Dorothee Staiger

Ultraviolet-B Radiation-Mediated Responses in Plants. Balancing Damage and Protection

Seven percent of the electromagnetic radiation emitted from the sun is in the UV range (200–400 nm). As it passes through the atmosphere, the total flux transmitted is greatly reduced, and the composition of the UV radiation is modified. Shortwave UV-C radiation (200–280 nm) is completely

A type III effector ADP-ribosylates RNA-binding proteins and quells plant immunity

The bacterial plant pathogen Pseudomonas syringae injects effector proteins into host cells through a type III protein secretion system to cause disease. The enzymatic activities of most of P. syringae effectors and their targets remain obscure. Here we show that the type III effector HopU1 is a mono-ADP-ribosyltransferase (ADP-RT). HopU1 suppresses plant innate immunity in a manner dependent on its ADP-RT active site. The HopU1 substrates in Arabidopsis thaliana extracts were RNA-binding proteins that possess RNA-recognition motifs (RRMs). A. thaliana knockout lines defective in the glycine-rich RNA-binding protein GRP7 (also known as AtGRP7), a HopU1 substrate, were more susceptible than wild-type plants to P. syringae. The ADP-ribosylation of GRP7 by HopU1 required two arginines within the RRM, indicating that this modification may interfere with GRP7’s ability to bind RNA. Our results suggest a pathogenic strategy where the ADP-ribosylation of RNA-binding proteins quells host immunity by affecting RNA metabolism and the plant defence transcriptome.

Alternative Splicing at the Intersection of Biological Timing, Development, and Stress Responses

High-throughput sequencing for transcript profiling in plants has revealed that alternative splicing (AS) affects a much higher proportion of the transcriptome than was previously assumed. AS is involved in most plant processes and is particularly prevalent in plants exposed to environmental stress. The identification of mutations in predicted splicing factors and spliceosomal proteins that affect cell fate, the circadian clock, plant defense, and tolerance/sensitivity to abiotic stress all point to a fundamental role of splicing/AS in plant growth, development, and responses to external cues. Splicing factors affect the AS of multiple downstream target genes, thereby transferring signals to alter gene expression via splicing factor/AS networks. The last two to three years have seen an ever-increasing number of examples of functional AS. At a time when the identification of AS in individual genes and at a global level is exploding, this review aims to bring together such examples to illustrate the extent and importance of AS, which are not always obvious from individual publications. It also aims to ensure that plant scientists are aware that AS is likely to occur in the genes that they study and that dynamic changes in AS and its consequences need to be considered routinely.

Photoreceptors in Arabidopsis thaliana: light perception, signal transduction and entrainment of the endogenous clock

Abstract. To keep track of fluctuations in spectral composition and intensity of incoming sunlight, plants engage a plethora of photosensory pigments. Absorption of light by these photoreceptors sets in motion signaling cascades that ultimately influence the plants physiology. Many light-controlled processes are based on modulation of gene activity in response to changes in irradiation. The molecular basis of this regulation and the downstream components transducing signals from the photoreceptors are not fully understood yet, but recent evidence suggests that some of those routes are rather short. The phytochrome photoreceptors have been found to influence light-responsive promoters by direct contact with transcription factors. Additionally, the cryptochrome blue-light receptors directly interact with a key repressor of photomorphogenesis, suggesting that light activation of photoreceptors could initiate photomorphogenesis through posttranslational regulation. This review focuses on recent insights into photosensory transduction mechanisms as well as on our current understanding of light entrainment of the endogenous clock.

