It's not easy being green tomatoes


Editor's Introduction

Uniform ripening encodes a Golden 2-like transcription factor regulating tomato fuit chloroplast development

When shopping for tomatoes, people often look for a uniform scarlet hue.  Plant breeders have selected for plants that have uniform light green fruit that will develop the characteristic red hue uniformly as it ripens. However, once these green tomatoes turn the perfect red they tend to lack the sweetness and flavor of those more imperfect tomatoes grown in the garden at home, and researchers have now identified the genetic reason behind this. The responsible gene is Golden 2-like (GLK) 2. When GLK2 is expressed, there is an increase in the fruit's photosynthetic capacity, resulting in higher sugar content and a favorable fruit.  Unfortunately, in uniformly colored tomatoes, GLK2 is inactivated.  Should tomato lovers start sacrificing beauty for taste?

Paper Details

Original title
Uniform ripening encodes a Golden 2-like transcription factor regulating tomato fuit chloroplast development
Ann Powell
Original publication date
Vol. 336 no. 6089 pp. 1711-1715
Issue name


Modern tomato (Solanum lycopersicum) varieties are bred for uniform ripening (u) light green fruit phenotypes to facilitate harvests of evenly ripened fruit. U encodes a Golden 2-like (GLK) transcription factorSlGLK2, which determines chlorophyll accumulation and distribution in developing fruit. In tomato, two GLKs—SlGLK1 and SlGLK2—are expressed in leaves, but only SlGLK2 is expressed in fruit. Expressing GLKs increased the chlorophyll content of fruit, whereas SlGLK2 suppression recapitulated the u mutant phenotype. GLK overexpression enhanced fruit photosynthesis gene expression and chloroplast development, leading to elevated carbohydrates and carotenoids in ripe fruit. SlGLK2 influences photosynthesis in developing fruit, contributing to mature fruit characteristics and suggesting that selection of u inadvertently compromised ripe fruit quality in exchange for desirable production traits.


For ~70 years, breeders have selected tomato varieties with uniformly light green fruit before ripening, a characteristic that facilitates maturity determinations and promotes even ripening at the stem end (12). However, light green fruit ripen with reduced sugars, compromising traits that are valuable for processed products and the flavor of fresh fruit (fig. S1) (35). The uniform ripening (u) locus determines the intensity and pattern of chlorophyll distribution in unripe fruit (357). The dominant U allele results in fruit with dark green shoulders at the stem end adjacent to the pedicel; u/u fruit are uniformly light green.

Chloroplast formation, chlorophyll synthesis, and photosystem assembly require exposure to light and genetically defined developmental cues (8). Developing tomato fruit are capable of photosynthesis and contribute up to 20% of the fruit’s photosynthate, with the remainder translocated from leaves (9). Light-harvesting electron transfer and CO2 fixation proteins are conserved in fruit (912) and regulated by light and developmental signals, as in leaves. However, differences suggest additional levels of fruit-specific regulation (1317).

Two GARP family Myb transcription factors [Golden 2-like 1 (GLK1) and GLK2] determine the capacity for light-stimulated photosynthesis by controlling chloroplast formation (1820). In C3 photosynthesizing tissuesGLK1 and GLK2 act redundantly, but each may have undefined specialized roles in C4 tissues (19). Although the Atglk2 and Atglk1-Atglk2 Arabidopsis mutants have pale siliques (18), the contributions of GLKs to fleshy fruit chloroplast development are unknown. Expression of AtGLK1 or AtGLK2 is sufficient for leaf chloroplast development through control of similar photosystem, light-harvesting complex, and chlorophyll biosynthetic genes (18, 20–22), although aspects of their function and/or regulation may have evolved distinctions (2324). We report the identification of tomato GLKs and show that the u phenotype results from a mutation in SlGLK2.

Using two interspecific populations segregating for the u locus (13,2527) (Fig. 1A), U mapped to a 60,507–base pair (bp) region on the short arm of chromosome 10 containing eight predicted genes, including SlGLK2, located at position Sl2.40chr10:2291209-2295578 (Fig. 2A). Sequencing full-length SlGLK2 transcripts predicted that in U genotypes, SlGLK2 encodes a 310–amino acid protein (fig. S2A), but in uSlglk2 encodes a truncated 80–amino acid protein because of a single base insertion that causes a frameshift and a premature stop codon (fig. S2B). The additional adenine (A) between SL2.40ch10:2292260-2292267 is the only difference in the SlGLK2 sequence that is common to all light green u/u varieties and absent in all dark-green shouldered U/U varieties (Fig. 2B).


