Global transcriptome analysis of constitutive resistance to the white pine weevil in spruce

Abstract

Constitutive defense mechanisms are critical to the understanding of defense mechanisms in conifers because they constitute the first barrier to attacks by insect pests. In interior spruce, trees that are putatively resistant and susceptible to attacks by white pine weevil (Pissodes strobi) typically exhibit constitutive differences in traits such as resin duct size and number, bark thickness, and terpene content. To improve our knowledge of their genetic basis, we compared globally the constitutive expression levels of 17,825 genes between 20 putatively resistant and 20 putatively susceptible interior spruce trees from the British Columbia tree improvement program. We identified 54 upregulated and 137 downregulated genes in resistant phenotypes, relative to susceptible phenotypes, with a maximum fold change of 2.24 and 3.91, respectively. We found a puzzling increase of resistance by downregulated genes, as one would think that "procuring armaments" is the best defense. Also, although terpenes and phenolic compounds play an important role in conifer defense, we found few of these genes to be differentially expressed. We found 15 putative small heat-shock proteins (sHSP) and several other stress-related proteins to be downregulated in resistant trees. Downregulated putative sHSP belong to several sHSP classes and represented 58% of all tested putative sHSP. These proteins are well known to be involved in plant response to various kinds of abiotic stress; however, their role in constitutive resistance is not yet understood. The lack of correspondence between transcriptome profile clusters and phenotype classifications suggests that weevil resistance in spruce is a complex trait.

FIG. 1.—

Parent trees' origin within the Prince George area. The color scale (S–R) indicates the level of resistance of the trees, from highly susceptible to highly resistant, blue to red, respectively. Filled circles represent origin of the tree families used in the present microarray study. Open circles represent the origin of tree families not used in the microarray study but used for the resistance ranking (map layers from MapPlace Web site http://www.empr.gov.bc.ca/MINING/GEOSCIENCE/MAPPLACE/Pages/default.aspx).

FIG. 2.—

Percentage of damaged trees among progenies. Progenies are ordered from the least damaged to the most damaged. Resistant and susceptible families are located on the left and on the right, respectively. White bars and black bars show selected families for the present study.

FIG. 3.—

Smoothed densities color representation of volcano plot, showing the differential expression levels of 18,725 genes between resistant and susceptible trees. Significant downregulated and upregulated genes are shown in blue and red, respectively.

FIG. 4.—

Heat map of the 191 significantly differently expressed genes between susceptible and resistant trees to the white pine weevil. Blue and red squares at the top of the heat map indicate susceptible and resistant trees, respectively. Tree labels are indicated at the bottom as follow: the tree phenotype (R = resistant, S = susceptible), the family rank in progeny tests for resistance (1 = the most resistant; 179 = the most susceptible), and then the family number.

FIG. 5.—

Significantly overrepresented GO terms of genes among significant upregulated or downregulated genes between susceptible and resistant trees, respectively. Fisher’s exact tests with multiple testing corrections were performed using Blast2GO software. Only GO categories with false detection rate lower than 0.05 are shown.

FIG. 6.—

Phylogenetic analysis of spruce sHSP. The tree was derived by Neighbor-Joining method with bootstrap analysis (1,000 replicates) from alignment of amino acid sequences of sHSP of rice, Arabidopsis, and poplar. Bootstrap values higher than 50% are shown next to the branches. Phylogenetic analyses were conducted in MEGA4. EST clones ID of Picea are indicated in bold and underlined. Downregulated sHSP are indicated by a closed black circle.