Reciprocal regulation of glycine-rich RNA-binding proteins via an interlocked feedback loop coupling alternative splicing to nonsense-mediated decay in Arabidopsis

The Arabidopsis RNA-binding protein AtGRP8 undergoes negative autoregulation at the post-transcriptional level. An elevated AtGRP8 protein level promotes the use of a cryptic 5′ splice site to generate an alternatively spliced transcript, as_AtGRP8, retaining the 5′ half of the intron with a premature termination codon. In mutants defective in nonsense-mediated decay (NMD) abundance of as_AtGRP8 but not its pre-mRNA is elevated, indicating that as_AtGRP8 is a direct NMD target, thus limiting the production of functional AtGRP8 protein. In addition to its own pre-mRNA, AtGRP8 negatively regulates the AtGRP7 transcript through promoting the formation of the equivalent alternatively spliced as_AtGRP7 transcript, leading to a decrease in AtGRP7 abundance. Recombinant AtGRP8 binds to its own and the AtGRP7 pre-mRNA, suggesting that this interaction is relevant for the splicing decision in vivo. AtGRP7 itself is part of a negative autoregulatory circuit that influences circadian oscillations of its own and the AtGRP8 transcript through alternative splicing linked to NMD. Thus, we identify an interlocked feedback loop through which two RNA-binding proteins autoregulate and reciprocally crossregulate by coupling unproductive splicing to NMD. A high degree of evolutionary sequence conservation in the introns retained in as_AtGRP8 or as_AtGRP7 points to an important function of these sequences.

The small glycine‐rich RNA binding protein AtGRP7 promotes floral transition in Arabidopsis thaliana

The RNA binding protein AtGRP7 is part of a circadian slave oscillator in Arabidopsis thaliana that negatively autoregulates its own mRNA, and affects the levels of other transcripts. Here, we identify a novel role for AtGRP7 as a flowering-time gene. An atgrp7-1 T-DNA mutant flowers later than wild-type plants under both long and short days, and independent RNA interference lines with reduced levels of AtGRP7, and the closely related AtGRP8 protein, are also late flowering, particularly in short photoperiods. Consistent with the retention of a photoperiodic response, the transcript encoding the key photoperiodic regulator CONSTANS oscillates with a similar pattern in atgrp7-1 and wild-type plants. In both the RNAi lines and in the atgrp7-1 mutant transcript levels for the floral repressor FLC are elevated. Conversely, in transgenic plants ectopically overexpressing AtGRP7, the transition to flowering is accelerated mainly in short days, with a concomitant reduction in FLC abundance. The late-flowering phenotype of the RNAi lines is suppressed by introducing the flc-3 loss-of-function mutation, suggesting that AtGRP7 promotes floral transition, at least partly by downregulating FLC. Furthermore, vernalization overrides the late-flowering phenotype. Retention of both the photoperiodic response and vernalization response are features of autonomous pathway mutants, suggesting that AtGRP7 is a novel member of the autonomous pathway.

T-DNA gene 5 of Agrobacterium modulates auxin response by autoregulated synthesis of a growth hormone antagonist in plants.

Oncogenes carried by the transferred DNA (T‐DNA) of Agrobacterium Ti plasmids encode the synthesis of plant growth factors, auxin and cytokinin, and induce tumour development in plants. Other T‐DNA genes regulate the tumorous growth in ways that are not yet understood. To determine the function of T‐DNA gene 5, its coding region was expressed in Escherichia coli. Synthesis of the gene 5 encoded protein (26 kDa) correlated with a 28‐fold increase in conversion of tryptophan to indole‐3‐lactate (ILA), an auxin analogue. Expression of chimeric gene 5 constructs in transgenic tobacco resulted in overproduction of ILA that enhanced shoot formation in undifferentiated tissues and increased the tolerance of germinating seedlings to the inhibitory effect of externally supplied auxin. Promoter analysis of gene 5 in plants revealed that its expression was inducible by auxin and confined to the vascular phloem cells. cis‐regulatory elements required for auxin regulation and phloem specific expression of gene 5 were mapped to a 90 bp promoter region that carried DNA sequence motifs common to several auxin induced plant promoters, as well as a binding site for a nuclear factor, Ax‐1. ILA was found to inhibit the auxin induction of the gene 5 promoter and to compete with indole‐3‐acetic acid (IAA) for in vitro binding to purified cellular auxin binding proteins. It is suggested therefore that ILA autoregulates its own synthesis and thereby modulates a number of auxin responses in plants.

Arabidopsis thaliana germin-like proteins: common and specific features point to a variety of functions.

