Genome-Wide Identification of MAPKK and MAPKKK Gene Families in Tomato and Transcriptional Profiling Analysis during Development and Stress Response

<div><p>Mitogen-activated protein kinase (MAPK) cascades have important functions in plant growth, development, and response to various stresses. The MAPKK and MAPKKK gene families in tomato have never been systematically analyzed. In this study, we performed a genome-wide analysis of the MAPKK and MAPKKK gene families in tomato and identified 5 MAPKK genes and 89 MAPKKK genes. Phylogenetic analyses of the MAPKK and MAPKKK gene families showed that all the MAPKK genes formed four groups (groups A, B, C, and D), whereas all the MAPKKK genes were classified into three subfamilies, namely, MEKK, RAF, and ZIK. Evolutionary analysis showed that whole genome or chromosomal segment duplications were the main factors responsible for the expansion of the MAPKK and MAPKKK gene families in tomato. Quantitative real-time RT-PCR analysis showed that the majority of MAPKK and MAPKKK genes were expressed in all tested organs with considerable differences in transcript levels indicating that they might be constitutively expressed. However, the expression level of most of these genes changed significantly under heat, cold, drought, salt, and <i>Pseudomonas syringae</i> treatment. Furthermore, their expression levels exhibited significant changes in response to salicylic acid and indole-3-acetic acid treatment, implying that these genes might have important roles in the plant hormone network. Our comparative analysis of the MAPKK and MAPKKK families would improve our understanding of the evolution and functional characterization of MAPK cascades in tomato.</p></div

Characteristics of MAPK kinase (MAPKKs) from <i>Solanum lycopersicum</i>.

<p>Characteristics of MAPK kinase (MAPKKs) from <i>Solanum lycopersicum</i>.</p

Additional file 2: Figure S1. of Identification and expression profiling of microRNAs involved in the stigma exsertion under high-temperature stress in tomato

Nucleotide bias at each position of identified miRNAs. Y-axis: frequency of A/U/G/C; X-axis: position in miRNAs. Figure S2. Real-time quantitative PCR validation of nine heat-responsive miRNAs in stamen (a) and pistil (b), respectively. Y-axis shows the log2 ratio of miRNAs expression in HS versus CK. SnoU6 was used as the internal control. Each bar represents the mean ± SE of triplicated assays. Figure S3. Distribution of tRNAs, snoRNAs, and snRNAs in stamen and pistil libraries. Y-axis: frequency of each category of small RNAs. Figure S4. qRT-PCR analysis of the expression of SlLAC4 in stamen (a) and pistil (b) under heat-stress treatment. SlUbi3 was used as the internal control. Each bar represents the mean ± SE of triplicated assays. (ZIP 508 kb

Additional file 1: Table S1. of Identification and expression profiling of microRNAs involved in the stigma exsertion under high-temperature stress in tomato

Evaluation of the correction of the reduplicate biological samples. Table S2. Known miRNAs identified from stamen and pistil libraries. Table S3. Differentially expressed known and novel miRNAs between stamen and pistil CK-2d libraries. Table S4. Novel miRNAs identified from stamen and pistil libraries. Table S5. Differentially expressed miRNAs shared between stamen and pistil under heat stress condition. Table S6. Stamen specific differentially expressed miRNAs. Table S7. Pistil specific differentially expressed miRNAs. Table S8. Target genes of differentially expressed miRNAs in stamen. Table S9. Target genes of differentially expressed miRNAs in pistil. Table S10. List of primers used for qRT-PCR analysis. Table S11. List of primers used for RLM-5′ RACE analysis. (XLS 299 kb

Expression profiles of ZIK subfamily genes with exogenous IAA (left) and SA (right) treatments.

<p>Expression profiles of ZIK subfamily genes with exogenous IAA (left) and SA (right) treatments.</p

Chromosomal distribution of SlMAPKKs and SlMAPKKKs genes in tomato genome.

<p>The names of each tandem duplicated gene cluster of the two families were indicated with black rectangles. The triangles indicate the upward or downward direction of transcription.</p

Characteristics of MAPK kinase kinase (MAPKKKs) from <i>S. lycopersicum</i>.

<p>Characteristics of MAPK kinase kinase (MAPKKKs) from <i>S. lycopersicum</i>.</p

Alignment of SlMAPKK proteins in tomato.

<p>The highlighted part shows the conserved signature motif obtained with the ClustalX program.</p

The numbers of SlMAPK, SlMAPKK, and SlMAPKKK in <i>Arabidopsis</i>, rice, tomato, and maize.

<p>The numbers of SlMAPK, SlMAPKK, and SlMAPKKK in <i>Arabidopsis</i>, rice, tomato, and maize.</p

Phylogenetic analysis (left), domain organization (middle) and exon-intron structures (right) of tomato MAPKKs.

<p>The unrooted phylogenetic tree was generated using MEGA4.1 program by the neighbor-joining method. Bootstrap supports from 1000 replicates are indicated at each branch. The gene names of each subfamily are indicated with the same color. The domain organizations are analyzed by scanning of the protein sequences for the presence of known motifs and domains using PlantsP. The exon-intron organization of corresponding SlMAPKK genes is represented by yellow boxes and lines, respectively.</p