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Many viral suppressors (VSRs) counteract antiviral RNA silencing, a central component of the plant's immune response by sequestration of virus-derived antiviral small interfering RNAs (siRNAs). Here, we addressed how VSRs affect the activities of cellular microRNAs (miRNAs) during a viral infection by characterizing the interactions of two unrelated VSRs, the Tombusvirus p19 and the Cucumovirus 2b, with miRNA 162 (miR162), miR168, and miR403. These miRNAs regulate the expression of the important silencing factors Dicer-like protein 1 (DCL1) and Argonaute proteins 1 and 2 (AGO1 and AGO2), respectively. Interestingly, while the two VSRs showed similar binding profiles, the miRNAs were bound with significantly different affinities, for example, with the affinity of miR162 greatly exceeding that of miR168. In vitro silencing experiments revealed that p19 and 2b affect miRNA-mediated silencing of the DCL1, AGO1, and AGO2 mRNAs in strict accordance with the VSR's miRNA-binding profiles. In Tombusvirus-infected plants, the miRNA-binding behavior of p19 closely corresponded to that in vitro Most importantly, in contrast to controls with a ?p19 virus, infections with wild-type (wt) virus led to changes of the levels of the miRNA-targeted mRNAs, and these changes correlated with the miRNA-binding preferences of p19. This was observed exclusively in the early stage of infection when viral genomes are proposed to be susceptible to silencing and viral siRNA (vsiRNA) concentrations are low. Accordingly, our study suggests that differential binding of miRNAs by VSRs is a widespread viral mechanism to coordinately modulate cellular gene expression and the antiviral immune response during infection initiation.IMPORTANCE Plant viruses manipulate their hosts in various ways. Viral suppressor proteins (VSRs) interfere with the plant's immune response by sequestering small, antivirally acting vsiRNAs, which are processed from viral RNAs during the plant's RNA-silencing response. Here, we examined the effects of VSRs on cellular microRNAs (miRNAs), which show a high degree of similarity with vsiRNAs. Binding experiments with two unrelated VSRs and three important regulatory miRNAs revealed that the proteins exhibit similar miRNA-binding profiles but bind different miRNAs at considerably different affinities. Most interestingly, experiments in plants showed that in the early infection phase, the Tombusvirus VSR p19 modulates the activity of these miRNAs on their target mRNAs very differently and that this differential regulation strictly correlates with the binding affinities of p19 for the respective miRNAs. Our data suggest that VSRs may specifically control plant gene expression and the early immune response by differential sequestration of miRNAs.

The female sex in honeybees (Apis spp.) comprises a reproductive queen and a sterile worker caste. Nurse bees feed all larvae progressively with a caste-specific food jelly until the prepupal stage. Only those larvae that are exclusively fed a large amount of royal jelly (RJ) develop into queens [1]. RJ is a composite secretion of two specialized head glands: the mandibular glands, which produce mainly fatty acids [2], and the hypopharyngeal glands, which contribute proteins, primarily belonging to the major royal jelly protein (MRJP) family [3]. Past research on RJ has focused on its nutritional function and overlooked its central role with regard to the orientation of the larva in the royal brood cell. Whereas workers are reared in the regular horizontal cells of the comb, the queen cells are specifically built outside of the normal comb area to accommodate for the larger queen [4, 5]. These cells hang freely along the bottom of the comb and are vertically oriented, opening downward [6]. Queen larvae are attached by their RJ diet to the cell ceiling. Thus, the physical properties of RJ are central to successful retention of larvae in the cell. Here, we show that the main protein of RJ (MRJP1) polymerizes in complex with another protein, apisimin, into long fibrous structures that build the basis for the high viscosity of RJ to hold queen larvae on the RJ surface.

Phage shock protein A (PspA) belongs to the highy conserved PspA/IM30 family and is a key component of the stress inducible Psp system in Escherichia coli. One of its central roles is the regulatory interaction with the transcriptional activator of this system, the σ54 enhancer binding protein PspF, a member of the AAA+ protein family. The PspA/F regulatory system has been intensively studied and serves as a paradigm for AAA+ enzyme regulation by trans-acting factors. However, the molecular mechanism of how exactly PspA controls the activity of PspF and hence σ54-dependent expression of the psp genes is still unclear. To approach this question, we identified the minimal PspF-interacting domain of PspA, solved its structure, determined its affinity to PspF and the dissociation kinetics, identified residues that are potentially important for PspF regulation and analyzed effects of their mutation on PspF in vivo and in vitro. Our data indicate that several characteristics of AAA+ regulation in the PspA·F complex resemble those of the AAA+ unfoldase ClpB, with both proteins being regulated by a structurally highly conserved coiled-coil domain. The convergent evolution of both regulatory domains points to a general mechanism to control AAA+ activity for divergent physiological tasks via coiled-coil domains.