Abstract. Germin-like proteins (GLPs) are ubiquitous plant proteins encoded by diverse multigene families. It is not known whether they share germins unusual biochemical properties and oxalate oxidase activity. Using specific antibodies, we have studied three GLPs (AtGER1, AtGER2 and AtGER3) in Arabidopsis thaliana (L.) Heynh. as well as in transgenic tobacco (Nicotiana tabacum L.) plants overexpressing these proteins. Like wheat (Triticum aestivum L.) germin, these Arabidopsis GLPs are associated with the extracellular matrix (ECM) and they also seem to exist as two glycosylated isoforms. However, none of them is an oxalate oxidase. Although GLPs display several conserved features, each has its specific characteristics. Both AtGER2 and AtGER3 are oligomeric proteins that share germins resistance to pepsin and to dissociation by heat and SDS. In contrast, AtGER1 seems to exist as a monomer. The GLPs may interact with the ECM in a variety of ways, since each is efficiently extracted by different conditions. In addition, germins and GLPs all bind Cibacron Blue, a dye often but not exclusively used for the purification of enzymes having nucleotide cofactors. In the case of AtGER2, binding to the dye is so tight that it almost allows a one-step purification of this protein. The variety of sequences, expression patterns and biochemical features indicates that GLPs could be a class of receptors localized in the ECM and involved in physiological and developmental processes as well as stress response.

Circadian Oscillations of a Transcript Encoding a Germin-Like Protein That Is Associated with Cell Walls in Young Leaves of the Long-Day Plant Sinapis alba L

As part of an attempt to analyze rhythmic phenomena in the long-day plant Sinapis alba L. at the molecular level, we have searched for mRNAs whose concentration varies as a function of time of day. Differential screening of a cDNA library established from mRNAs expressed at the end of the daily light phase with probes representing transcripts expressed predominantly in the morning or evening has identified one major transcript. The cDNA, Saglp, encodes a predicted 22-kD protein with an N-terminal signal sequence. The protein shows homology to germin, a protein expressed in wheat embryos after onset of germination. The Saglp mRNA level undergoes circadian oscillations in light/dark cycles with maxima between 8 and 12 PM (zeitgeber time [zt] 12-zt16) and minima around 8 AM (zt0). In plants grown from seed in constant light, transcript levels are constitutive. In constant light regular temperature shifts function as an alternative “zeitgeber” to initiate Saglp transcript oscillations. At the cellular level, Saglp transcripts are expressed in the epidermis and spongy parenchyma of young leaves, and in distinct regions of the epidermis and the cortex in stems and petioles. Strong signals are observed in these tissues around zt12, whereas little expression is found around zt20, suggesting that the underlying oscillatory mechanism(s) operate(s) synchronously in different plant organs. The SaGLP steady-state protein concentration remains constant over light/dark cycles. Immunogold labeling shows that the SaGLP protein is associated with primary cell walls.

Emerging role for RNA‐based regulation in plant immunity

Infection by phytopathogenic bacteria triggers massive changes in plant gene expression, which are thought to be mostly a result of transcriptional reprogramming. However, evidence is accumulating that plants additionally use post-transcriptional regulation of immune-responsive mRNAs as a strategic weapon to shape the defense-related transcriptome. Cellular RNA-binding proteins regulate RNA stability, splicing or mRNA export of immune-response transcripts. In particular, mutants defective in alternative splicing of resistance genes exhibit compromised disease resistance. Furthermore, detection of bacterial pathogens induces the differential expression of small non-coding RNAs including microRNAs that impact the host defense transcriptome. Phytopathogenic bacteria in turn have evolved effector proteins to inhibit biogenesis and/or activity of cellular microRNAs. Whereas RNA silencing has long been known as an antiviral defense response, recent findings also reveal a major role of this process in antibacterial defense. Here we review the function of RNA-binding proteins and small RNA-directed post-transcriptional regulation in antibacterial defense. We mainly focus on studies that used the model system Arabidopsis thaliana and also discuss selected examples from other plants.