Gibberellic acid (GA) promotes seed germination, elongation growth, and flowering time in plants. GA responses are repressed by DELLA proteins, which contain an N-terminal DELLA domain essential for GA-dependent proteasomal degradation of DELLA repressors. Mutations of or within the DELLA domain of DELLA repressors have been described for species including Arabidopsis thaliana, wheat (Triticum aestivum), maize (Zea mays), and barley (Hordeum vulgare), and we show that these mutations confer GA insensitivity when introduced into the Arabidopsis GA INSENSITIVE (GAI) DELLA repressor. We also demonstrate that Arabidopsis mutants lacking the three GA INSENSITIVE DWARF1 (GID1) GA receptor genes are GA insensitive with respect to GA-promoted growth responses, GA-promoted DELLA repressor degradation, and GA-regulated gene expression. Our genetic interaction studies indicate that GAI and its close homolog REPRESSOR OF ga1-3 are the major growth repressors in a GA receptor mutant background. We further demonstrate that the GA insensitivity of the GAI DELLA domain mutants is explained in all cases by the inability of the mutant proteins to interact with the GID1A GA receptor. Since we found that the GAI DELLA domain alone can mediate GA-dependent GID1A interactions, we propose that the DELLA domain functions as a receiver domain for activated GA receptors.
Brassinosteroids (BRs) are plant steroid hormones that control many aspects of plant growth and development. BRs are perceived at the cell-surface by the plasma membrane-localized receptor complex composed of the receptor kinase BRI1 and its co-receptor BAK1. Here we show that BRI1 is post-translationally modified by K63 polyubiquitin chains in vivo. Artificially ubiquitinated BRI1 is recognized at the trans-Golgi Network/Early Endosomes (TGN/EE) and rapidly routed for vacuolar degradation. Mass spectrometry analyses identified residue K866 as an in vivo ubiquitination target in BRI1 involved in the negative regulation of BRI1. Model prediction revealed several redundant ubiquitination sites required for the endosomal sorting and vacuolar targeting of BRI1. Using total internal reflection fluorescence microscopy (TIRF), we also uncovered a role for BRI1 ubiquitination in promoting internalization from the cell-surface. Finally, we demonstrate that the control of BRI1 protein dynamics by ubiquitination is a fundamental control mechanism for BR responses in plants. Altogether, our results identify K63-linked polyubiquitin chain formation as a dual targeting signal for BRI1 internalization and sorting along the endocytic pathway, and highlight its role in hormonally controlled plant development.
The COP9 signalosome (CSN) was originally identified based on the constitutively photomorphogenic/de-etiolated/fusca (cop/det/fus) mutants from Arabidopsis thaliana. CSN is evolutionary conserved, and its subunit 5 (CSN5) mediates the deconjugation of NEDD8 from the cullin subunit of E3 ubiquitin ligases (deneddylation). Here, we report on Arabidopsis mutants deficient in CSN5 function. We show that these mutants are phenotypically indistinguishable from the previously described cop/det/fus mutants of other CSN subunits. However, we also show that these mutants retain the CSN complex (lacking CSN5), and this finding is in contrast with the previously described CSN subunit mutants, which lack the CSN complex. We therefore conclude that loss of CSN5 as part of CSN is sufficient to cause the cop/det/fus mutant phenotype. Furthermore, we show that mutants defective in CSN5 as well as mutants defective in CSN are unable to deneddylate the Arabidopsis cullins AtCUL1, AtCUL3A, and AtCUL4. Because these are representative cullin subunits of the three cullincontaining E3 families present in Arabidopsis, we postulate that the cop/det/fus mutant phenotype may be the result of the defects caused by impaired CSN5-dependent deneddylation of cullin-containing E3s.
