Genome-wide study of C2H2 zinc finger gene family in Medicago truncatula

Abstract

Background: C2H2 zinc finger proteins (C2H2 ZFPs) play vital roles in shaping many aspects of plant growth and adaptation to the environment. Plant genomes harbor hundreds of C2H2 ZFPs, which compose one of the most important and largest transcription factor families in higher plants. Although the C2H2 ZFP gene family has been reported in several plant species, it has not been described in the model leguminous species Medicago truncatula.

Results: In this study, we identified 218 C2H2 type ZFPs with 337 individual C2H2 motifs in M. truncatula. We showed that the high rate of local gene duplication has significantly contributed to the expansion of the C2H2 gene family in M. truncatula. The identified ZFPs exhibit high variation in motif arrangement and expression pattern, suggesting that the short C2H2 zinc finger motif has been adopted as a scaffold by numerous transcription factors with different functions to recognize cis-elements. By analyzing the public expression datasets and quantitative RT-PCR (qRT-PCR), we identified several C2H2 ZFPs that are specifically expressed in certain tissues, such as the nodule, seed, and flower.

Conclusion: Our genome-wide work revealed an expanded C2H2 ZFP gene family in an important legume M. truncatula, and provides new insights into the diversification and expansion of C2H2 ZFPs in higher plants.

Keywords: C2H2; EAR motif; Expression; Gene family; Local gene duplication; Zinc finger.

Fig. 1

MapChart diagram illustrating the distribution of C2H2 ZFPs in the M. truncatula genome. The black lines within the chromosomes indicate C2H2 ZFPs. The line linking two C2H2 ZFPs represents a pair of C2H2 ZFPs resulting from WGD. Different colors of the line and gene ID indicate different pairs of C2H2 ZFPs from WGD

Fig. 2

Phylogenetic analysis and motif arrangement of TFIIIA and TRM1 homologs. The top part of (a) and (b) were the unrooted maximum-likelihood tree of TFIIIA and TRM1 homologs, respectively. The blue and green branches indicated monocot and dicot plants respectively. The bottom part of (a) and (b) are motif arrangements of TFIIIA and TRM1, respectively. The black line represents the protein sequence. The black rectangle indicates the C2H2 motif, and the red rectangle indicates the EAR motif

Fig. 3

IDD-type C2H2 motifs. a Alignment of IDD-type C2H2 motifs from M. truncatula. b Signature of IDD-type C2H2 motifs

Fig. 4

Representative single C2H2 ZFP in M. truncatula. a Single C2H2 ZFP without the EAR motif. b IDD homologs. c PALM1 and RSD type C2H2 ZFPs. The black line represents the protein sequence. The black and red rectangles indicate C2H2 and EAR motifs, respectively

Fig. 5

ZFPs with an array of C2H2 motifs in M. truncatula. The black line represents the protein sequence and the black rectangle indicates C2H2 motifs

Fig. 6

Tissue-specific expression analysis of C2H2 ZFPs in M. truncatula. a - c The transcript abundances of the C2H2 ZFPs showed by heatmaps generated from the M. truncatula Gene Expression Atlas data. a shows the genes with low transcript abundances, (b) shows the genes with medium transcript abundances, and (c) shows the genes with high transcript abundances. d The qRT-PCR result of six selected C2H2 genes. Shown are means ± standard deviations for three biological replicates and three technical replicates of each biological replicate. The relative expression levels of tested genes were normalized by the geometric mean of three endogenous control genes. ‘N.A.’ indicated undetectable expression. The lowercase letters above the bar indicate significant differences (P < 0.05) using ANOVA and Tukey’s test among samples

Fig. 7

Confirmation of C2H2 ZFPs specifically expressed in M. truncatula symbiotic nodules. The qRT-PCR result of six selected C2H2 genes. Shown are means ± standard deviations for three biological replicates and three technical replicates of each biological replicate. The relative expression levels of tested genes were normalized by the geometric mean of three endogenous control genes. ‘N.A.’ indicated undetectable expression. The lowercase letters above the bar indicate significant differences (P < 0.05) using ANOVA and Tukey’s test among samples