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In addition to well-known roles in RNA metabolism, the nucleolus and Cajal bodies (CBs), both located within the nucleus, are involved in plant responses to biotic and abiotic stress. Previously we showed that plants in which expression of the CB protein coilin is downregulated are more susceptible to certain viruses including tobacco rattle virus (TRV), suggesting a role of coilin in antiviral defence. Experiments with coilin-deficient plants and the deletion mutant of the TRV 16K protein showed that both 16K and coilin are required for restriction of systemic TRV infection. The potential mechanisms of coilin-mediated antiviral defence were elucidated via experiments involving co-immunoprecipitation, use of NahG transgenic plants deficient in salicylic acid (SA) accumulation, measurement of endogenous SA concentrations and assessment of SA-responsive gene expression. Here we show that TRV 16K interacts with and relocalizes coilin to the nucleolus. In wild-type plants these events are accompanied by activation of SA-responsive gene expression and restriction of TRV systemic infection. By contrast, viral systemic spread was enhanced in NahG plants, implicating SA in these processes. Our findings suggest that coilin is involved in plant defence, responding to TRV infection by recognition of the TRV-encoded 16K protein and activating SA-dependent defence pathways., (© 2019 The Authors. New Phytologist © 2019 New Phytologist Trust.)

Human B-cell receptor-associated protein BAP31 (HsBAP31) is the endoplasmic reticulum-resident protein involved in protein sorting and transport as well as pro-apoptotic signaling. Plant orthologs of HsBAP31 termed 'plant BAP-like proteins' (PBL proteins) have thus far remained unstudied. Recently, the PBL protein from Nicotiana tabacum (NtPBL) was identified as an interactor of Nt-4/1, a plant protein known to interact with plant virus movement proteins and affect the long-distance transport of potato spindle tuber viroid (PSTVd) via the phloem. Here, we have compared the sequences of PBL proteins and studied the biochemical properties of NtPBL. Analysis of a number of fully sequenced plant genomes revealed that PBL-encoding genes represent a small multigene family with up to six members per genome. Two conserved motifs were identified in the C-terminal region of PBL proteins. The NtPBL C-terminal hydrophilic region (NtPBL-C) was expressed in bacterial cells, purified, and used for analysis of its RNA binding properties in vitro. In gel shift experiments, NtPBL-C was found to bind several tested RNAs, showing the most efficient binding to microRNA precursors (pre-miRNA) and less efficient interaction with PSTVd. Mutational analysis suggested that NtPBL-C has a composite RNA-binding site, with two conserved lysine residues in the most C-terminal protein region being involved in binding of pre-miRNA but not PSTVd RNA. Virus-mediated transient expression of NtPBL-C in plants resulted in stunting and leaf malformation, developmental abnormalities similar to those described previously for blockage of miRNA biogenesis/function. We hypothesize that the NtPBL protein represents a previously undiscovered component of the miRNA pathway., (Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

Cajal bodies (CBs) are distinct nuclear bodies physically and functionally associated with the nucleolus. In addition to their traditional function in coordinating maturation of certain nuclear RNAs, CBs participate in cell cycle regulation, development, and regulation of stress responses. A key "signature" component of CBs is coilin, the scaffolding protein essential for CB formation and function. Using an RNA silencing (loss-of-function) approach, we describe here new phenomena whereby coilin also affects, directly or indirectly, a variety of interactions between host plants and viruses that have RNA or DNA genomes. Moreover, the effects of coilin on these interactions are manifested differently: coilin contributes to plant defense against tobacco rattle virus (tobravirus), tomato black ring virus (nepovirus), barley stripe mosaic virus (hordeivirus), and tomato golden mosaic virus (begomovirus). In contrast, with potato virus Y (potyvirus) and turnip vein clearing virus (tobamovirus), coilin serves to increase virus pathogenicity. These findings show that interactions with coilin (or CBs) may involve diverse mechanisms with different viruses and that these mechanisms act at different phases of virus infection. Thus, coilin (CBs) has novel, unexpected natural functions that may be recruited or subverted by plant viruses for their own needs or, in contrast, are involved in plant defense mechanisms that suppress host susceptibility to the viruses.

The Arabidopsis thaliana 4/1 (At-4/1) protein has a highly α-helical structure with potential to interact both with itself and other protein ligands, including the movement proteins of some plant viruses; the Nicotiana tabacum ortholog (Nt-4/1) has similar structure. Here we describe localization of GUS expression in transgenic N. tabacum seedlings under control of the Nt-4/1 promoter, which indicates that transcription is associated with the veins at certain developmental stages, and especially in the hypocotyl. Viroid accumulation and movement was altered in plants in which 4/1 expression was reduced by virus-induced gene silencing. These localization studies support a role of 4/1 in signaling in the vasculature,including mobility of pathogen-related and cellular RNAs.

Arabidopsis thaliana At-4/1 is the protein of unknown function capable of polar localization in plant cells and intercellular trafficking. In this work, we cloned cDNAs and chromosomal genes of At-4/1 orthologues from several Nicotiana species. Similarly to the 4/1 genes of A. thaliana and Oryza sativa, Nicotiana 4/1 genes have eight exons and seven introns but are considerably longer due to their larger introns. The allotetraploid genome of Nicotiana tabacum, which is known to consist of the 'S genome' originated from Nicotiana sylvestris and the 'T genome' derived from Nicotiana tomentosiformis, encodes two 4/1 genes. The T genome-encoded 4/1 gene, but not that of the S genome, contains a SINE-like transposable element in its intron 2. The 4/1 genes of Nicotiana hesperis and Nicotiana benthamiana lack such an element in the intron 2, but possess a related SINE-like sequence in their intron 4. Collectively, the sequence analysis data provide an insight into the organization of 4/1 genes in flowering plants and the patterns of evolution in the genus Nicotiana. The Nicotiana 4/1 proteins and those of other flowering plants show a significant level of sequence similarity. Computer-assisted analysis was further used to compare their predicted secondary structures. Several algorithms confidently predicted the presence of several coiled-coil domains occupying similar positions in different 4/1 proteins. Analysis of circular dichroism spectra carried out for bacterially expressed N. tabacum 4/1 protein (Nt-4/1) and its N- and C-terminally truncated mutants confirmed that the secondary structure of Nt-4/1 is generally alpha-helical. The C-terminal region of Nt-4/1 was found to undergo a partial proteolysis in Escherichia coli cells. Differential scanning calorimetry of Nt-4/1 protein and its mutants revealed three calorimetric domains most probably corresponding to the N-terminal, central, and C-terminal structural domains of the protein., (Copyright © 2011 Elsevier Masson SAS. All rights reserved.)