The COP9 signalosome (CSN) is required for the full activity of cullin-RING E3 ubiquitin ligases (CRLs) in eukaryotes. CSN exerts its function on CRLs by removing the ubiquitin-related NEDD8 conjugate from the cullin subunit of CRLs. CSN seems, thereby, to control CRL disassembly or CRL subunit stability. In Arabidopsis thaliana, loss of CSN function leads to constitutive photomorphogenic (cop) seedling development and a post-germination growth arrest. The underlying molecular cause of this growth arrest is currently unknown. Here, we show that Arabidopsis csn mutants are delayed in G2 phase progression. This cell cycle arrest correlates with the induction of the DNA damage response pathway and is suggestive of the activation of a DNA damage checkpoint. In support of this hypothesis, we detected gene conversion events in csn mutants that are indicative of DNA doublestrand breaks. DNA damage is also apparent in mutants of the NEDD8 conjugation pathway and in mutants of the E3 ligase subunits CULLIN4, COP1 and DET1, which share phenotypes with csn mutants. In summary, our data suggest that Arabidopsis csn mutants undergo DNA damage, which might be the cause of the delay in G2 cell cycle progression.KEY WORDS: COP9 signalosome, Cell cycle, DNA damage Development 135, 2013Development 135, -2022Development 135, (2008 (Miséra et al., 1994; Chory et al., 1989;Deng et al., 1991). COP1 is a RING-type E3 ubiquitin ligase that mediates the degradation of several positive photomorphogenesis regulators (Osterlund et al., 2000;Seo et al., 2003;Seo et al., 2004). The human COP1 ortholog (RFWD2) has been implicated in c-JUN degradation and in DNA damage response following irradiation Wertz et al., 2004). Human COP1 is inactivated in response to DNA damage by ATM-dependent phosphorylation, and the consequent stabilization of its degradation target p53 induces a G1 cell cycle arrest . The function of DET1 only became clear when it was recognized that it is a subunit of a CULLIN4 (CUL4)-containing CRL, designated DCX DET1COP1 or CUL4-DDB1 DET1COP1 , which also includes COP1 and the adaptor subunit DAMAGED DNA-BINDING PROTEIN1 (DDB1) (Benvenuto et al., 2002;Dornan et al., 2004;Wertz et al., 2004;Yanagawa et al., 2004;Chen et al., 2006). Although these findings suggest that COP1 and DET1 function together in a CUL4-containing CRL, the COP1 monomer alone has in vitro E3 ligase activity. It is therefore presently unclear which functions of COP1 require the E3 complex (and DET1) and which functions are mediated by COP1 alone.Arabidopsis csn mutants arrest growth at the seedling stage. The underlying molecular cause of this growth arrest remains to be identified. Here we show that csn mutant cells have a delay in G2 phase progression. This delay correlates with the activation of the DNA damage response pathway but is not exclusively induced by the DNA damage signaling kinases ATAXIA TELANGIECTASIA MUTATED (ATM) or WEE1. Our observation that gene conversion events can occur in csn mutants strongly argues that DNA doublestrand break...
In the Arabidopsis root meristem, polar auxin transport creates a transcriptional auxin response gradient that peaks at the stem cell niche and gradually decreases as stem cell daughters divide and differentiate [1-3]. The amplitude and extent of this gradient are essential for both stem cell maintenance and root meristem growth [4, 5]. To investigate why expression of some auxin-responsive genes, such as the essential root meristem growth regulator BREVIS RADIX (BRX) [6], deviates from this gradient, we combined experimental and computational approaches. We created cellular-level root meristem models that accurately reproduce distribution of nuclear auxin activity and allow dynamic modeling of regulatory processes to guide experimentation. Expression profiles deviating from the auxin gradient could only be modeled after intersection of auxin activity with the observed differential endocytosis pattern and positive autoregulatory feedback through plasma-membrane-to-nucleus transfer of BRX. Because BRX is required for expression of certain auxin response factor targets, our data suggest a cell-type-specific endocytosis-dependent input into transcriptional auxin perception. This input sustains expression of a subset of auxin-responsive genes across the root meristem's division and transition zones and is essential for meristem growth. Thus, the endocytosis pattern provides specific positional information to modulate auxin response.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
customersupport@researchsolutions.com
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
This site is protected by reCAPTCHA and the Google Privacy Policy and Terms of Service apply.
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.