Optimization of Tomato Shoot Architecture by Combined Mutations in the Floral Activators FUL2/MBP20 and the Repressor SP

Abstract

Shoot determinacy is a key trait affecting productivity in tomato, quantitatively governed by genes within the flowering pathway. Achieving an optimal balance of flowering signals is essential for shaping plant architecture and maximizing yield potential. However, the genetic resources and allelic diversity available for fine-tuning this balance remain limited. In this work, we demonstrate the potential for directly manipulating shoot architecture by simultaneously targeting the flowering activating FRUITFULL(FUL)-like genes, FUL2 and MADS-BOX PROTEIN 20 (MBP20), and the flowering-repressing gene SELFPRUNING (SP). Loss of MBP20 in the sp background leads to additional inflorescences, while determinacy is largely maintained. However, additional mutation of FUL2 results in mainly indeterminate plants, which have faster sympodial cycling, leading to more compact growth and increased flower production. Our results provide a path to quantitative tuning of the flowering signals with a direct impact on shoot architecture and productivity.

Keywords: flowering; shoot architecture; sympodial growth; tomato.

Figure 1

Mutations in SP and FUL2/MBP20 can be combined to create variation in shoot architecture compactness. (A) Sequences of SP alleles (a) obtained with CRISPR/cas9 using three guide RNAs (gRNAs), namely, sp (a1, a2), sp (a3, a4) mbp20, and sp (a5) ful2 mbp20. The mbp20 and ful2 mbp20 lines were previously generated [19]. The gRNA and protospacer-adjacent motif (PAM) sequences are shown in bold red and black, respectively. Deletions and insertions are indicated by blue dashes and blue font, respectively, and the lengths of sequence gaps are indicated in parentheses. (B) Quantification of primary shoot flowering time for wild-type (WT) and mutant plants. n: numbers of individual plants measured. quad ful: ful1 ful2 mbp10 mbp20 quadruple mutant (generated in [19]). (C) Representative main shoots from all genotypes. Three-month-old plants are shown. L: leaf; D/ID: determinate and indeterminate growth. White bar: 5 cm. (D) Average leaf number in sympodial shoots across all genotypes, measured for the first five successive sympodial units. (E) Proportion of shoot determinacy of the genotypes. (F) Quantification of inflorescence numbers in determinate plants of sp, sp mbp20, and sp ful2 mbp20 mutants. In (B,D,F), mean values (±SE) were compared between genotypes using one-way ANOVA followed by a post hoc LSD test. Statistical significance in (F) was assessed using the Wilcoxon rank-sum test. Different letters indicate significant differences at the p < 0.05 level.

Figure 2

Quantification of flower production. (A) Proportion of branched inflorescences per branching category for the indicated genotypes. The numbers (0–3) indicate the number of branching events. (B,C): Quantification of flower numbers per inflorescence and the total flower number per plant. In (B,C), mean values (±SD) were analyzed for statistical significance using a t-test. Significant differences compared to WT plants (B) and sp plants (C) are represented by asterisks: ** p < 0.01. ns: non-significant.

Figure 3

Gene expression analysis in the sympodial vegetative meristems. (A) Microdissection of the SYM stage was performed for gene expression analysis. Dashed line represents the boundary of the dissected tissue. White bar: 200 μm. (B,C) Gene expression of FUL-like genes (B) and AP2-like genes (C) in SYM detected by qRT-PCR. The values shown (mean ± SE) are the average of three replicates. Significant differences were calculated using a one-tailed Student’s t test (* p < 0.05 and ** p < 0.01).

Figure 4

Model of shoot determinacy in relation to FUL-like flowering signals. When flowering signals are reduced, sympodial flowering is delayed, resulting in an indeterminate shoot growth habit.