DELLA-dependent and -independent gibberellin signaling

ABSTRACT

DELLA proteins act as negative regulators in gibberellin (GA) signal transduction. GA-induced DELLA degradation is a central regulatory system in GA signaling pathway. Intensive studies have revealed the degradation mechanism of DELLA and the functions of DELLA as a transcriptional regulator. Meanwhile, recent studies suggest the existence of a DELLA-independent GA signaling pathway. In this review, we summarized the DELLA-independent GA signaling pathway together with the well-analyzed DELLA-dependent pathway.

Gibberellins (GAs) are phytohormones that control diverse aspects of plant growth and development, including seed germination, stem elongation, leaf expansion, and flower and seed development.¹ DELLA proteins act as growth repressors by inhibiting GA signaling in response to developmental and environmental cues.² Rice (Oryza sativa) contains one DELLA, SLENDER1, while Arabidopsis thaliana contains five DELLAs, GIBBERELLIN-INSENSITIVE (GAI), REPRESSOR OF ga1–3 (RGA), RGA-LIKE1 (RGL1), RGL2, and RGL3, which display partially overlapping but distinct functions in repressing GA responses.^3-7 SLEEPY1 (SLY1) and GIBBERELLIN INSENSITIVE DWARF2 (GID2) are F-box proteins in Arabidopsis and rice, respectively.^8-10 Upon GA-binding to a soluble GA receptor, GIBBERELLIN INSENSITIVE DWARF1 (GID1), DELLAs are recruited to the SCF^SLY1/GID2 ubiquitin E3 ligase complex for polyubiquitination and is subsequently degraded by the 26S proteasome.^11-13 Proteolysis-independent regulations of DELLA have been also reported. GID1 overexpression in sly1 mutant partially rescues dwarf and infertility phenotypes without degradation of DELLAs suggesting the proteolysis-independent downregulation of DELLAs.¹⁴ A portion of DELLAs is conjugated to the small ubiquitin-like modifier (SUMO) protein.¹⁵ GA-independent interaction of SUMOylated DELLAs with GID1 causes an accumulation of non-SUMOylated DELLAs, restraining plant growth.

DELLAs modulate gene expression by interacting with transcription factors (reviewed in refs^16,17). For example, DELLAs interact with PHYTOCHROME INTERACTING FACTORs (PIFs) and BRASSINAZOLE RESISTANT1 (BZR1) to inhibit their DNA-binding activity, resulting in repression of hypocotyl elongation.^18-20 DELLAs also interact with jasmonic acid ZIM-domain proteins (JAZs), causing the release of MYC2 from JAZs.²¹ Moreover, DELLAs are recruited to the promoters by DNA-binding transcription factors, such as INDETERMINATE1 domain proteins (IDDs) and type-B ARABIDOPSIS RESPONSE REGULATORS (ARRs), and act as transcriptional coactivators.^22-26 SPINDLY (SPY) and its paralog, SECRET AGENT (SEC), affect the binding affinity of DELLA for PIFs and BZR1. SPY was identified in a genetic screen for negative regulators of GA signaling in Arabidopsis.²⁷ Loss-of-function spy alleles are able to germinate in the presence of GA biosynthesis inhibitors, and to partially rescue the dwarf phenotype of GA-deficient mutants. Based on sequence comparison, SPY and SEC were predicted to encode O-linked N-acetylglucosamine (O-GlcNAc) transferases.^28,29 Recent studies revealed that SPY and SEC act as an O-fucosyltransferase and an O-GlcNAc transferase of DELLA proteins, respectively.^30,31 O-fucosylation by SPY promotes the interaction of DELLA with PIFs and BZR1, whereas O-GlcNAcylation by SEC inhibits these interaction. In addition to these glycosylation, phosphorylation and dephosphorylation are reported to inhibit and promote the degradation of DELLA, respectively,^32,33 but it is unclear whether phosphorylation state affects O-fucosylation and O-GlcNAcylation of DELLA.

