Edited by: Manuel Rodriguez-Concepcion, Centre for Research in Agricultural Genomics (CRAG), Spain
Reviewed by: Gianfranco Diretto, Energy and Sustainable Economic Development (ENEA), Italy; Gabriela Toledo-Ortiz, Lancaster University, United Kingdom
*Correspondence: Lauren Stanley,
This article was submitted to Plant Metabolism and Chemodiversity, a section of the journal Frontiers in Plant Science
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In plants, the carotenoid biosynthesis pathway (CBP) is essential for the production of photosynthetic and protective pigments, plant hormones, and visual/olfactory attractants for animal pollinators and seed dispersers. The regulation of carotenoid biosynthesis at the transcriptional level is vitally important for all of these functions and has been the subject of intensive research. Many putative transcriptional regulators, both direct and indirect, have been identified through conventional mutant analysis, transcriptome profiling, yeast one-hybrid screening, and candidate gene approaches. Despite this progress, our understanding of the transcriptional regulation of carotenoid biosynthesis remains fragmented and incomplete. Frequently, a stimulus or regulator is known, but the mechanism by which it affects transcription has not been elucidated. In other cases, mechanisms have been proposed (such as direct binding of a CBP gene promoter by a transcription factor), but function was tested only
Carotenoids are red, orange, and yellow pigments produced by photoautotrophic organisms. In the green tissues of plants, carotenoids are essential for light capture, photoprotection, and stabilization of the photosynthetic apparatus (
Because of their critical importance in the physiology, development, ecology, and evolution of plants, carotenoid metabolism and function have been intensively studied. The highly conserved carotenoid biosynthesis pathway (CBP) has been characterized in many plants (reviewed in
In this review, we will focus on the transcriptional regulation of carotenoid biosynthesis genes. For other aspects of carotenoid regulation, we refer readers to several recent reviews (
Transcriptional regulation of carotenoid biosynthesis pathway (CBP) genes in photosynthetic tissues. The regulation of CBP genes in response to light (sun), senescence (leaf), and high temperature (thermometer) and by epigenetic controls (DNA) is shown. The carotenoid biosynthesis pathway is in black, with carotenoid biosynthesis genes indicated in dark blue. Carotenoid regulators discussed in the paper are shown in light blue, with other regulators in purple. Green arrows indicate positive regulation, while blunt red arrows indicate negative regulation. Solid lines show direct interactions, while dotted lines show indirect/unknown interactions. GGPP, geranylgeranyl pyrophosphate;
We have organized this review by tissue type because carotenoids serve unique functions in photosynthetic tissues, fruits, flowers, seeds, and roots and because the literature is already somewhat structured in this manner. For example, tomatoes are considered the model system for carotenoid biosynthesis in fruits, and
Carotenoids are an integral part of the light harvesting apparatus, capturing light energy and protecting the photosynthetic apparatus from damaging reactive oxygen species (ROS) formed during photosynthesis (
The light signaling machinery of plants has been extensively characterized in
Another important player in light signaling is the bZIP transcription factor Long Hypocotyl 5 (HY5), which acts antagonistically to PIF1 during photomorphogenesis. HY5 activates carotenoid and chlorophyll biosynthesis genes, as well as genes involved in chloroplast development and cotyledon expansion. Unlike PIF1, which is stabilized in the dark by the DET1/DDB1/CUL4 complex, HY5 is stabilized by light (the COP1/DDB1/CUL4 complex targets HY5 for degradation in the dark) (
PIFs are also involved in shade-triggered reduction of carotenoid accumulation in
Carotenoid biosynthesis is also induced when greening is de-repressed in the dark, which can be achieved through the blockage of gibberellic acid (GA) biosynthesis (
While the PIF1/HY5 regulatory mechanism is relatively well understood, there is still much to be learned about the transcriptional regulation of carotenoid biosynthesis during deetiolation. For instance, many other carotenoid biosynthesis genes are de-repressed during photomorphogenesis in
Indeed, a chromatin immunoprecipitation–microarray (ChIP–chip) analysis in
Another thing to consider is that PIF1 is certainly not a specific carotenoid regulator: it has been shown to directly regulate the chlorophyll biosynthesis gene
The intensity of light affects both carotenoid concentration and composition (
Ultraviolet B (UV-B) light also triggers the production of carotenoids, which are directly linked to photoprotection of the photosynthetic apparatus (
The PIF1/HY5 switch can also control
The sensitivity of
Leaf senescence is a developmentally controlled process leading eventually to organ death. The breakdown and recycling of macromolecules from senescing leaves allow plants to reallocate resources to reproduction or new growth (
The transcription of CBP genes changes dramatically during leaf senescence. In an
The only known potential regulator of CBP genes during leaf senescence is
Besides
As described above, only a few regulators of carotenoid biosynthesis in green tissues, such as PIF1 and HY5, have been well characterized and shown to directly regulate
The ripening developmental program of fleshy fruits involves changes in texture (alteration of cell wall composition, reduction in turgor pressure), flavor and aroma (alteration of volatiles, sugar/starch, and acid metabolism), and color (alteration of chlorophyll, carotenoid, and flavonoid content) (
The foremost model for carotenoid regulation during fruit ripening is tomato (
Transcriptional regulation of CBP genes in tomato fruits: “ripening quartet”–related proteins.
