A telomere-to-telomere gapless genome reveals SlPRR1 control of circadian rhythm and photoperiodic flowering in tomato

Abstract

Cultivated tomato (Solanum lycopersicum) is a major vegetable crop of high economic value that serves as an important model for studying flowering time in day-neutral plants. A complete, continuous, and gapless genome of cultivated tomato is essential for genetic research and breeding programs. Here, we report the construction of a telomere-to-telomere (T2T) gap-free genome of S. lycopersicum cv. VF36 using a combination of sequencing technologies. The 815.27-Mb T2T "VF36" genome contained 600.23 Mb of transposable elements. Through comparative genomics and phylogenetic analysis, we identified structural variations between the "VF36" and "Heinz 1706" genomes and found no evidence of a recent species-specific whole-genome duplication in the "VF36" tomato. Furthermore, a core circadian oscillator, SlPRR1, was identified, which peaked at night in a circadian rhythm. CRISPR/Cas9-mediated knockdown of SlPRR1 in tomatoes demonstrated that slprr1 mutant lines exhibited significantly earlier flowering under long-day condition than wild type. We present a hypothetical model of how SlPRR1 regulates flowering time and chlorophyll biosynthesis in response to photoperiod. This T2T genomic resource will accelerate the genetic improvement of large-fruited tomatoes, and the SlPRR1-related hypothetical model will enhance our understanding of the photoperiodic response in cultivated tomatoes, revealing a regulatory mechanism for manipulating flowering time.

Keywords: SlPRR1; chlorophyll biosynthesis; cultivated tomato T2T genome; flowering time; photoperiod.

Figure 1:

Complete genome assembly and annotation of the VF36 tomato. (A) Circos plot of VF36 genome annotation. Quantitative tracks are aggregated in a 10-kb window. Track a, chromosomes information. Track b, gene density. Track c, GC content. Track d, repeat coverage. Track e, collinearity information. (B) Track displaying density of Gypsy, Copia, CACTA, PIF-Harbinger, Tc1, hAT, LINE, and Helitron elements. (C) Map of centromere prediction for VF36 genome. (D) Distribution of VF36 genomic features.

Figure 2:

Comparative genomic analysis of the VF36 genome. (A) Structural variations between Heinz_1706 and VF36 genomes. (B) Estimation of divergence time and gene family expansion/contraction. The blue blocks represent the published whole-genome duplication (WGD) events. The dark yellow blocks represent the published whole-genome triplication events. (C) Venn diagram of gene family clustering. (D) The GO enrichment on the genes in SV regions.

Figure 3:

Gene duplication and evolution. distribution from orthologs and paralogs among S. pimpinellifolium, S. tuberosum, N. benthamiana, V. vinifera, and S. lycopersicum.

Figure 4:

Expression and functional analysis of SlPRR1 in circadian rhythm. (A) Expression of the SlPRR1 gene in various stages of tomato tissues. (B) Expression of SlPRR1 gene from tomato plants grown in 12L/12D. Shading indicates the dark period.

Figure 5:

Functional analysis of SlPRR1 in the regulation of tomato flowering time. (A) The 2 sgRNA target sites in SlPRR1 locus used for the CRISPR/Cas9 gene-editing system. (B) The mutation types of SlPRR1 in slprr1-35, slprr1-5, slprr1-6, and slprr1-10 line. Red letters indicate the substitution sites, green letters indicate the insertion sites, and blue letters indicate the PAM. (C–E) Flowering phenotype from 40-day-old (C) and 73-day-old (D) tomato plants and flowering time (E) of the WT, slprr1-6, and slprr1-35 lines under LD (16L/8D) and SD (8L/16D) conditions. (F) The content of sucrose, fructose, and glucose of red ripening tomato fruits in WT and slprr1 lines under SD or LD conditions. (G, H) The chlorophyll (G) and nitrogen (H) content of tomato leaves in WT and slprr1 lines under SD or LD conditions during the photoperiod.

Figure 6:

Chlorophyll synthesis and degradation of tomato leaves in WT and slprr1 lines under SD or LD conditions. (A) The potential simplified chlorophyll synthesis pathway in tomato leaves is depicted. (B) The potential simplified chlorophyll degradation pathway in tomato leaves is depicted.

Figure 7:

Relative expression levels of photosynthesis and chlorophyll synthesis-related genes in tomato leaves. Error bars represent the averages of 3 biological replicates ± SD. Asterisks indicate statistical significance (*P < 0.05, **P < 0.01, Student’s t-test).

Figure 8:

Relative expression levels of flowering-related genes in tomato. Error bars represent the averages of 3 biological replicates ± SD. Asterisks indicate statistical significance (*P < 0.05, **P < 0.01, Student’s t-test).

Figure 9:

Model for the regulation of flowering time and chlorophyll synthesis by knockout SlPRR1 in tomato under LD and SD conditions.