein termed ECS1 (formerly named CXc750; Aufsatz and Grimm 1994). Subsequent work using transgenic plants and polyclonal antibodies generated to ECS1 revealed it is associated with the cell wall (Aufsatz et al., 1998). Although ECS1 had been tightly correlated with the resistance phenotype, definitive genetic and transgenic experiments have revealed ECS1 is not a resistance gene for Xcc750 but instead is linked to a locus involved in resistance to Xcc750.

Hypersensitive Response: Interactions with Xcc147

The response of Arabidopsis to the HR-inducing strain 147 of Xcc has been described in a series of papers by the Roby laboratory and other researchers. Xcc147 induces a HR on resistant accessions of Arabidopsis such as Col-0 (Lummerzheim et al., 1993). At the molecular level, a correlation between mRNA accumulation of two defense genes, phenylalanine ammonia lyase (PAL) and β-1,3-glucanase, and the HR was observed (Lummerzheim et al., 1993). Lummerzheim et al. (1993) also examined transcript accumulation of a basic class I chitinase and an ascorbate peroxidase and although they were up-regulated upon infection with Xcc147, they were also expressed in the compatible interaction with Xcc8004 revealing a lack of specificity with the HR. There are four classes of chitinase (I to IV) and Gerhardt et al. (1997) were able to demonstrate that an extracellular class IV chitinase was expressed in Arabidopsis leaves following challenge with Xcc147. Unfortunately, class IV chitinase expression was not examined in leaves challenged with a compatible Xcc isolate and no conclusion can be made regarding the specificity of class IV chitinase expression in the HR. In healthy plants, the class IV chitinase was only expressed in siliques, suggesting a specific developmental pattern for this gene.

While the above studies with Xcc147 were performed using whole plant studies, a number of new genes involved in the HR to Xcc147 have been identified using cell suspension cultures. A cDNA clone encoding a putative sulfotransferase was identified from a library constructed from pathogen-challenged cell suspension cultures. The sulfotransferase, RaR047, is similar to flavonol sulfotransferases from Flaveria species (Lacomme and Roby 1996). RaR047 is developmentally regulated as mRNA could only be detected in cell cultures and in developing aerial tissues (Lacomme and Roby 1996). Expression of RaR047 was strongly induced in young seedlings by treatment with several elicitors of defense responses, including salicylic acid and jasmonic acid. The induction of RaR047 expression by elicitors is consistent with the induction of expression observed with pathogen treatment of Arabidopsis leaves (Lacomme and Roby 1996). Inoculation with either Xcc147 or an incompatible strain of Pseudomonas syringae pv maculicola (M2) resulted in accumulation of RaR047 mRNA. Compatible strains of Xcc (Xcc8004) and P. s. maculicola (M4) were also able to induce accumulation of RaR047 although the extent of accumulation was substantially less than that in incompatible interactions. Although there is a correlation of RaR407 expression with resistance (Lacomme and Roby 1996), the function of the encoded protein is unknown. It is speculated that RaR047 may function in synthesis of a molecule that is involved in signal transduction or that RaR047 functions directly in the suppression of pathogen growth (Lacomme and Roby 1996). Definitive biochemical studies will establish an enzymatic function for RaR047 and allow placement of this sulfotransferase in the defense response pathway.

In addition to sulfotransferase, cinnamoyl-CoA reductase, an enzyme involved in lignification, is associated with the development of the HR to Xcc147. Two cinnamoyl-CoA reductase genes, AtCCR1 and AtCCR2, exhibit different substrate specificities and do not exhibit coordinate regulation during development and pathogen challenge (Lauvergeat et al., 2001). AtCCR1 is expressed throughout normal development and is expressed more in highly lignified tissues such as stems in comparison to leaf tissue. In contrast, AtCCR2 is weakly expressed in developing tissue. Following challenge with Xcc147, AtCCR2 and not AtCCR1, was highly upregulated. AtCCR2 expression was correlated with the HR as AtCCR2 expression was not detectable in compatible tissues. AtCCR2 was also inducible by treatment with salicylic acid, further supporting its role in defense responses. Although these two genes share substantial similarity (81.6 % identity at the amino acid level) they clearly have distinct expression profiles in planta. With the completion of the Arabidopsis Genome Initiative (2000), it would be interesting to examine the regulatory regions of these genes for promoter sequences that may reflect the differential expression patterns.

