A functionally required unfoldome from the plant kingdom: intrinsically disordered N-terminal domains of GRAS proteins are involved in molecular recognition during plant development
The intrinsic disorder is highly abundant in eukaryotic genomes. In the animal kingdom, numerous intrinsically disordered proteins (IDPs) have been characterized, especially in cell signalling and transcription regulation. An intrinsically disordered region often folds in different structures allowing an IDP to recognize and bind different partners at various binding interfaces. In contrast, there have only been a few reports of IDPs from the plant kingdom. Plant-specific GRAS proteins play critical and diverse roles in plant development and signalling and often act as integrators of signals from multiple plant growth regulatory inputs. Using computational and bioinformatics tools, we demonstrate here that the GRAS proteins are intrinsically disordered, thus forming the first functionally required unfoldome in the plant kingdom. Furthermore, the N-terminal domains of GRAS proteins are predicted to contain numerous Molecular Recognition Features (MoRFs), short interaction-prone segments that are located within extended disorder regions and are able to recognize their interacting partners and to undergo disorder-to-order transitions upon binding to these specific partners. Overlapping with the relatively conserved motifs in the N-terminal domains of GRAS proteins, these predicted MoRFs represent the potential protein–protein binding sites and may be involved in molecular recognition during plant development. This study enables us to propose a conceptual framework that guides future experimental approaches to understand structure–function relationships of the entire GRAS family.

title: GRAS proteins: the versatile roles of intrinsically disordered proteins in plant signalling.
abstract: IDPs (intrinsically disordered proteins) are highly abundant in eukaryotic proteomes and important for cellular functions, especially in cell signalling and transcriptional regulation. An IDR (intrinsically disordered region) within an IDP often undergoes disorder-to-order transitions upon binding to various partners, allowing an IDP to recognize and bind different partners at various binding interfaces. Plant-specific GRAS proteins play critical and diverse roles in plant development and signalling, and act as integrators of signals from multiple plant growth regulatory and environmental inputs. Possessing an intrinsically disordered N-terminal domain, the GRAS proteins constitute the first functionally required unfoldome from the plant kingdom. Furthermore, the N-terminal domains of GRAS proteins contain MoRFs (molecular recognition features), short interaction-prone segments that are located within IDRs and are able to recognize their interacting partners by undergoing disorder-to-order transitions upon binding to these specific partners. These MoRFs represent potential protein-protein binding sites and may be acting as molecular bait in recognition events during plant development. Intrinsic disorder provides GRAS proteins with a degree of binding plasticity that may be linked to their functional versatility. As an overview of structure-function relationships for GRAS proteins, the present review covers the main biological functions of the GRAS family, the IDRs within these proteins and their implications for understanding mode-of-action.

title: N-terminal domains of DELLA proteins are intrinsically unstructured in the absence of interaction with GID1/gibberellic acid receptors.
abstract: The plant growth-repressing DELLA proteins (DELLAs) are known to represent a convergence point in integration of multiple developmental and environmental signals in planta, one of which is hormone gibberellic acid (GA). Binding of the liganded GA receptor (GID1/GA) to the N-terminal domain of DELLAs is required for GA-induced degradation of DELLAs via the ubiquitin-proteasome pathway, thus derepressing plant growth. However, the conformational changes of DELLAs upon binding to GID1/GA, which are the key to understanding the precise mechanism of GID1/GA-mediated degradation of DELLAs, remain unclear. Using biophysical, biochemical, and bioinformatics approaches, we demonstrated for the first time that the unbound N-terminal domains of DELLAs are intrinsically unstructured proteins under physiological conditions. Within the intrinsically disordered N-terminal domain of DELLAs, we have identified several molecular recognition features, sequences known to undergo disorder-to-order transitions upon binding to interacting proteins in intrinsically unstructured proteins. In accordance with the molecular recognition feature analyses, we have observed the binding-induced folding of N-terminal domains of DELLAs upon interaction with AtGID1/GA. Our results also indicate that DELLA proteins can be divided into two subgroups in terms of their molecular compactness and their interactions with monoclonal antibodies.

title: Multifarious Roles of Intrinsic Disorder in Proteins Illustrate Its Broad Impact on Plant Biology
abstract: Intrinsically disordered proteins (IDPs) are highly abundant in eukaryotic proteomes. Plant IDPs play critical roles in plant biology and often act as integrators of signals from multiple plant regulatory and environmental inputs. Binding promiscuity and plasticity allow IDPs to interact with multiple partners in protein interaction networks and provide important functional advantages in molecular recognition through transient protein–protein interactions. Short interaction-prone segments within IDPs, termed molecular recognition features, represent potential binding sites that can undergo disorder-to-order transition upon binding to their partners. In this review, we summarize the evidence for the importance of IDPs in plant biology and evaluate the functions associated with intrinsic disorder in five different types of plant protein families experimentally confirmed as IDPs. Functional studies of these proteins illustrate the broad impact of disorder on many areas of plant biology, including abiotic stress, transcriptional regulation, light perception, and development. Based on the roles of disorder in the protein–protein interactions, we propose various modes of action for plant IDPs that may provide insight for future experimental approaches aimed at understanding the molecular basis of protein function within important plant pathways.