Transcriptional regulation of CBP genes in tomato fruits: other proteins. The regulation of CBP genes in tomato fruits. The carotenoid biosynthesis pathway is shown in black, with carotenoid biosynthesis genes indicated in dark blue. Carotenoid regulators discussed in the paper are shown in light blue. Green arrows indicate positive regulation, while blunt red arrows indicate negative regulation. Solid lines show direct interactions, while dotted lines show indirect interactions.
Several MADS-box ripening regulators affect the expression of CBP genes in tomatoes, and a ripening model similar to the floral quartet model has been proposed (
These MADS-box proteins exert their effects over CBP gene transcription both directly by binding the promoters of some genes and indirectly by unknown mechanisms. Various studies have shown the promoter of
It should be mentioned that ChIP studies assessing RIN binding to target gene promoters have produced inconsistent results. Some studies have shown that the
Other CBP-regulating MADS-box genes in tomato that interact with or regulate the ripening quartet are
Many other regulators play a role in CBP gene regulation during fruit ripening, especially those involved in hormone synthesis and signaling. SlAP2a (an APETALA2/ERF protein) positively regulates fruit ripening, promoting the expression of
NAC family transcription factors involved in ethylene biosynthesis also affect the transcription of CBP genes: SlNAC4 positively regulates
Additionally, overexpression of
Besides ethylene and ABA, other plant hormones are also involved in tomato fruit ripening, with complex actions and interactions. Auxin acts antagonistically to ethylene, delaying ripening. Exogenous application of an auxin inhibitor to tomato fruits produces an effect similar to ethylene application, indicating that perhaps the presence vs. absence of auxin, not ethylene
Brassinosteroid (BR) application to pericarp discs induces lycopene formation (
Light is also an important regulator of CBP genes during tomato fruit development. Interestingly, the shading response seen in
Other light signaling components have been examined in tomato fruits. Mutations in the high pigment genes
Another light-responsive CBP regulator in tomato is
Other proteins appear to indirectly affect the transcription of CBP genes through plastid processes. Overexpression of the B-box protein BBX20 increases the chlorophyll and carotenoid content in tomato leaves and fruits, inducing
The Clp protease
Epigenetic regulation is also crucial to fruit ripening and carotenoid biosynthesis in tomato.
Colorless non-ripening (Cnr) tomato mutants do not express
Another epigenetic regulator, the tomato histone deacetylase gene
Putative transcriptional regulators have also been identified in other climacteric fruits (
Transcriptional regulation of CBP genes in other fruits. The regulation of CBP genes in citrus, peach, papaya, and orange kiwi. The carotenoid biosynthesis pathway is shown in black, with carotenoid biosynthesis genes indicated in dark blue. Carotenoid regulators discussed in the paper are shown in light blue. Green arrows indicate positive regulation, while blunt red arrows indicate negative regulation. Solid lines show direct interactions, while dotted lines show indirect/unknown interactions.
