A cucumber protein, Phloem Phosphate Stress-Repressed 1, rapidly degrades in response to a phosphate stress condition.

Under depleted external phosphate (Pi), many plant species adapt to this stress by initiating downstream signaling cascades. In plants, the vascular system delivers nutrients and signaling agents to control physiological and developmental processes. Currently, limited information is available regarding the direct role of phloem-borne long-distance signals in plant growth and development under Pi stress conditions. Here, we report on the identification and characterization of a cucumber protein, Cucumis sativus Phloem Phosphate Stress-Repressed 1 (CsPPSR1), whose level in the phloem translocation stream rapidly responds to imposed Pi-limiting conditions. CsPPSR1 degradation is mediated by the 26S proteasome; under Pi-sufficient conditions, CsPPSR1 is stabilized by its phosphorylation within the sieve tube system through the action of CsPPSR1 kinase. Further, we discovered that CsPPSR1 kinase was susceptible to Pi starvation-induced degradation in the sieve tube system. Our findings offer insight into a molecular mechanism underlying the response of phloem-borne proteins to Pi-limited stress conditions.

Plasmodesmal endoplasmic reticulum proteins regulate intercellular trafficking of cucumber mosaic virus in Arabidopsis.

Plasmodesmata (PD) are plasma membrane-lined cytoplasmic nanochannels that mediate cell-to-cell communication across the cell wall. A range of proteins are embedded in the PD plasma membrane and endoplasmic reticulum (ER), and function in regulating PD-mediated symplasmic trafficking. However, knowledge of the nature and function of the ER-embedded proteins in the intercellular movement of non-cell-autonomous proteins is limited. Here, we report the functional characterization of two ER luminal proteins, AtBiP1/2, and two ER integral membrane proteins, AtERdj2A/B, which are located within the PD. These PD proteins were identified as interacting proteins with cucumber mosaic virus (CMV) movement protein (MP) in co-immunoprecipitation studies using an Arabidopsis-derived plasmodesmal-enriched cell wall protein preparation (PECP). The AtBiP1/2 PD location was confirmed by TEM-based immunolocalization, and their AtBiP1/2 signal peptides (SPs) function in PD targeting. In vitro/in vivo pull-down assays revealed the association between AtBiP1/2 and CMV MP, mediated by AtERdj2A, through the formation of an AtBiP1/2-AtERdj2-CMV MP complex within PD. The role of this complex in CMV infection was established, as systemic infection was retarded in bip1/bip2w and erdj2b mutants. Our findings provide a model for a mechanism by which the CMV MP mediates cell-to-cell trafficking of its viral ribonucleoprotein complex.

Getting to the roots of N, P, and K uptake.

The soil contributes to the main pool of essential mineral nutrients for plants. These mineral nutrients are critical elements for the building blocks of plant biomolecules, play fundamental roles in cell processes, and act in various enzymatic reactions. The roots are the main entry point for mineral nutrients used within the plant to grow, develop, and produce seeds. In this regard, a suite of plant nutrient transport systems, sensors, and signaling proteins function in acquiring mineral nutrients through the roots. Mineral nutrients from chemical fertilizers, composed mainly of nitrogen, phosphorus, and potassium (NPK), are added to agricultural land to maximize crop yields, worldwide. However, improving nutrient uptake and use within crops is critical for economically and environmentally sustainable agriculture. Therefore, we review the molecular basis for N, P, and K nutrient uptake into the roots. Remarkably, plants are responsive to heterogeneous nutrient distribution and align root growth and nutrient uptake with nutrient-rich patches. We highlight the relationship between nutrient distribution in the growth environment and root system architecture. We discuss the exchange of information between the root and shoot systems through the xylem and phloem, which coordinates nutrient uptake with photosynthesis. The size and structure of the root system, along with the abundance and activity of nutrient transporters, largely determine the nutrient acquisition rate. Lastly, we discuss connections between N, P, and K uptake and signaling.

Plant lncRNAs are enriched in and move systemically through the phloem in response to phosphate deficiency.

In response to phosphate (Pi) deficiency, it has been shown that micro-RNAs (miRNAs) and mRNAs are transported through the phloem for delivery to sink tissues. Growing evidence also indicates that long non-coding RNAs (lncRNAs) are critical regulators of Pi homeostasis in plants. However, whether lncRNAs are present in and move through the phloem, in response to Pi deficiency, remains to be established. Here, using cucumber as a model plant, we show that lncRNAs are enriched in the phloem translocation stream and respond, systemically, to an imposed Pi-stress. A well-known lncRNA, IPS1, the target mimic (TM) of miRNA399, accumulates to a high level in the phloem, but is not responsive to early Pi deficiency. An additional 24 miRNA TMs were also detected in the phloem translocation stream; among them miRNA171 TMs and miR166 TMs were induced in response to an imposed Pi stress. Grafting studies identified 22 lncRNAs which move systemically into developing leaves and root tips. A CU-rich PTB motif was further identified in these mobile lncRNAs. Our findings revealed that lncRNAs respond to Pi deficiency, non-cell-autonomously, and may act as systemic signaling agents to coordinate early Pi deficiency signaling, at the whole-plant level.

