Regulation of Root Angle and Gravitropism

Abstract

Regulation of plant root angle is critical for obtaining nutrients and water and is an important trait for plant breeding. A plant's final, long-term root angle is the net result of a complex series of decisions made by a root tip in response to changes in nutrient availability, impediments, the gravity vector and other stimuli. When a root tip is displaced from the gravity vector, the short-term process of gravitropism results in rapid reorientation of the root toward the vertical. Here, we explore both short- and long-term regulation of root growth angle, using natural variation in tomato to identify shared and separate genetic features of the two responses. Mapping of expression quantitative trait loci mapping and leveraging natural variation between and within species including Arabidopsis suggest a role for PURPLE ACID PHOSPHATASE 27 and CELL DIVISION CYCLE 73 in determining root angle.

Keywords: gravitropism; root; tomato.

Figure 1

Average absolute angle and gravitropism in M82 and S. pennellii. (A) Roots of M82 (left) and S. pennellii (right). Scale bar = 1 cm. (B) Quantification of average absolute angle. (C) S. pennellii has a significantly greater average absolute angle relative to M82. ***= P < 0.001 (ANOVA). (D) Gravitropism was measured as the rate of tip angle change over two hours following a 90° rotation, with 0° as horizontal to the right and positive angle clockwise (as the tip bends downward). (E) Mean tip angle of M82 (n = 89) and S. pennellii (n = 18) as a function of time after rotation by 90 degrees. Error bars represent standard error of the mean. (F) Mean tip angle curve for all rotated (Rot) roots (solid) has the same response curve shape as, and is intermediate between, curves for roots rotated (rot) through or away from the gravity vector (dashed). M82: n = 36 away, n = 53 through; S. pennellii: n = 10 away, n = 8 through. Error bars represent standard error of the mean. (G) Mean rate of change of tip angle (swing rate), per time point in the two hours following 90 degree rotation of M82 (n = 89) and S. pennellii (n = 18). Error bars represent standard error of the mean.

Figure 2

The genetic architecture of root angle is largely distinct from that of root gravitropism. (A) ILs showing significant differences in swing rate over time. Time points are separated by 3.75 min. Early response = T0-T6, middle response = T7-T19; late response = T20-T33. Color indicates magnitude of effect (difference of means of swing rates of ILs vs. M82), and only ILs with p_FDR ≤ 0.05 (ANOVA) are colored. (B) Overlap in QTL for root angle and QTL for gravitropism at three or more consecutive time points (p_FDR ≤ 0.05, ANOVA) relative to M82.

Figure 3

Differentially expressed genes associated with cis- and trans-eQTL in the root tips of ILs. (A) Number of differentially expressed genes (y-axis) between each IL (x-axis) and M82 at p_FDR ≤ 0.01 (negative binomial distribution tests with edgeR). Blue = cis-eQTL; Gray = trans-eQTL. (B) Tomato eFP browser with chromosome positions of introgressions in ILs. Expression of Solyc07g008570 (SlPAP27) is indicated in each IL. Asterisks indicate significant differences from M82 (p_FDR ≤ 0.01).

Figure 4

Solyc07g008570 is localized to a genetic interval associated with regulating root angle and its expression correlates with root growth angle in ILs with introgressions on chromosome 7. (A) Bin d-7B-3 contains 19 genes. Chromosome 7 position (Mbp) is indicated on the x-axis, and each S. pennelli introgression within the IL or sub-IL is colored green for a significantly increased avgAbsAng relative to M82 or red if it is not different from M82. Light red lines indicate approximate boundaries of each bin. (B) Expression of Solyc07g008570 is positively correlated with avgAbsAng using the ILs defining bin d-7B and as determined using RNA-seq (Pearson correlation), error bars are standard error of the mean, n_EXPR: M82:4, S. pennellii:3; IL7-4:4; IL7-4-1:4, 7-5;3; n_ANGLE: M82:64, S. pennellii:78; IL7-4:34; IL7-4-1:38; IL7-5:29. (C) Quantitative real-time PCR confirmation of the increased expression of Solyc07g008570 in ILs with increased avgAbsAng, n = 3 for each genotype, error bars represent standard deviation. (D) An increase of AtPAP27 expression is positively correlated with an increased root angle using multiple independent insertion lines (R² = 0.66, P ≤ 0.05, ANOVA). Error bars represent standard error of the mean, n(Col-0)=275, n(35S:AtPAP27 ∑A:D)=277. (E) The average absolute angle of four independent insertion lines overexpressing AtPAP27 is increased relative to Col-0 (**P ≤ 0.01, ANOVA). Error bars represent standard error of the mean, n_Angle: Col-0:275, 35S:AtPAP27/lineA:11, lineB:20, lineD:83, lineF:100; n_Expr: Col-0:19, 35S:AtPAP27/lineA:11, lineB:20, lineD:22, lineF:7.

Figure 5

AtCDC73 regulates root angle in Arabidopsis. (A) GWAS Manhattan Plot for SNPs associated with median direction index. Black box indicates associated genomic region of CDC73. (B) Loss of CDC73 function in two alleles (cdc73-1, cdc73-2) leads to an increased root growth angle relative to Col-0 or the FRI control. *P ≤ 0.05 **P ≤ 0.01 ***P ≤ 0.001 as determined using an ANOVA. n(Col-0)=144, n(cdc73-1)=85, n(cdc73-2)=26, n(FRI)=22, n(FRI/cdc73-1)=32.