Fig. 1.  Fruit phenotypes. (A) Immature green fruit (IM, 15 dpa) from theS. lycopersicum “M82” x S. pennellii introgression population andS. lycopersicum “Moneymaker” x S. pimpinellifolium backcross lines. (B) IM fruit from S. lycopersicum “T63” containing the CaMV35S promoter (Tcontrol) or AtGLK1 or AtGLK2 with theCa35S (p35S), RbcS (pRbcS), LTP (pLTP), or PDS (pPDS) promoter. (C) Mature green (32 dpa) fruit from “Ailsa Craig” U/U and “Craigella” u/u. (D) Mature green fruit from Ailsa Craig U/containing p35S::SlGLK2 with overexpression (OE) or cosuppression (CS) of SlGLK2 and from M82 u/u overexpressing p35S::SlGLK2.

Phenotypes of tomato fruits

Phenotypes of tomato fruits resulting from breeding experiments and introducing transgenes. 

The purpose of this figure is to show images of the immature fruits resulting from the experiments in this paper as a reference to the reader. 

Panel A

Photos of immature fruit of hybrids from breeding experiments.

Parent plants S. lycopersicuum (homozygous for u allele = M82) and S. pennellii (Wildtype for U allele = LA0716) were bred to generate hybrids, which were then used to generate S. lycopersicum that has the wildtype genotype (homozygous U/U allele).

Three of these lines, IL10-1, IL10-1-1, and IL10-2  have this and the resulting phenotype is dark-green shoulders.

Breeding experiments of S. lycopersicum (homozygous for u allele = Moneymaker) and S. pimpinellifollium (Wildtype for U allele = TO-973).

The first-generation hybrid (F1 = U/u genotype), looks like an intermediate for size and has dark green shoulders.

Breeding the F1 plants with the S. lycopersicum parent (called a backcross) results in two separate phenotypes: BC2S1 49-B has wildtype genotype and dark green shoulders; backcross BC2S1 13-A has u/u genotype and pale green fruit.

Panel B

Fig 1B: S. lycopersicum immature containing transgenes. 

T63 is the control showing that simply introducing the delivery system for the transgene (p35S) doesn’t affect phenotype.

Plants that contain the transgenes p35S::AtGLK1 and p35S::AtGLK2, where the transgene is expressed all the time, or pRbcS::AtGLK1, pRbcS::AtGLK2, pLTP::AtGLK1 and pLTP::AtGLK2, where the transgene is expressed early in development, have dark green fruit .

When the transgene is expressed later in development (pPDS::AtGLK1 or pPDS::AtGLK2), the fruit remains light green.

Panel C

Immature fruit phenotypes for S. lycopersicum “Alisa Craig” (genotype U/U) and S. lycopersicum “Craigella” (genotype u/u). 

Panel D

Immature fruit phenotypes for S. lycopersicum “Alisa Craig” (genotype U/U) containing transgenes that overexpress or suppress GLK2.

Overexpression of GLK2 (OE p35S::SlGLK2) results in very dark green fruit. Inhibition of gene expression of GLK2 (CS p35S::SlGLK2) results in light-green fruit.  

Increased gene expression of GLK2 (OE p35S::SlGLK2) in S. lycopersicum “M82” (genotype u/u) results in dark green fruit.


Fig. 2.  Map position and sequence of SlGLK2 alleles. (A) Left to right, the classical morphological map (26), IL map (27), maps derived from the IL 10-1 F2 and the MM x “T0937” populations, candidate genes in the 60,507-bp region defined by markers, and gene model of SlGLK2 with the additional A underlined and the resulting stop codon in Slglk2/u allele circled. (B) SlGLK2/U and Slglk2/u coding sequences. Nucleic acid calls between SL2.40ch10:2292143-2295824 (SGN tomato release 2.4) from cDNA and genomic sequencing of the u/u S. lycopersicum varieties “Heinz 1706” (HEI), M82, T63, Moneymaker (MM), “Castlemart” (CSM), “E6203” (E6), “Fireball” (FB), “Long Red” (uLR), “N93,” S. lycopersicum var. cerasiforme PI114490 (Cer), and Craigella (CRG) and the U/U S. pimpinellifolium (Spi), S. pennellii (Spe), and S. lycopersicoides (Sha) wild tomato relatives and S. lycopersicum varieties Ailsa Craig (AC), “73X,” “T91,” and a “Cuatomate” (CAU) landrace. Translated reverse transcription polymerase chain reaction (RT-PCR) products and Basic Local Alignment Search Tool (BLAST) searches predicted the start codon at 2292050; no differences were detected until position 2292143. The sequence differences at 2292260-2292267 of all u/u varieties compared with all U/U varieties is boxed in orange.