title: A facile and efficient strategy for one-step in situ preparation of hydrophobic organic monolithic stationary phases by click chemistry and its application on protein separation.
abstract: A simple one-step in situ "click" modification strategy was developed for the preparation of hydrophobic organic monolithic columns for the first time. The column morphology and surface chemistry of the fabricated monolithic columns were characterized by scanning electron microscopy, Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy, respectively. The chromatographic performances of the C8/C18 "click" monoliths were evaluated through the separation of a mixture of five proteins such as ribonuclease A, soybean trypsin inhibitor, cytochrome c, bovine haemoglobin and bovine serum albumin. Compared with the blank column, the higher hydrophobicity stationary phases obtained from the "clicked" modification have longer retention times and higher resolution for the five proteins. The separation of five proteins mixture on click C18 monolith with gradient elution at different flow rates was also investigated, the baseline separation of five proteins could be achieved at three different flow rates.

title: Heterozygous deletion of Atbf1 by the Cre-loxP system in mice causes preweaning mortality.
abstract: ATBF1 is a large nuclear protein that contains multiple zinc-finger motifs and four homeodomains. In mammals, ATBF1 regulates differentiation, and its mutation and/or downregulation is involved in tumorigenesis in several organs. To gain more insight into the physiological functions of ATBF1, we generated and validated a conditional allele of mouse Atbf1 in which exons 7 and 8 were flanked by loxP sites (Atbf1(flox) ). Germline deletion of a single Atbf1 allele was achieved by breeding to EIIa-cre transgenic mice, and Atbf1 heterozygous mice displayed reduced body weight, preweaning mortality, increased cell proliferation, and attenuated cytokeratin 18 expression, indicating haploinsufficiency of Atbf1. Floxed Atbf1 mice will help us understand such biological processes as neuronal differentiation and tumorigenesis.

title: Meta-analysis of randomized trials for health care interventions: one for all?
abstract: Meta-analysis of randomized trials stands at the top of the epidemiological designs to test the effect of health care interventions. Ideally, it systematically and comprehensively collects the relevant randomized trials, and pools the trials to generate a single estimate of effects. However, the rationale for meta-analysis may not always be thoughtfully considered, and overlooking underlying principles may cause illusions. This article revisits the rationale for meta-analysis, discusses how meta-analysis can be misleading, and summarizes approaches to critically and judiciously interpret and use the results of meta-analysis.

title: Antisense HIF-1α prevents acquired tumor resistance to angiostatin gene therapy
abstract: Angiostatin is a naturally occurring inhibitor of angiogenesis that is being developed as a drug to fight cancer. In this study we reveal that EL-4 tumors established in mice rapidly develop resistance to angiostatin gene therapy by upregulating hypoxia-inducible pathways. Angiostatin initially delayed tumor growth for 6 days by reducing blood vessel density. However, tumors quickly responded by upregulating the production of hypoxia-inducible factor-1α (HIF-1α) and its effector vascular endothelial growth factor (VEGF) in response to increasing tumor hypoxia, leading to restored angiogenesis and rapid tumor growth. Theoretically, blockade of HIF-1 should prevent resistance to anti-angiogenic therapy by preventing a tumor from responding to induced hypoxia. Antisense HIF-1α inhibited the expression of HIF-1α and of the HIF-1 effectors VEGF, glucose transporter-1 and lactate dehydrogenase. As a monotherapy, it was effective in eradicating small 0.1 cm diameter tumors, but only delayed the growth of large 0.4 cm diameter tumors. In contrast, timed injection of a combination of angiostatin and antisense HIF-1α plasmids completely eradicated large EL-4 tumors within 2 weeks, and prevented upregulation of hypoxia-inducible pathways induced by angiostatin. The data indicate that blocking hypoxia-inducible pathways by antisense HIF-1α can circumvent hypoxia-induced drug resistance and thereby augment the efficacy of anti-angiogenic therapies.

title: Synthetic cannabinoids and potential reproductive consequences.
abstract: Increases in emergency room visits due to abuse of designer drugs, popularly known by the street names "K2" and "Spice," are a cause for social, judicial, and clinical concerns. The psychoactive components in these herbal drugs mainly consist of different synthetic cannabinoids, and users of these street drugs are primarily within the age group of 12 to 20years old. The abusive use of synthetic cannabinoids results in anxiety, nausea, vomiting, tachycardia, elevated blood pressure, tremors, seizures, hallucinations, and paranoid behavior, but the effects of maternal use of synthetic cannabinoids during pregnancy are ambiguous due to limited studies in humans and a relative short history of the drugs. In this review, we discuss the known and potential adverse effects of synthetic cannabinoids on human pregnancy using knowledge gathered from studies in mice and limited studies in humans. In mice, multiple sites and stages of pregnancy are potential targets of synthetic cannabinoids, including preimplantation embryo development, oviductal embryo transport, implantation, placentation, and parturition. It is anticipated that maternal use of synthetic cannabinoids would result in severely compromised female fertility and pregnancy outcome.