The E3 Ubiquitin Ligase AFF1 targets ARF19 to the proteasome
Ontology highlight
ABSTRACT: Auxin critically regulates nearly every aspect of plant growth and development. Auxin-driven transcriptional responses are mediated through the AUXIN RESPONSE FACTOR (ARF) family of transcription factors. Although ARF protein stability is regulated via the 26S proteasome, molecular mechanisms underlying ARF stability and turnover are unknown. Here, we report the identification and functional characterization of an F-Box E3 ubiquitin ligase, which we have named AUXIN RESPONSE FACTOR F-BOX1 (AFF1). AFF1 directly interacts with ARF19 and regulates its accumulation. Mutants defective in AFF1 display ARF19 protein hyperaccumulation, mild auxin resistance, and developmental defects. Together, our data suggest a new mechanism, namely control of ARF protein stability, in regulating auxin responsiveness.
Project description:The phytohormone auxin plays crucial roles in nearly every aspect of plant growth and development. The AUXIN RESPONSE FACTOR (ARF) family of transcription factors regulates auxin-responsive gene expression and exhibit nuclear localization in regions of high auxin responsiveness. Here we show that activating ARF7 and ARF19 proteins accumulate in micron-sized assemblies within the cytoplasm of tissues with attenuated auxin responsiveness. The intrinsically disordered middle region and the folded PB1 interaction domain of ARFs drive protein assembly formation. Mutation of a single lysine within the PB1 domain abrogates cytoplasmic assemblies, promotes ARF nuclear localization, and results in an altered transcriptome and morphological defects. Our data suggest a model in which ARF nucleo-cytoplasmic partitioning regulates auxin responsiveness, thus providing a mechanism for cellular competence for auxin signaling.
Project description:RNA samples were extracted from liquid cultured seedlings treated with or without auxin (5µM IAA) for 2 h. Lines used for this study: Columbia wild-type nph4-1(arf7) single mutant arf19-1 single mutant nph4-1 arf19-1 double mutant Treatment: Control (EtOH) Auxin treated (5µM IAA) Keywords = Auxin Keywords = Auxin response factor Keywords: parallel sample
Project description:RNA samples were extracted from liquid cultured seedlings treated with or without auxin (5µM IAA) for 2 h. Lines used for this study: Columbia wild-type nph4-1(arf7) single mutant arf19-1 single mutant nph4-1 arf19-1 double mutant Treatment: Control (EtOH) Auxin treated (5µM IAA) Keywords = Auxin Keywords = Auxin response factor Keywords: parallel sample
Project description:Lateral roots (LRs) are formed post-embryonically and contribute to root architecture formation in vascular plants. LATERAL ORGAN BOUNDARIES-DOMAIN 16 (LBD16) is a key transcription factor to initiate LR formation. LBD16 functions downstream of AUXIN RESPONSE FACTOR 7 (ARF7) and ARF19, and overexpression of LBD16 partially restores LR formation in the absence of ARF7 and ARF19. To identify downstream targets of LBD16, we engineered a transgenic line with inducible LBD16 activity by overexpressing a fusion protein of LBD16 and rat glucocorticoid receptor (GR) in arf7 arf19 mutant. Here we identified primary response genes of LBD16 from transcriptome analysis of 35Spro:LBD16:GR arf7 arf19 line.
Project description:Arabidopsis thaliana AF7/ARF19 double knockout with ARF7 reintroduced under its own promotor with a glucocorticoid receptor added were treated with Auxin, Dexamethazone and cycloheximide to determine primary and secondary ARF7 auxin sensitive downstream targets
Project description:The P-TEFb-containing super elongation complex (SEC) plays the essential role in transcriptional elongation control. The AF4/FMR2 family members AFF1 and AFF4 are the central scaffold proteins of SEC, associated with different human diseases. However, their specific roles in transcriptional control remain unclear. Here, we report that AFF1 and AFF4 show distinct genomic distribution patterns around TSS. AFF1 binds upstream of TSS, while AFF4 is enriched downstream of TSS. Pol II occupancies are reduced genome-widely after depletion of AFF1, but not AFF4. Interestingly, in a subset of active genes with broad AFF4 binding signature, AFF4 disruption causes slow elongation and early termination, while AFF1 deletion mirrors the transcriptional defects observed in the fast Pol II mutant. Furthermore, AFF4 knockdown leads to increased AFF1 levels at chromatin, and vice versa. In summary, our data demonstrate that AFF1 and AFF4 function, to some extent, antagonistically to ensure proper Pol II transcription.
Project description:Up-regulation of the neuropeptide NTS in a subgroup of lung cancers has been linked to poor prognosis. However, the regulatory pathway centered on NTS in lung cancer remains unclear. Here we identified the NTS specific enhancer in lung adenocarcinoma cells. The AF4/FMR2 (AFF) family protein AFF1 occupies the NTS enhancer and inhibits NTS transcription. Clustering analysis of lung adenocarcinoma gene expression data demonstrated that NTS is highly positively correlated with the expression of the oncogenic factor CPS1. Detailed analyses demonstrated that NTS antagonizes the IL6 pathway in regulating CPS1. Thus, our analyses revealed a novel NTS centered regulatory axis, consisting of AFF1 as a master transcription suppressor and IL6 as an antagonist in lung adenocarcinoma cells.
Project description:The P-TEFb-containing super elongation complex (SEC) plays the essential role in transcriptional elongation control. The AF4/FMR2 family members AFF1 and AFF4 are the central scaffold proteins of SEC, associated with different human diseases. However, their specific roles in transcriptional control remain unclear. Here, we report that AFF1 and AFF4 show distinct genomic distribution patterns around TSS. AFF1 binds upstream of TSS, while AFF4 is enriched downstream of TSS. Pol II occupancies are reduced genome-widely after depletion of AFF1, but not AFF4. Interestingly, in a subset of active genes with broad AFF4 binding signature, AFF4 disruption causes slow elongation and early termination, while AFF1 deletion mirrors the transcriptional defects observed in the fast Pol II mutant. Furthermore, AFF4 knockdown leads to increased AFF1 levels at chromatin, and vice versa. In summary, our data demonstrate that AFF1 and AFF4 function, to some extent, antagonistically to ensure proper Pol II transcription.
Project description:The P-TEFb-containing super elongation complex (SEC) plays the essential role in transcriptional elongation control. The AF4/FMR2 family members AFF1 and AFF4 are the central scaffold proteins of SEC, associated with different human diseases. However, their specific roles in transcriptional control remain unclear. Here, we report that AFF1 and AFF4 show distinct genomic distribution patterns around TSS. AFF1 binds upstream of TSS, while AFF4 is enriched downstream of TSS. Pol II occupancies are reduced genome-widely after depletion of AFF1, but not AFF4. Interestingly, in a subset of active genes with broad AFF4 binding signature, AFF4 disruption causes slow elongation and early termination, while AFF1 deletion mirrors the transcriptional defects observed in the fast Pol II mutant. Furthermore, AFF4 knockdown leads to increased AFF1 levels at chromatin, and vice versa. In summary, our data demonstrate that AFF1 and AFF4 function, to some extent, antagonistically to ensure proper Pol II transcription.