Proteomics and metabolomics analyses reveal the cucurbit sieve tube system as a complex metabolic space.

The plant vascular system, and specifically the phloem, plays a pivotal role in allocation of fixed carbon to developing sink organs. Although the processes involved in loading and unloading of sugars and amino acids are well characterized, little information is available regarding the nature of other metabolites in the sieve tube system (STS) at specific sites along the pathway. Here, we elucidate spatial features of metabolite composition mapped with phloem enzymes along the cucurbit STS. Phloem sap (PS) was collected from the loading (source), unloading (apical sink region) and shoot-root junction regions of cucumber, watermelon and pumpkin. Our PS analyses revealed significant differences in the metabolic and proteomic profiles both along the source-sink pathway and between the STSs of these three cucurbits. In addition, metabolite profiles established for PS and vascular tissue indicated the presence of distinct compositions, consistent with the operation of the STS as a unique symplasmic domain. In this regard, at various locations along the STS we could map metabolites and their related enzymes to specific metabolic pathways. These findings are discussed with regard to the function of the STS as a unique and highly complex metabolic space within the plant vascular system.

A tomato phloem-mobile protein regulates the shoot-to-root ratio by mediating the auxin response in distant organs.

The plant vascular system serves as a conduit for delivery of both nutrients and signaling molecules to various distantly located organs. The anucleate sieve tube system of the angiosperm phloem delivers sugars and amino acids to developing organs, and has recently been shown to contain a unique population of RNA and proteins. Grafting studies have established that a number of these macromolecules are capable of moving long distances between tissues, thus providing support for operation of a phloem-mediated inter-organ communication network. Currently, our knowledge of the roles played by such phloem-borne macromolecules is in its infancy. Here, we show that, in tomato, translocation of a phloem-mobile cyclophilin, SlCyp1, from a wild-type scion into a mutant rootstock results in restoration of vascular development and lateral root initiation. This process occurs through reactivation of auxin response pathways and reprogramming of the root transcriptome. Moreover, we show that long-distance trafficking of SlCyp1 is associated with regulation of the shoot-to-root ratio in response to changing light intensities, by modulating root growth. We conclude that long-distance trafficking of SlCyp1 acts as a rheostat to control the shoot-to-root ratio, by mediating root development to integrate photosynthesis and light intensity with requirements for access to water and mineral nutrients.

Systemic delivery of siRNA in pumpkin by a plant PHLOEM SMALL RNA-BINDING PROTEIN 1-ribonucleoprotein complex.

In plants, the vascular system, specifically the phloem, functions in delivery of small RNA (sRNA) to exert epigenetic control over developmental and defense-related processes. Although the importance of systemic sRNA delivery has been established, information is currently lacking concerning the nature of the protein machinery involved in this process. Here, we show that a PHLOEM SMALL-RNA BINDING PROTEIN 1 (PSRP1) serves as the basis for formation of an sRNA ribonucleoprotein complex (sRNPC) that delivers sRNA (primarily 24 nt) to sink organs. Assembly of this complex is facilitated through PSRP1 phosphorylation by a phloem-localized protein kinase, PSRPK1. During long-distance transport, PSRP1-sRNPC is stable against phloem phosphatase activity. Within target tissues, phosphatase activity results in disassembly of PSRP1-sRNPC, a process that is probably required for unloading cargo sRNA into surrounding cells. These findings provide an insight into the mechanism involved in delivery of sRNA associated with systemic gene silencing in plants.

Overexpression of Arabidopsis plasmodesmata germin-like proteins disrupts root growth and development.