In kiwifruit (
Watermelon (
In citrus, a yeast one-hybrid screen using the promoters of
The coordinated transcriptional regulation of CBP genes is largely responsible for the coloration of carotenoid-pigmented flowers (e.g.,
Transcriptional regulation of CBP genes in flowers. The regulation of CBP genes in
The F-box protein CORONATINE INSENSITIVE 1 (COI1) is necessary for the perception of jasmonic acid JA. In addition to its many other functions, COI1-mediated JA signaling has been implicated in the production of floral and extrafloral nectar. Silencing of
In the monkeyflower species
Seed carotenoids are critical for ABA biosynthesis and seed dormancy, as well as protecting seeds from ROS damage. Therefore, carotenoids contribute to successful germination (
Very few transcriptional regulators of seed carotenoid biosynthesis have been identified (
Transcriptional regulation of CBP genes in roots. The regulation of CBP genes in root tissues. Icons indicate the species (carrot, cassava, maize, rice, and
It is perhaps surprising that so little is known about CBP transcriptional regulation in seeds, given the developmental and economic importance of seed carotenoids. This may be because many carotenoid-containing seeds primarily accumulate lutein (e.g., wheat, maize, millet, sunflower, pumpkin, canola), and the regulation of the α-carotene branch of the pathway is little understood (
Although most roots do not produce carotenoids in appreciable amounts, the CBP is active to provide the precursors for ABA biosynthesis (
Because roots are responsible for water and nutrient acquisition, root tissues must be able to respond to environmental cues. Of particular relevance to carotenoid biosynthesis is the sensing of and response to drought and salt stress (
In cassava (
In
It appears that the transcriptional regulation of
In carrots (
In
A fairly large number of putative transcriptional regulators of carotenoid biosynthesis have been identified from various species and tissue types (
The endogenous functions of some of these putative transcriptional regulators have not been verified through knockout or knockdown experiments (e.g.,
Most putative regulators were identified from ripening fruits, especially tomato (e.g.,
Current major model systems for carotenoid regulation are somewhat unusual or at least not representative. For example, the best genetic model system,
Minimal effort has been put into testing whether the function of a certain regulator identified from one species is conserved in another species. So far, only the PIF1/HY5 regulatory module has been shown to play a role in carotenoid biosynthesis during both
In addition to these challenges, there are also many gaps in our understanding of transcriptional regulation of carotenoid biosynthesis. For example, we know very little about what regulates most CBP genes downstream of
The challenges and knowledge gaps discussed above present wonderful opportunities for future carotenoid research. We think the following research directions will be fruitful in understanding how carotenoid biosynthesis is controlled at the transcriptional level:
Testing the function of known putative carotenoid regulators (
Identifying regulators of late pathway CBP genes using promoter screens. The recent study in maize (
Discovering key CREs of the CBP genes in various species. Databases such as PLACE (
Integrating multi-omics data (genomics, transcriptomics, proteomics, metabolomics, etc.) towards a more comprehensive understanding of CBP gene expression. With the rapid advances in generating large quantities of high-quality data as well as sophisticated bioinformatics methods and analytical tools, the systems biology approach will allow us to uncover correlations between metabolome and transcriptome profiles, to identify candidate transcriptional regulators of biosynthetic genes in co-expression modules, and to map regulatory network interactions (e.g.,
Broadening the diversity of “model” systems. For example, citrus would be an excellent system to complement the existing tomato fruit model because it is non-climacteric and accumulates various carotenoids beyond lycopene. In addition, carotenoid-containing, embryogenic citrus calli can be readily produced and transformed (
We believe that these research avenues will lead to many more exciting discoveries in the coming years, which will not only contribute new knowledge on the transcriptional regulation of carotenoid biosynthesis but also likely have a significant impact on carotenoid biofortification of crop plants. So far, most of the efforts towards enhancing carotenoid biosynthesis or engineering novel carotenoid products in staple crops have focused on CBP genes (e.g., aSTARice;
LS and Y-WY wrote the manuscript.
Our work on carotenoids is supported by the National Science Foundation (IOS-1558083, IOS-1827645).
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
We thank members of our laboratory and Drs. Foen Peng, Pam Diggle, Jeff Seemann, and Qinlong Zhu for discussions.
The Supplementary Material for this article can be found online at: