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Enamul Huq

Publications and source records attributed to Enamul Huq.

10 recordsLinked to original sources
PubMed2006

A new CULLIN 1 mutant has altered responses to hormones and light in Arabidopsis.

Regulated protein degradation contributes to plant development by mediating signaling events in many hormone, light, and developmental pathways. Ubiquitin ligases recognize and ubiquitinate target proteins for subsequent degradation by the 26S proteasome. The multisubunit SCF is the best-studied class of ubiquitin ligases in Arabidopsis (Arabidopsis thaliana). However, the extent of SCF participation in signaling networks is unclear. SCFs are composed of four subunits: CULLIN 1 (CUL1), ASK, RBX1, and an F-box protein. Null mutations in CUL1 are embryo lethal, limiting insight into the role of CUL1 and SCFs in later stages of development. Here, we describe a viable and fertile weak allele of CUL1, called cul1-6. cul1-6 plants have defects in seedling and adult morphology. In addition to reduced auxin sensitivity, cul1-6 seedlings are hyposensitive to ethylene, red, and blue light conditions. An analysis of protein interactions with the cul1-6 gene product suggests that both RUB (related to ubiquitin) modification and interaction with the SCF regulatory protein CAND1 (cullin associated and neddylation dissociated) are disrupted. These findings suggest that the morphological defects observed in cul1-6 plants are caused by defective SCF complex formation. Characterization of weak cul1 mutants provides insight into the role of SCFs throughout plant growth and development.

Arabidopsis↗
PubMed2005

Degradation of negative regulators: a common theme in hormone and light signaling networks?

Signal transduction pathways often modulate both positively and negatively acting components to optimize the efficiency of a signal. Recent results have shown that plants make extensive use of regulated proteolysis to modulate signal transduction pathways. An emerging theme from hormone (e.g. auxin and gibberellin) and light signaling pathways is signal or stimulus-induced degradation of negative regulators to optimize plant growth and development.

Light↗
PubMed2005

PIF1 is regulated by light-mediated degradation through the ubiquitin-26S proteasome pathway to optimize photomorphogenesis of seedlings in Arabidopsis.

Light signals perceived by the phytochrome (phy) family of sensory photoreceptors control multiple aspects of plant development. Recently, PIF1, a phy-interacting basic helix-loop-helix (bHLH) transcription factor, has been shown to negatively regulate facets of the photomorphogenesis of seedlings. Moreover, the transcriptional activation activity of PIF1 is reduced in a phy-dependent manner. In this study we use the luciferase (LUC) activity of the LUC-PIF1 fusion protein as an indicator of the stability of PIF1 in various light conditions. We found that the activity of LUC-PIF1 in both transient and stable transgenic lines is rapidly reduced in light, while the LUC-only control is stable under the same conditions, suggesting that PIF1 is degraded in response to light. Fluence-rate response curves indicate that PIF1 degradation is very sensitive to the quality and quantity of light. The half-life of PIF1 is about 16 min under 10 micromol m-2 sec-1 red light. PIF1 reaccumulates in the subsequent dark period after light-induced degradation, signifying that PIF1 not only functions in the dark and during the transition from etiolated to de-etiolated growth, but may also function during diurnal cycles. Inhibitors of the 26S proteasome increased the stability of PIF1, indicating that degradation of PIF1 is mediated by the ubiquitin-26S proteasome pathway. Further, de novo protein synthesis is not required for degradation of PIF1, as the presence of cycloheximide does not prevent degradation of PIF1 in the light. Taken together, these results suggest that the light signals perceived by phys induce the degradation of PIF1 and other phy-interacting factors to optimize photomorphogenesis.

Arabidopsis↗
PubMed2004

A novel molecular recognition motif necessary for targeting photoactivated phytochrome signaling to specific basic helix-loop-helix transcription factors.

The phytochrome (phy) family of sensory photoreceptors (phyA to phyE) in Arabidopsis thaliana control plant developmental transitions in response to informational light signals throughout the life cycle. The photoactivated conformer of the photoreceptor Pfr has been shown to translocate into the nucleus where it induces changes in gene expression by an unknown mechanism. Here, we have identified two basic helix-loop-helix (bHLH) transcription factors, designated PHYTOCHROME-INTERACTING FACTOR5 (PIF5) and PIF6, which interact specifically with the Pfr form of phyB. These two factors cluster tightly with PIF3 and two other phy-interacting bHLH proteins in a phylogenetic subfamily within the large Arabidopsis bHLH (AtbHLH) family. We have identified a novel sequence motif (designated the active phytochrome binding [APB] motif) that is conserved in these phy-interacting AtbHLHs but not in other noninteractors. Using the isolated domain and site-directed mutagenesis, we have shown that this motif is both necessary and sufficient for binding to phyB. Transgenic expression of the native APB-containing AtbHLH protein, PIF4, in a pif4 null mutant, rescued the photoresponse defect in this mutant, whereas mutated PIF4 constructs with site-directed substitutions in conserved APB residues did not. These data indicate that the APB motif is necessary for PIF4 function in light-regulated seedling development and suggest that conformer-specific binding of phyB to PIF4 via the APB motif is necessary for this function in vivo. Binding assays with the isolated APB domain detected interaction with phyB, but none of the other four Arabidopsis phys. Collectively, the data suggest that the APB domain provides a phyB-specific recognition module within the AtbHLH family, thereby conferring photoreceptor target specificity on a subset of these transcription factors and, thus, the potential for selective signal channeling to segments of the transcriptional network.

Amino Acid Motifs↗
PubMed2004

Phytochrome-interacting factor 1 is a critical bHLH regulator of chlorophyll biosynthesis.

Photosynthetic organisms must achieve a delicate balance between the light energy absorbed by chlorophyll and their capacity to channel that energy into productive photochemical reactions. Release of excess absorbed energy in the cell can cause lethal photooxidative damage. We identified a basic helix-loop-helix (bHLH) transcription factor, designated PHYTOCHROME-INTERACTING FACTOR 1 (PIF1), that negatively regulates chlorophyll biosynthesis. pif1 mutant seedlings accumulate excess free protochlorophyllide when grown in the dark, with consequent lethal bleaching upon exposure to light. PIF1 interacts specifically with the photoactivated conformer of phytochromes A and B, suggesting a signaling pathway by which chlorophyll biosynthetic rates are tightly controlled during the critical initial emergence of seedlings from subterranean darkness into sunlight.

Arabidopsis↗
PubMed2003

Nuclear translocation of the photoreceptor phytochrome B is necessary for its biological function in seedling photomorphogenesis.

The phytochrome (phy) family of sensory photoreceptors (phyA to phyE in Arabidopsis) enables plants to optimize their growth and development under natural light environments. Subcellular localization studies have shown that the photoreceptor molecule is induced to translocate from cytosol to nucleus by light, but direct evidence of the functional relevance of this translocation has been lacking. Here, using a glucocorticoid receptor-based fusion protein system, we demonstrate that both photoactivation and nuclear translocation combined are necessary and sufficient for the biological function of phyB. Conversely, neither artificial nuclear translocation of non-photoactivated phyB nor artificial retention of photoactivated phyB in the cytosol provides detectable biological activity. Together these data indicate that signal transfer from photoactivated phyB to its primary signaling partner(s) is localized in the nucleus, and conversely suggest the absence of a cytosolic pathway from photoactivated phyB to light-responsive genes.

Arabidopsis↗
PubMed2003

The Arabidopsis basic/helix-loop-helix transcription factor family.

The basic/helix-loop-helix (bHLH) proteins are a superfamily of transcription factors that bind as dimers to specific DNA target sites and that have been well characterized in nonplant eukaryotes as important regulatory components in diverse biological processes. Based on evidence that the bHLH protein PIF3 is a direct phytochrome reaction partner in the photoreceptor's signaling network, we have undertaken a comprehensive computational analysis of the Arabidopsis genome sequence databases to define the scope and features of the bHLH family. Using a set of criteria derived from a previously defined consensus motif, we identified 147 bHLH protein-encoding genes, making this one of the largest transcription factor families in Arabidopsis. Phylogenetic analysis of the bHLH domain sequences permits classification of these genes into 21 subfamilies. The evolutionary and potential functional relationships implied by this analysis are supported by other criteria, including the chromosomal distribution of these genes relative to duplicated genome segments, the conservation of variant exon/intron structural patterns, and the predicted DNA binding activities within subfamilies. Considerable diversity in DNA binding site specificity among family members is predicted, and marked divergence in protein sequence outside of the conserved bHLH domain is observed. Together with the established propensity of bHLH factors to engage in varying degrees of homodimerization and heterodimerization, these observations suggest that the Arabidopsis bHLH proteins have the potential to participate in an extensive set of combinatorial interactions, endowing them with the capacity to be involved in the regulation of a multiplicity of transcriptional programs. We provide evidence from yeast two-hybrid and in vitro binding assays that two related phytochrome-interacting members in the Arabidopsis family, PIF3 and PIF4, can form both homodimers and heterodimers and that all three dimeric configurations can bind specifically to the G-box DNA sequence motif CACGTG. These data are consistent, in principle, with the operation of this combinatorial mechanism in Arabidopsis.

Amino Acid Sequence↗
PubMed2002

A light-switchable gene promoter system.

Regulatable transgene systems providing easily controlled, conditional induction or repression of expression are indispensable tools in biomedical and agricultural research and biotechnology. Several such systems have been developed for eukaryotes. Most of these rely on the administration of either exogenous chemicals or heat shock. Despite the general success of many of these systems, the potential for problems, such as toxic, unintended, or pleiotropic effects of the inducing chemical or treatment, can impose limitations on their use. We have developed a promoter system that can be induced, rapidly and reversibly, by short pulses of light. This system is based on the known red light-induced binding of the plant photoreceptor phytochrome to the protein PIF3 and the reversal of this binding by far-red light. We show here that yeast cells expressing two chimeric proteins, a phytochrome-GAL4-DNA-binding-domain fusion and a PIF3-GAL4-activation-domain fusion, are induced by red light to express selectable or "scorable" marker genes containing promoters with a GAL4 DNA-binding site, and that this induction is rapidly abrogated by subsequent far-red light. We further show that the extent of induction can be controlled precisely by titration of the number of photons delivered to the cells by the light pulse. Thus, this system has the potential to provide rapid, noninvasive, switchable control of the expression of a desired gene to a preselected level in any suitable cell by simple exposure to a light signal.

Arabidopsis Proteins↗
PubMed2002

PIF4, a phytochrome-interacting bHLH factor, functions as a negative regulator of phytochrome B signaling in Arabidopsis.

Plants sense and respond to red and far-red light using the phytochrome (phy) family of photoreceptors. However, the mechanism of light signal transduction is not well defined. Here, we report the identification of a new mutant Arabidopsis locus, srl2 (short under red-light 2), which confers selective hypersensitivity to continuous red, but not far-red, light. This hypersensitivity is eliminated in srl2phyB, but not srl2phyA, double mutants, indicating that this locus functions selectively and negatively in phyB signaling. The SRL2 gene encodes a bHLH factor, designated PIF4 (phytochrome-interacting factor 4), which binds selectively to the biologically active Pfr form of phyB, but has little affinity for phyA. Despite its hypersensitive morphological phenotype, the srl2 mutant displays no perturbation of light-induced expression of marker genes for chloroplast development. These data suggest that PIF4 may function specifically in a branch of the phyB signaling network that regulates a subset of genes involved in cell expansion. Consistent with this proposal, PIF4 localizes to the nucleus and can bind to a G-box DNA sequence motif found in various light-regulated promoters.

Amino Acid Sequence↗