Genome-wide association studies meta-analysis uncovers NOJO and SGS3 novel genes involved in Arabidopsis thaliana primary root development and plasticity

Abstract

Background: Arabidopsis thaliana primary root growth has become a model for evo-devo studies due to its simplicity and facility to record cell proliferation and differentiation. To identify new genetic components relevant to primary root growth, we used a Genome-Wide Association Studies (GWAS) meta-analysis approach using data published in the last decade. In this work, we performed intra and inter-studies analyses to discover new genetic components that could participate in primary root growth.

Methods and results: We used 639 accessions from nine different studies under control conditions and performed different GWAS tests. We found that primary root growth changes were associated with 41 genes, of which six (14.6%) have been previously described as inhibitors or promoters of primary root growth. The knockdown lines of two genes, Suppressor of Gene Silencing (SGS3), involved in tasiRNA processing, and a gene with a Sterile Alpha Motif (SAM) motif named NOJOCH MOOTS (NOJO), confirmed their role as repressors of primary root growth, none has been shown to participate in this developmental process before.

Conclusions: In summary, our GWAS analysis of different available studies identified new genes that participate in primary root growth; two of them were identified as repressors of primary root growth.

Keywords: Arabidopsis thaliana; NOJO; SGS3; Accessions; GWAS; Primary root growth.

Fig. 1

Comparison of the PR length between studies and the frequency of accessions used. (A). The box plot represents the distribution of the mean PR length of all accessions used in the different studies, the boxes indicate the first and third quartiles, the vertical line is the median, and the whiskers go from each quartile to the maximum or minimum value. The letter corresponds to each study, and the number means the DAS evaluated. Lachowiec et al., 2015 (A7) [21], Julkowska et al., 2017 (B8) [10], Ristova et al., 2018 (C10) [13], Li et al., 2019 (F3-F8, F10, F13) [16]; Justamante et al., 2019 (H6) [12], Deolu-Ajayi et al., 2019 (I6-I9) [11] or DAG: Bouain, et al., 2018 (D1-D7) [14], Bouain, et al., 2019 (E4-E5) [15], Ogura et al., 2019 (G5) [23], (B). Frequency of accessions shared between studies. (C). Primary root length of KIN-0, the only accession shared between all the studies and seedling ages

Fig. 2

Correlation coefficients among the studies and the different seedling ages used in each one. The abbreviation of each study is depicted in S1 Table. On the right side of the correlogram, the legend colour shows the correlation coefficients and the corresponding colours; positive correlations are displayed in blue and negative correlations in red colour. The black cross in each cell depicts no association. The letters on the left side of the figure correspond to the different studies: Lachowiec et al., 2015 (A7); Julkowska et al., 2017 (B8); Ristova et al., 2018 (C10); Bouain et al., 2018 (D1-D7). Bouain, et al., 2019 (E3-E5), Li et al., 019 (F3-F8, F10, F13); Ogura et al., 2019 (G5), Justamante et al., 2019 (H6), Deolu-Ajayi et al., 2019 (I6-I9).

Fig. 3

SNPs obtained from individual GWAS and SNPs associated to the genes involved in PR length (A). Manhattan plot using the top 0.1% SNPs (n = 205 SNPs for each study, 5,535 SNPs in total) and their association [-log10(P-value)]. The 10% false discovery rate (FDR) threshold after the Benjamini-Hochberg correction is plotted as a red horizontal line. (B). 10-kbp region surrounding SNPs whose associated genes show significant changes in PR length

Fig. 4

NOJO and SGS3 are novel genes involved in PR length. (A) PR length at 12 DAS of two alleles of Col-0, SGS3 and NOJO (n = 90 seedlings used per allele). Significant value is indicated (***) p < 0.001, (*) p < 0.05 using ANOVA one-way followed by Tukey’s post-hoc tests (B) Representative seedling at 12 DAS of two alleles of NOJO and SGS3 that were grown in the same plate with Col-0. Scale bar = 1 cm. (C) Relative expression of SAM domain-containing protein (NOJO) and SGS3 in two T-DNA lines. Data represent the means ± SD (n = 9). For nojo lines and WT, ANOVA one-way followed by Tukey’s post-hoc tests were performed, for sgs3-13 and WT, unpaired two-tail Student t test was used (***) p < 0.001)

Fig. 5

SNPs observed in SGS3 and NOJO from accessions with large and short PR length (A) The red rectangles indicate the SNPs found in SGS3 and NOJO genes and each nucleotide displays different colours. In SGS3, the main changes are kept in the first and second exon and first intron in accessions with a short PR. In the gene that encodes a SAM domain-containing protein, the SNPs are detected in the distal part of the single exon in some accessions with long PR length and in the proximal part of the exon for accessions with short PR. (B) SGS3 protein. The amino acid change is indicated in the XS domain. (C) NOJO protein. The amino acid changes were found in the SAM domain and in a sequence without a described domain. In B and C, the main domains are depicted with different colours, and regions with uncommon domains are shown in grey