An additional set of genes involved in the HR to Xcc147 was described by Lacomme and Roby in 1999. A total of 27 cDNA clones (Athsr) was identified by differential screening of a cDNA library that was constructed from cell suspension cells challenged with Xcc147. The 27 clones were then grouped into “cDNA clone families” based on cross-hybridization and sequencing results. The Athsr2 cDNA family was the most heavily represented family with 16 clones identified. The Athsr2 cDNA family encodes voltage-dependent anion channel proteins (Lacomme and Roby 1999) that are localized in the mitochondrion and function to transport small molecules across the mitochondrial membrane. Membrane integrity and control over solute movement across the membrane are central components in apoptotic cell death and voltage-dependent anion channel proteins have been well studied in mammalian apoptosis (for review see Green and Reed 1998; Boya et al., 2001). The Athsr3 cDNA family, represented by 6 clones from the screening, encodes an alterative oxidase (Lacomme and Roby 1999). In tobacco, Chivasa et al. (1997) demonstrated that alternative oxidase was a component of the signal transduction pathway that leads to the HR following challenge with tobacco mosaic virus. While Athsr2 and Athsr3 were associated with the mitochondrion, Athsr4 (represented by a single clone) encodes a protein with similarity to a Rab GDP-dissociation inhibitor protein. Rab proteins are involved in membrane/vesicle trafficking and Rab GDP-dissociation inhibitor proteins are intimately involved in the regulation of Rab proteins and ultimately in the regulation of membrane/vesicle trafficking (for a review on Rab proteins and their regulation see Stenmark and Olkkonen, 2001). The other 3 cDNA families (Athsr5, 6, 7), also represented by a single clone, did not have significant similarity to any entries in the database (Lacomme and Roby 1999).

The expression patterns of the Athsr2-7 cDNAs were assessed in cell suspension cultures challenged with differential isolates of Xanthomonas: Xcc147 (avirulent), Xcc8004 (virulent), and Xcc 8B2 (control). All of the cDNAs were exclusively or preferentially expressed in cells challenged with Xcc147, confirming the success of the differential screening and suggesting a role for these genes in the HR (Lacomme and Roby 1999). Further research using a biochemical and a functional genomics approach will be essential for defining the roles of these genes in the HR.

The last gene identified from Xcc147-challenged cell suspension cells is the AtMYB30 gene which encodes an orthologue of the myb oncogene (Daniel et al., 1999; Lacomme and Roby 1999). MYB proteins are transcription factors and are involved in many cellular functions including plant defense responses (Yang and Klessig 1996). AtMYB30 encodes a 323 amino acid protein and consistent with other MYB proteins contains MYB repeats in its N-terminus (Daniel et al., 1999). At the developmental level, AtMYB30 is expressed at extremely low levels and is detectable only in developing seedlings. Expression of AtMYB30 was induced by challenge with avirulent Xcc147 cells and avirulent P. syringae strains. Expression was correlated with phenotypic expression of the HR, as AtMYB30 expression was not detectable in the susceptible accession Sf-2 or with virulent strains of Xcc or P. syringae. Expression of AtMYB30 was also examined in various lesion-mimic mutants (lsd) which develop HR-like lesions in the absence of pathogens. In three lsd mutants (lsd3, lsd4, lsd5), a positive correlation between AtMYB30 expression and lesion presence was found (Daniel et al., 1999), further supporting the hypothesis that AtMYB30 is a positive regulator of cell death.

Using differential display, Cordeiro et al., (1998) were able to identify two novel genes involved in the early stages of the HR of Arabidopsis to Xcc147. One gene, ap3.3a, encodes a protein with similarity to centrin, a cytoskeletal protein. Similar to the other centrins, the Arabidopsis ap3.3a protein contains four regions that are hypothesized to be involved in Ca²⁺-binding (Cordeiro et al., 1998). Expression of ap3.3a was measured in compatible and incompatible interactions with Xcc. A rapid (1 hr) up-regulation of ap3.3a was observed in the incompatible interaction with Xcc147 whereas accumulation of ap3.3a was not observed in the compatible interaction until 1–2 dpi. A second gene identified through differential display is the ap4.3a gene which encodes a protein with multiple kinase domains. ap4.3a is rapidly induced (15 min) upon challenge with Xcc, although specificity of the induction was not apparent between compatible and incompatible Xcc strains.

Mutational Approaches to Characterize Signal Transduction and Defense Pathway Components in Arabidopsis-Xanthomonas Interactions

In a complementary approach to differential screening of cDNA libraries, Roby and colleagues identified a mutant of Arabidopsis that is deficient in manifestation of the HR following challenge with Xcc147. In a screen of 20,700 M₂ plants, a single recessive mutant (hxc-2) was identified that is unable to mount a full HR (Godard et al., 2000). Inoculation of hxc-2 plants results initially in necrosis (48 hr) at the infection site which is followed by a chlorotic halo and ultimately, spreading chlorosis. In the hxc-2 plants, bacterial growth is unrestricted and is indistinguishable from levels in the susceptible accession Sf-2. The hxc-2 mutant is able to mount an effective HR against avirulent strains of P. syringae, suggesting that the wild-type HXC2 locus is specific for Xcc-mediated HR. At the biochemical level, the hxc-2 mutants retain full responsiveness to salicylic acid which is an inducer of systemic acquired resistance (Godard et al., 2000). However, accumulation of salicylic acid is impaired following challenge with Xcc147. At the molecular level, expression levels of several defense genes (PAL-1, PR-1, PDF-1.2) are altered in hxc-2, suggesting a role for hxc-2 in the signal transduction pathway leading to resistance. Genetic mapping experiments place HXC2 on chromosome 3 near the Major Recognition Gene Complex MRC-F (Godard et al., 2000). Through allelism tests, it was demonstrated that HXC2 is not the determinant of specificity to Xcc147 and that a second gene, RXC5, governs recognition of the pathogen.

Ethylene has been implicated in numerous plant responses, including the contrasting responses of defense in incompatible interactions and symptom formation in compatible interactions (Abeles et al., 1992). The availability of mutants in ethylene-associated developmental processes has provided valuable reagents to dissect the role of ethylene in plant-pathogen interactions (for a recent review on the ethylene response pathway see Bleecker and Kende 2000). Inoculation of the ethylene insensitive2 (ein2) perception mutant with a virulent isolate of Xcc resulted in plants with reduced symptoms (Bent et al., 1992), suggesting EIN2 is involved in symptom formation. EIN2 has been cloned and encodes a novel protein (Alonso et al., 1999). However, although EIN2 has been placed in the ethylene response pathway through epistasis tests, its function at the biochemical and molecular level is still unknown (Bleecker and Kende 2000). Another ethylene-related mutant, hookless1 (hls1-1), is unable to form an apical hook during the triple response in germinating seedlings (Guzman and Ecker 1990). Infection of hls1-1 with Xcc2D520 results in disease symptoms and an increase in bacterial growth in comparison to the asymptomatic parental accession (Col-0), suggesting HLS1 functions in resistance, not tolerance, to Xcc2D520 (Buell 1998; Buell, unpublished). The HLS1 gene encodes an acetyltransferase-like protein yet a specific enzymatic function for the protein has not been determined either in apical hook formation or disease resistance (Lehman et al., 1988). A second mutant in apical hook formation, constitutive photomorphogenic 2 (cop2; Hou et al., 1993), is also suppressed in resistance to Xcc2D520 (Buell, C.R., unpublished) consistent with the phenotype observed in the hls1-1 apical hook mutant. COP2 has not been cloned but with the recent completion of the Arabidopsis genome (Arabidopsis Genome Initiative, 2000), this can be accomplished in the near future. Continued work focused on resolving the biochemical function of these proteins will be valuable to dissecting their role in pathogen responses.