Questions being asked

What is u?

What is the change in the DNA sequence that results in a difference in the function of that protein?

Animations on genetic mapping

SlGLK2 and an additional GLK homolog (SlGLK1) were identified in tomato and related Solanaceae species (fig. S2A). The amino acid sequences of the Solanaceae GLKs are similar to other dicot GLKs and are approximately 45% identical to their Arabidopsis counterparts (fig. S2, A, C, and D).

SlGLK1 and SlGLK2 expression predicts their roles in leaves and fruit. Transcripts from both SlGLK1 and SlGLK2 accumulate in cotyledons, sepals, and leaves, but only SlGLK2 transcripts accumulate in green fruit (Fig. 3A).SlGLK2 is eightfold more abundant in the pedicel (shoulder) than in the blossom (stylar) portions, suggesting that SlGLK2 contributes to the pattern and intensity of chlorophyll accumulation (table S2). Maturation in the absence of light reduces, but does not eliminate, SlGLK2 expression, which remains 40- to 300-fold greater than that ofSlGLK1 (table S3). Therefore, SlGLK2 expression is partially regulated by light. In u/u fruit, Slglk2 expression was 26 to 40% of SlGLK2 levels (Fig. 3A and table S2). Fruit that develop in the dark with either SlGLK2 allele are pale (fig. S4), confirming that light is essential for fruit chloroplast development and chlorophyll synthesis, but the pattern and extent of chloroplast development is determined by SlGLK2 (table S3).


Fig. 3.  GLK expression and chlorophyll accumulation. (A) SlGLK1 and SlGLK2 RT-PCR products from Ailsa Craig U/U and Craigella u/cotyledons (C), young leaves (YL), developed leaves (OL), flower petals (P), stamens (S), immature fruit (IM, 10-15 dpa), and the pedicel shoulders (Pd) or the stylar ends (St) of mature green (MG, 32 dpa) fruit and ripe fruit (RR, 46 dpa). Results are typical of replicated RNA samples. (B) Chlorophyll in the outer pericarp and epidermis of IM fruit from AtGLK1- or AtGLK2-expressing lines. Statistical significance by means of general linear model (GLM) and Bonferroni multiple comparison test (MCT) at P < 0.05 are indicated. (C) Quantitative RT-PCR of AtGLK1 and AtGLK2 expression in IM fruit. Statistical significance by means of GLM and Tukey’s honestly significant difference (HSD) test at P < 0.05 are indicated by roman (AtGLK1) and italic (AtGLK2) letters, respectively. (D) Chlorophyll in leaves from AtGLK-expressing lines. Statistical significance determined by means of GLM and Bonferroni MCT at P< 0.05 are indicated.

Question being asked

Which tissues are SlGLK1 and SlGLK2 expressed in?

Does their expression affect the amount of chlorophyll (impacting how dark green the immature fruits are) in these tissues?

Panel A

Reverse Transcriptase PCR (RT-PCR), a method where RNA is harvested from the organism of interest, and it's abundance is measured by comparing it with a DNA ladder containing DNA fragments of defined length to determine whether this gene is expressed.

The RNA needs to be converted to complementary DNA (cDNA) in order for PCR to take place.

This experiment shows that for the S. lycopersicum “Aillsa Craig” plant (genotype = U/U), SlGLK1 and SlGLK2  are expressed in leaves, and SlGLK2, but not SlGLK1, is also expressed in the fruits.

Useful animations:


PCR learning lab, Univ. Utah

Panel B

Total chlorophyll content in the immature fruits of plants containing transgenes.

TControl is the plant that only has the delivery system p35S.

Tcontrol has only the 35S promoter but no gene linked to it. All of the plants also have antibiotic resistance genes as part of the suite of genes introduced into them.

Panel C

Quantitative RT-PCR is used to measure expression of the transgenes in the immature fruits.   

Useful animation about Reverse Transcription

Panel D

Total chlorophyll content in the leaves of plants containing transgenes.

TControl is the plant that only has the delivery system p35S.

As you can see, there's no significant difference in chlorophyll content.

We tested whether the u phenotype is altered by GLK expression. Expression of AtGLK1 or AtGLK2 in a u/tomato variety (28) with promoters expressed before ripening resulted in homogeneously dark green unripe fruit (Fig. 1B and fig. S3, A to C), with three- to sixfold elevated chlorophyll contents (Fig. 3B). With a promoter expressed later in fruit development, AtGLK1 or AtGLK2 mRNA was not detected, and chlorophyll was not elevated (Figs. 1B and 3, B and C). The chlorophyll contents of the leaves were not different from controls (Fig. 3D). The chlorophyll a/b ratio was unchanged by the expression of AtGLK1 or AtGLK2. Fruit set, size, overall color, and time to ripen were indistinguishable from the control fruit (fig. S3). We also examined the effects of ectopic expression of SlGLK2 in U/U and u/u genotypes. Expression of a full-length SlGLK2 cDNA in either genotype resulted in homogeneously dark green unripe fruit (Fig. 1D). Cosuppression of SlGLK2 in four U/U transgenic lines (28) converted the dark green shoulder U trait to light green, confirming that SlGLK2 is U (Fig. 1Dand fig. S5). The dark green fruit phenotype is confined to the shoulder region, where SlGLK2 is more highly expressed in U/U varieties (Fig. 1, C and D), and all lines expressing GLKs with promoters expressed throughout the fruit produced homogeneously dark green fruit, affirming that the manifestation and intensity of the phenotype depends on the spatial pattern and level of GLK expression.

Transmission electron microscopy (TEM) revealed that AtGLK1 or AtGLK2 expression increased the number (twofold) and size of green fruit chloroplasts and promoted accumulation and development of grana thylakoids (Fig. 4A). Chloroplasts of green fruit expressing AtGLKs had 6.6 ± 0.45 thylakoids/granum; green control fruit had 3.1 ± 0.84 thylakoids/granum. No obvious alterations of leaf chloroplasts were observed (Fig. 4A).


Fig. 4.  AtGLK-expressing fruit characteristics. (A) TEM of IM (15 dpa) green fruit (top row) and leaf (bottom row) chloroplasts, 0.5 μm scale. (B) Cellular components Gene Ontology (GO) terms for significantly (P < 0.05, fold change >2) overexpressed genes in IM fruit identified in microarray hybridizations. The total number of genes with known GO terms is shown below bars. (C) GO categories of overrepresented genes (P < 0.05, n > 3 genes) whose transcript abundance were different (P < 0.05, fold change >2). *P < 0.0001; **0.0001 < P < 0.001; ***0.001 < P <0.01. (D) Starch in IM fruit pericarp and epidermis. (E) Glucose (dark bar) and fructose (light bar) in red ripe (RR, 42 dpa) fruit. (F) Total soluble solids in RR fruit. Statistical significance determined by means of GLM and Tukey’s HSD at P < 0.05 are indicated.

Question being asked

Does the mutation in SlGLK2 impact the way chloroplasts develop?

Does it affect the components that lead to good-tasting fruit (starch and sugar content)?

Panel A

Visualizing chloroplasts from immature fruit and leaves for plants overexpressing AtGLK1 (p35S::AtGLK1) and AtGLK2 (p35S::AtGLK2) using a technique called Transmission Electron Microscopy (TEM).

Panel B

Microarray analysis to assess differences in gene expression between a control plant and one that contains transgenes p35S::AtGLK1 and p35S::AtGLK2, where the transgene is expressed all the time and at high levels.

Specifically, the authors are interested in identifying genes were expression is higher  (= up-regulated) in immature fruits compared to wildtype when overexpressing GLK1, GLK2 or both. 

Panel C

List of types of overrepresented genes where mRNA generated significantly more compared with the control.

Panel D

Total starch content in fruits overexpressing GLK1 and GLK2.

Panel E

Percent glucose and fructose in fruits overexpressing GLK1 and GLK2.

Panel F

Total soluable solid content in fruits overexpressing GLK1 and GLK2.

Analysis of transcript abundance in immature green fruit (28) demonstrated that constitutive expression of AtGLK1 or AtGLK2 in u/u increased accumulation of transcripts from 672 genes (fig. S6), especially those with photosynthetic functions (Fig. 4C). Of the 127 genes with known cellular compartments, 47% of those up-regulated by AtGLK1 and AtGLK2 are predicted to function in chloroplasts (Fig. 4B). Expression of genes associated with chlorophyll biosynthesis, light-harvesting complexes, photosystem, and starch metabolism increased owing to expression of AtGLK1 or AtGLK2 (Fig. 4C and tables S4 to S6). In atglk1-atglk2 lines constitutively overexpressing rice or Arabidopsis GLKs, similar classes of genes are up-regulated (2022). Expression of the photomorphogenic regulators DE-ETIOLATED1 (DET1), UV-DAMAGED DNA-BINDING PROTEIN 1 (DDB1), and ELONGATED HYPOCOTYL5 (HY5) was not altered by AtGLK expression (table S4). Constitutive photomorphogenesis in the tomato hp1 (DDB1) mutant does not reduce the low levels of SlGLK1 expression (29) and only slightly elevates SlGLK2 expression, suggesting distinct routes to plastid regulation via photomorphogenesis and GLK expression. Chloroplast development is exaggerated with costs to yield when the negative regulators of photomorphogenesis, DET1 and DDB1, are down-regulated, with or without GLK expression (3031). Expression of AtGLK1 or AtGLK2 did not affect fruit yield.

In further characterizing the effects of GLKs on fruit biology and quality, increased starch levels in green fruit were observed in response to AtGLK expression (28). Furthermore, AtGLK expression increased fructose and glucose 40% in red fruit (Fig. 4, D and E). Ripe fruit expressing AtGLK1 or AtGLK2 had a 21% increase in soluble solids (Fig. 4F). A quantitative trait locus for increased soluble solids was reported near the U locus (5). The increase in soluble solids in U/U compared with that of u/u was 10% (fig. S1), presumably because SlGLK2 expression was enhanced in the green fruit shoulders. Lycopene (28) increased 10 to 60% with ectopic expression of AtGLKs (fig. S8), supporting the conclusion that GLK expression regulating chloroplast development in unripe fruit affects sugars and the predominant carotenoid in ripe fruit.

Whereas u—that is, Slglk2—results in reduced soluble solids in ripe fruit (Fig. 4F and fig. S1), the soluble solids in red u/u fruit that develop in the dark decreased by an additional 30% (fig. S7). Thus, both GLK activity and light regulate photosynthetic capacity in green fruit through their regulation of chlorophyll accumulation and chloroplast development and ultimately contribute to sugars that accumulate in ripe fruit.

As in many other plants, two GLK genes are present and expressed in tomato, but in fruit, SlGLK2 mRNA predominates and accumulates in a spatial pattern consistent with chlorophyll biosynthesis and chloroplast development. All u/u cultivars examined contain a Slglk2 allele encoding a truncated loss-of-function GLK protein. Our results suggest that breeding selections for the u fruit trait that is helpful for harvesting methods may have had an unintended negative impact on fruit quality because suboptimal chloroplasts develop, and consequently, ripe fruit sugar and lycopene levels decrease. Manipulation of GLK levels or spatial expression patterns represents an opportunity to recover and enhance production and quality traits in tomato and other crop species.

Supplementary Materials

Materials and Methods

Figs. S1 to S8

Tables S1 to S8

References (3258)

References and Notes

  1. L. Butler, The linkage map of the tomato. J. Hered. 43, 25 (1952).

  2. F. Yeager, The uniform fruit color gene in the tomato. Proc. Am. Soc. Hort. Sci. 33, 512 (1935

  3. S. M. Kinzer, S. J. Schwager, M. A. Mutschler, Mapping of ripening-related or -specific cDNA clones of tomato (Lycopersicon esculentum). Theor. Appl. Genet. 79, 489 (1990).

  4. C. M. Rick, High soluble-solids content in large-fruited tomato lines derived from a wild green-fruit species. Hilgardia 42, 493 (1974).

  5. S. D. Tanksley, J. Hewitt, Use of molecular markers in breeding for soluble solids content in tomato—A re-examination. Theor. Appl. Genet. 75, 811 (1988).

  6. G. A. Kemp, I. L. Nonnecke, Differences in intensity of unripe fruit colour in the tomato. Can. J. Plant Sci. 40, 306 (1960).

  7. C. M. Rick, L. Butler, Cytogenetics of the tomato. Adv. Genet. Incorp. Mol. Gen. Med. 8, 267 (1956).

  8. M. T. Waters, J. A. Langdale, The making of a chloroplast. EMBO J. 28, 2861 (2009).

  9. S. Hetherington, R. Smillie, W. Davies, Photosynthetic activities of vegetative and fruiting tissues of tomato. J. Exp. Bot. 49, 1173 (1998).

  10. M. M. Blanke, F. Lenz, Fruit photosynthesis. Plant Cell Environ. 12, 31 (1989).

  11. S. Carrara, A. Pardossi, G. F. Soldatini, F. Tognoni, L. Guidi, Photosynthetic activity of ripening tomato fruit. Photosynthetica 39, 75 (2001).

  12. J. Mataset al., Tissue- and cell-type specific transcriptome profiling of expanding tomato fruit provides insights into metabolic and regulatory specialization and cuticle formation. Plant Cell 23, 3893 (2011).

  13. T. Manzara, P. Carrasco, W. Gruissem, Developmental and organ-specific changes in DNA-protein interactions in the tomato rbcS1, rbcS2 and rbcS3A promoter regions. Plant Mol. Biol. 21, 69 (1993).

  14. M. Sugita, W. Gruissem, Developmental, organ-specific, and light-dependent expression of the tomato ribulose-1,5-bisphosphate carboxylase small subunit gene family. Proc. Natl. Acad. Sci. U.S.A. 84, 7104 (1987).

  15. L. A. Wanner, W. Gruissem, Expression dynamics of the tomato rbcS gene family during development. Plant Cell 3, 1289 (1991).

  16. B. Piechulla, W. Gruissem, Diurnal mRNA fluctuations of nuclear and plastid genes in developing tomato fruits. EMBO J. 6, 3593 (1987).

  17. B. Piechulla, R. E. Glick, H. Bahl, A. Melis, W. Gruissem, Changes in photosynthetic capacity and photosynthetic protein pattern during tomato fruit ripening. Plant Physiol. 84, 911 (1987).

  18. D. W. Fitter, D. J. Martin, M. J. Copley, R. W. Scotland, J. A. Langdale, GLK gene pairs regulate chloroplast development in diverse plant species. Plant J. 31, 713 (2002).

  19. J. A. Langdale, C4 cycles: Past, present, and future research on C4 photosynthesis. Plant Cell 23, 3879 (2011).

  20. M. T. Waters et al., GLK transcription factors coordinate expression of the photosynthetic apparatus in Arabidopsis. Plant Cell 21, 1109 (2009).

  21. M. T. Waters, E. C. Moylan, J. A. Langdale, GLK transcription factors regulate chloroplast development in a cell-autonomous manner. Plant J. 56, 432 (2008).

  22. H. Nakamura et al., Ectopic overexpression of the transcription factor OsGLK1 induces chloroplast development in non-green rice cells. Plant Cell Physiol. 50, 1933 (2009).

  23. Bravo-Garcia, Y. Yasumura, J. A. Langdale, Specialization of the Golden2-like regulatory pathway during land plant evolution. New Phytol. 183, 133 (2009).

  24. Y. Yasumura, E. C. Moylan, J. A. Langdale, A conserved transcription factor mediates nuclear control of organelle biogenesis in anciently diverged land plants. Plant Cell 17, 1894 (2005).

  25. H. Paterson et al., Resolution of quantitative traits into Mendelian factors by using a complete linkage map of restriction fragment length polymorphisms. Nature 335, 721 (1988).

  26. S. D. Tanksley, M. A. Mutschler, C. M. Rick, in Genetic Maps, S. J. O’Brien, Ed. (Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, 1987), pp. 655–669.

  27. Y. Eshed, D. Zamir, An introgression line population of Lycopersicon pennellii in the cultivated tomato enables the identification and fine mapping of yield-associated QTL. Genetics 141, 1147 (1995).

  28. Materials and methods are available as supplementary materials on Science Online.

  29. J. Rohrmann et al., Combined transcription factor profiling, microarray analysis and metabolite profiling reveals the transcriptional control of metabolic shifts occurring during tomato fruit development. Plant J. 68, 999 (2011).

  30. E. M. A. Enfissi et al., Integrative transcript and metabolite analysis of nutritionally enhanced DE-ETIOLATED1 downregulated tomato fruit. Plant Cell 22, 1190 (2010).

  31. Y. Liu et al., Manipulation of light signal transduction as a means of modifying fruit nutritional quality in tomato. Proc. Natl. Acad. Sci. U.S.A. 101, 9897 (2004).

  32. M. T. Waters et al., GLK transcription factors coordinate expression of the photosynthetic apparatus in Arabidopsis. Plant Cell 21, 1109 (2009).

  33. L. V. Savitch, R. Subramaniam, G. C. Allard, J. Singh, The GLK1 ‘regulon’ encodes disease defense related proteins and confers resistance to Fusarium graminearum in Arabidopsis. Biochem. Biophys. Res. Commun. 359, 234 (2007).

  34. J. Rohrmann et al., Combined transcription factor profiling, microarray analysis and metabolite profiling reveals the transcriptional control of metabolic shifts occurring during tomato fruit development. Plant J. 68, 999 (2011).

  35. E. M. A. Enfissi et al., Integrative transcript and metabolite analysis of nutritionally enhanced DE-ETIOLATED1 downregulated tomato fruit. Plant Cell 22, 1190 (2010).

  36. L. Darby, D. B. Ritchie, I. B. Taylor, in The Glasshouse Crops Research Institute Annual Report (Glasshouse Crops Research Institute, Littlehampton, UK, 1977), pp. 168–184.

  37. M. B. Trevino, M. A. O’Connell, Three drought-responsive members of the nonspecific lipid-transfer protein gene family in Lycopersicon pennellii show different developmental patterns of expression. Plant Physiol. 116, 1461 (1998).

  38. G. Giuliano, G. E. Bartley, P. A. Scolnik, Regulation of carotenoid biosynthesis during tomato development. Plant Cell 5, 379 (1993).

  39. L. A. Wanner, W. Gruissem, Expression dynamics of the tomato rbcS gene family during development. Plant Cell 3, 1289 (1991).

  40. G. Sandmann, S. Römer, P. D. Fraser, Understanding carotenoid metabolism as a necessity for genetic engineering of crop plants. Metab. Eng. 8, 291 (2006).

  41. R. Brent, M. Ptashne, A eukaryotic transcriptional activator bearing the DNA specificity of a prokaryotic repressor. Cell 43, 729 (1985).

  42. N.-C. Lin, R. B. Abramovitch, Y. J. Kim, G. B. Martin, Diverse AvrPtoB homologs from several Pseudomonas syringae pathovars elicit Pto-dependent resistance and have similar virulence activities. Appl. Environ. Microbiol. 72, 702 (2006).

  43. J. J. Fillatti, J. Kiser, R. Rose, L. Comai, Efficient transfer of a glyphosate tolerance gene into tomato using a binary Agrobacterium tumefaciens vector. Nat. Biotechnol. 5, 726 (1987).

  44. Drummond et al., in (2010).

  45. Y. Eshed, D. Zamir, An introgression line population of Lycopersicon pennellii in the cultivated tomato enables the identification and fine mapping of yield-associated QTL. Genetics 141, 1147 (1995).

  46. D. Cantu et al., Ripening-regulated susceptibility of tomato fruit to Botrytis cinerea requires NOR but not RIN or ethylene. Plant Physiol. 150, 1434 (2009).

  47. S. Chang, J. Puryear, J. Cairney, A simple and efficient method for isolating RNA from pine trees. Plant Mol. Biol. Rep. 11, 113 (1993).

  48. J. Vrebalov et al., Fleshy fruit expansion and ripening are regulated by the Tomato SHATTERPROOF gene TAGL1. Plant Cell 21, 3041 (2009).

  49. K. J. Livak, T. D. Schmittgen, Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Δ Δ C(T)) Method. Methods 25, 402 (2001).

  50. R. A. Irizarry et al., Summaries of Affymetrix GeneChip probe level data. Nucleic Acids Res. 31, e15 (2003).

  51. C. Eisenhart, The assumptions underlying the analysis of variance. Biometrics 3, 1 (1947).

  52. K. J. Cheung, V. Badarinarayana, D. W. Selinger, D. Janse, G. M. Church, A microarray-based antibiotic screen identifies a regulatory role for supercoiling in the osmotic stress response of Escherichia coli. Genome Res. 13, 206 (2003).

  53. R. Porra, W. Thompson, P. Kriedmann, Determination of accurate extinction coefficients and simultaneous equations for assaying chlorophylls a and b extracted with four different solvents: verification of the concentration of chlorophyll standards by atomic absorption spectroscopy. Biochim. Biophys. Acta 975, 384 (1989).

  54. D. A. T. Southgate, Determination of Food Carbohydrates (Applied Science Publishers, Barking, Essex, ed. 178, 1976).

  55. W. A. Russin, C. L. Trivett, in Microwave: Techniques and Protocols, R. T. Giberson, R. S. Demaree, Eds. (Humana Press, Totowa, NJ, 2001), pp. 25–35.

  56. J. M. Lee et al., Combined transcriptome, genetic diversity and metabolite profiling in tomato fruit reveals that the ethylene response factor SlERF6 plays an important role in ripening and carotenoid accumulation. Plant J. 70, 191 (2012).

  57. J. Javanmardi, C. Kubota, Variation of lycopene, antioxidant activity, total soluble solids and weight loss of tomato during postharvest storage. Postharvest Biol. Technol. 41, 151 (2006).

  58. Martínez-Valverde, M. J. Periago, G. Provan, A. Chesson, Phenolic compounds, lycopene and antioxidant activity in commercial varieties of tomato (Lycopersicum esculentum). J. Sci. Food Agric. 82, 323 (2002).

  59. Acknowledgments: Minimum Information About a Microarray Experiment (MIAME)–compliant microarray data are available at and at (accession E-MEXP-3652). F. Carrari and A. Fernie provided S. pennellii SlGLK2, and J. Maloof provided S. habrochaites SlGLK2 sequences. The U.S. Department of Agriculture (USDA)/National Institute of Food and Agriculture Solanaceae Coordinated Agricultural Project provided potato data. We are grateful to the Tomato Genome Consortium and the SOL Genomics Network for prepublication access to the tomato genome sequence. The S. pennellii introgression lines were provided by the C. M. Rick Tomato Genetics Resource Center; the S. pimpinellifolium populations were provided by the Instituto de Hortofruticultura Subtropical y Mediterranea “La Mayora,” Consejo Superior de Investigaciones Cientificas; and both populations are available by request from the sources. The AtGLK-expressing lines were provided by Mendel Biotechnology and Seminis/Monsanto Vegetable Seeds. SlGLK2, the corresponding lines, and the F2 10-1 IL x M82 population lines and seeds are available from J.J.G. without restriction. Seminis/Monsanto will make available, upon request, and under a material transfer agreement indicating they are to be used for noncommercial purposes, the following lines: LexA:AtGLK1:p35S:LexA-Gal4; LexA:AtGLK1:pLTP:LexA-Gal4; LexA:AtGLK1:pRbcS:LexA-Gal4; LexA:AtGLK1:pPDS:LexA-Gal4; LexA:AtGLK2:p35S:LexA-Gal4; LexA:AtGLK2:pLTP:LexA-Gal4; LexA:AtGLK2:pRbcS:LexA-Gal4; LexA:AtGLK2:pPDS:LexA-Gal4; plus the T63 control line. Other biological materials are available by request from A.L.T.P. or J.J.G. A.L.T.P., T.H., K.L.-C., R.F.-B., and A.B.B. have filed a provisional U.S. patent application UC #2011-841, “Introduction of wild species GLK genes for improved ripe tomato fruit quality,” through the University of California. A.L.T.P. and A.B.B. have filed the U.S. patent application #2010/0154078, “Transcription factors that enhance traits in plant organs,” through Mendel Biotechnology. Assistance from B. Blanco-Ulate, S. Phothiset, S. Reyes, A. Abraham, L. Gilani, and G. Arellano is gratefully acknowledged. J. Langdale provided helpful advice regarding GLK phylogeny and nomenclature. G. Adamson and P. Kysar, Electron Microscopy (EM) Laboratory, University of California Davis Medical Center did the EM work. University of California Discovery and partners funded the pepper analysis and the initial investigations of the Arabidopsis GLKs. The Vietnam Education Foundation supported C.N. Fundación Genoma España ESPSOL Project provided partial funding to A.G. USDA–Agricultural Research Service, USDA–National Research Initiative (2007-02773), and NSF (Plant Genome Program IOS-0923312) provided support to J.J.G.