In plants, a population of non-cell-autonomous proteins (NCAPs), including numerous transcription factors, move cell to cell through plasmodesmata (PD). In many cases, the intercellular trafficking of these NCAPs is regulated by their interaction with specific PD components. To gain further insight into the functions of this NCAP pathway, coimmunoprecipitation experiments were performed on a tobacco (Nicotiana tabacum) plasmodesmal-enriched cell wall protein preparation using as bait the NCAP, pumpkin (Cucurbita maxima) PHLOEM PROTEIN16 (Cm-PP16). A Cm-PP16 interaction partner, Nt-PLASMODESMAL GERMIN-LIKE PROTEIN1 (Nt-PDGLP1) was identified and shown to be a PD-located component. Arabidopsis thaliana putative orthologs, PDGLP1 and PDGLP2, were identified; expression studies indicated that, postgermination, these proteins were preferentially expressed in the root system. The PDGLP1 signal peptide was shown to function in localization to the PD by a novel mechanism involving the endoplasmic reticulum-Golgi secretory pathway. Overexpression of various tagged versions altered root meristem function, leading to reduced primary root but enhanced lateral root growth. This effect on root growth was corrected with an inability of these chimeric proteins to form stable PD-localized complexes. PDGLP1 and PDGLP2 appear to be involved in regulating primary root growth by controlling phloem-mediated allocation of resources between the primary and lateral root meristems.

Phloem long-distance delivery of FLOWERING LOCUS T (FT) to the apex.

Cucurbita moschata FLOWERING LOCUS T-LIKE 2 (hereafter FTL2) and Arabidopsis thaliana (Arabidopsis) FLOWERING LOCUS T (FT), components of the plant florigenic signaling system, move long-distance through the phloem from source leaves to the vegetative apex where they mediate floral induction. The mechanisms involved in long-distance trafficking of FT/FTL2 remain to be elucidated. In this study, we identified the critical motifs on both FT and FTL2 required for cell-to-cell trafficking through mutant analyses using a zucchini yellow mosaic virus expression vector. Western blot analysis, performed on phloem sap collected from just beneath the vegetative apex of C. moschata plants, established that all mutant proteins tested retained the ability to enter the phloem translocation stream. However, immunolocalization studies revealed that a number of these FTL2/FT mutants were defective in the post-phloem zone, suggesting that a regulation mechanism for FT trafficking exists in the post-phloem unloading step. The selective movements of FT/FTL2 were further observed by microinjection and trichome rescue studies, which revealed that FT/FTL2 has the ability to dilate plasmodesmata microchannels during the process of cell-to-cell trafficking, and various mutants were compromised in their capacity to traffic through plasmodesmata. Based on these findings, a model is presented to account for the mechanism by which FT/FTL2 enters the phloem translocation stream and subsequently exits the phloem and enters the apical tissue, where it initiates the vegetative to floral transition.

The angiosperm phloem sieve tube system: a role in mediating traits important to modern agriculture.

The plant vascular system serves a vital function by distributing water, nutrients and hormones essential for growth and development to the various organs of the plant. In this review, attention is focused on the role played by the phloem as the conduit for delivery of both photosynthate and information macromolecules, especially from the context of its mediation in traits that are important to modern agriculture. Resource allocation of sugars and amino acids, by the phloem, to specific sink tissues is of importance to crop yield and global food security. Current findings are discussed in the context of a hierarchical control network that operates to integrate resource allocation to competing sinks. The role of plasmodesmata that connect companion cells to neighbouring sieve elements and phloem parenchyma cells is evaluated in terms of their function as valves, connecting the sieve tube pressure manifold system to the various plant tissues. Recent studies have also revealed that plasmodesmata and the phloem sieve tube system function cooperatively to mediate the long-distance delivery of proteins and a diverse array of RNA species. Delivery of these information macromolecules is discussed in terms of their roles in control over the vegetative-to-floral transition, tuberization in potato, stress-related signalling involving miRNAs, and genetic reprogramming through the delivery of 24-nucleotide small RNAs that function in transcriptional gene silencing in recipient sink organs. Finally, we discuss important future research areas that could contribute to developing agricultural crops with engineered performance characteristics for enhance yield potential.

Creating saponin-free yellow pea seeds by CRISPR/Cas9-enabled mutagenesis on ß-amyrin synthase.