GA-induced degradation of DELLA proteins acts as a central regulatory switch for GA signal transduction. However, some studies suggest the existence of a DELLA-independent GA pathway. Fruit growth in Arabidopsis is partially determined by a DELLA-independent GA response, and this response requires both GID1-mediated GA reception and 26S proteasome activity.³⁴ A microarray study using the Arabidopsis quadruple-DELLA mutant, which lacks GAI, RGA, RGL1, and RGL2, suggested that some GA-regulated genes are not regulated by DELLAs.³⁵ Tomato (Solanum lycopersicum) has only one DELLA gene, PROCERA (PRO). RNA sequencing using the pro mutant suggests that 5% of all GA-regulated genes in tomato are DELLA independent.³⁶

Besides these studies, we also found a DELLA-independent GA pathway.³⁷ Ca²⁺ is a ubiquitous second messenger involved in signal transduction of various environmental and developmental stimuli in eukaryote.^38,39 One physiological response of plant cells to GAs is increase in cytosolic Ca²⁺ ([Ca²⁺]_cyt).⁴⁰ An increase in [Ca²⁺]_cyt was observed previously after several hours of GA application.⁴¹ We reexamined the effects of GAs on [Ca²⁺]_cyt using the Ca²⁺ sensor protein aequorin in Arabidopsis.³⁷ [Ca²⁺]_cyt increased within a few minutes of GA treatment, even in transgenic plants expressing a mutated degradation-resistant version of RGA (RGAΔ17) and in della pentuple mutant plants. These results suggested that the GA-induced increase in [Ca²⁺]_cyt occurs via a DELLA-independent pathway.

Ca²⁺-dependent protein kinases (CDPKs) are multifunctional Ser/Thr protein kinases possessing both a Ca²⁺-sensing function and kinase activity within a single gene product and found only in plants and some protozoans.^42-44 Tobacco (Nicotiana tabacum) NtCDPK1 is involved in GA feedback regulation through the phosphorylation of the transcription factor, REPRESSION OF SHOOT GROWTH (RSG).^45-47 The translocation of RSG from the nucleus to the cytoplasm is promoted by NtCDPK1.^47,48 We previously found that NtCDPK1 is phosphorylated in response to GAs in plants and autophosphorylated in vitro.^47,49,50 NtCDPK1 is possibly activated by the increase in [Ca²⁺]_cyt via a DELLA-independent GA pathway.

Unlike DELLA proteins, GID1 GA receptors are localized in both the nucleus and the cytoplasm.¹¹ Therefore, GID1 could be involved in GA-induced increase in [Ca²⁺]_cyt. GID1-GA complex in the cytoplasm could cause the activation of Ca²⁺ channels or promote the degradation of a Ca²⁺ channel repressor via the ubiquitin-proteasome pathway, similar to DELLAs, leading to the GA-induced increase in [Ca²⁺]_cyt. Another possibility is that an unidentified accessory GA receptor other than GID1 is involved in the GA-induced increase in [Ca²⁺]_cyt. Although GID1 plays essential roles in GA signaling, physiological studies suggest an alternative signaling pathway related to a membrane-localized GA receptor.^41,51 Therefore, determining the involvement of GID1 in the GA-induced increase in [Ca²⁺]_cyt is an important issue. In addition to GID1 receptors, SPY is also reported to be a nucleocytoplasmic protein. Nuclear-export signal-fused SPY complements the spy mutation, but nuclear-localization signal-fused SPY does not.⁵² This result suggests a DELLA-independent function of cytosolic SPY. It is unlikely that SPY is involved in GA reception, but GA-induced increase in [Ca²⁺]_cyt might possibly affect SPY activity in cytoplasm. The importance of DELLA-independent pathway in GA signaling is still largely unknown. Further studies of DELLA-dependent and -independent GA signaling will not only provide new insights into each signaling pathway but also promote better understanding of each other.

Funding Statement

This work was supported by the Ministry of Education, Culture, Sports, Science and Technology (24118004) Japan Society for the Promotion of Science (17K15143) Japan Society for the Promotion of Science (17K07449) Japan Society for the Promotion of Science (15H04392).

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.