Dry pea (Pisum sativum) seeds are valuable sources of plant protein, dietary fiber, and starch, but their uses in food products are restricted to some extent due to several off-flavor compounds. Saponins are glycosylated triterpenoids and are a major source of bitter, astringent, and metallic off-flavors in pea products. ß-amyrin synthase (BAS) is the entry point enzyme for saponin biosynthesis in pea and therefore is an ideal target for knock-out using CRISPR/Cas9 genome editing to produce saponin deficient pea varieties. Here, in an elite yellow pea cultivar (CDC Inca), LC/MS analysis identified embryo tissue, not seed coat, as the main location of saponin storage in pea seeds. Differential expression analysis determined that PsBAS1 was preferentially expressed in embryo tissue relative to seed coat and was selected for CRISPR/Cas9 genome editing. The efficiency of CRISPR/Cas9 genome editing of PsBAS1 was systematically optimized in pea hairy roots. From these optimization procedures, the AtU6-26 promoter was found to be superior to the CaMV35S promoter for gRNA expression, and the use of 37°C was determined to increase the efficiency of CRISPR/Cas9 genome editing. These promoter and culture conditions were then applied to stable transformations. As a result, a bi-allelic mutation (deletion and inversion mutations) was generated in the PsBAS1 coding sequence in a T1 plant, and the segregated psbas1 plants from the T2 population showed a 99.8% reduction of saponins in their seeds. Interestingly, a small but statistically significant increase (~12%) in protein content with a slight decrease (~5%) in starch content was observed in the psbas1 mutants under phytotron growth conditions. This work demonstrated that flavor-improved traits can be readily introduced in any pea cultivar of interest using CRISPR/Cas9. Further field trials and sensory tests for improved flavor are necessary to assess the practical implications of the saponin-free pea seeds in food applications.

Phloem-Mobile MYB44 Negatively Regulates Expression of PHOSPHATE TRANSPORTER 1 in Arabidopsis Roots.

Phosphorus (P) is an essential plant macronutrient; however, its availability is often limited in soils. Plants have evolved complex mechanisms for efficient phosphate (Pi) absorption, which are responsive to changes in external and internal Pi concentration, and orchestrated through local and systemic responses. To explore these systemic Pi responses, here we identified AtMYB44 as a phloem-mobile mRNA, an Arabidopsis homolog of Cucumis sativus MYB44, that is responsive to the Pi-starvation stress. qRT-PCR assays revealed that AtMYB44 was up-regulated and expressed in both shoot and root in response to Pi-starvation stress. The atmyb44 mutant displayed higher shoot and root biomass compared to wild-type plants, under Pi-starvation conditions. Interestingly, the expression of PHOSPHATE TRANSPORTER1;2 (PHT1;2) and PHT1;4 was enhanced in atmyb44 in response to a Pi-starvation treatment. A split-root assay showed that AtMYB44 expression was systemically regulated under Pi-starvation conditions, and in atmyb44, systemic controls on PHT1;2 and PHT1;4 expression were moderately disrupted. Heterografting assays confirmed graft transmission of AtMYB44 transcripts, and PHT1;2 and PHT1;4 expression was decreased in heterografted atmyb44 rootstocks. Taken together, our findings support the hypothesis that mobile AtMYB44 mRNA serves as a long-distance Pi response signal, which negatively regulates Pi transport and utilization in Arabidopsis.

CmRBP50 protein phosphorylation is essential for assembly of a stable phloem-mobile high-affinity ribonucleoprotein complex.

RNA-binding proteins (RBPs) form ribonucleoprotein (RNP) complexes that play crucial roles in RNA processing for gene regulation. The angiosperm sieve tube system contains a unique population of transcripts, some of which function as long-distance signaling agents involved in regulating organ development. These phloem-mobile mRNAs are translocated as RNP complexes. One such complex is based on a phloem RBP named Cucurbita maxima RNA-binding protein 50 (CmRBP50), a member of the polypyrimidine track binding protein family. The core of this RNP complex contains six additional phloem proteins. Here, requirements for assembly of this CmRBP50 RNP complex are reported. Phosphorylation sites on CmRBP50 were mapped, and then coimmunoprecipitation and protein overlay studies established that the phosphoserine residues, located at the C terminus of CmRBP50, are critical for RNP complex assembly. In vitro pull-down experiments revealed that three phloem proteins, C. maxima phloem protein 16, C. maxima GTP-binding protein, and C. maxima phosphoinositide-specific phospholipase-like protein, bind directly with CmRBP50. This interaction required CmRBP50 phosphorylation. Gel mobility-shift assays demonstrated that assembly of the CmRBP50-based protein complex results in a system having enhanced binding affinity for phloem-mobile mRNAs carrying polypyrimidine track binding motifs. This property would be essential for effective long-distance translocation of bound mRNA to the target tissues.

Vascular expression in Arabidopsis is predicted by the frequency of CT/GA-rich repeats in gene promoters.

Phloem-transported signals play an important role in regulating plant development and in orchestrating responses to environmental stimuli. Among such signals, phloem-mobile RNAs have been shown to play an important role as long-distance signaling agents. At maturity, angiosperm sieve elements are enucleate, and thus transcripts in the phloem translocation stream probably originate from the nucleate companion cells. In the present study, a pumpkin (Cucurbita maxima) phloem transcriptome was used to test for the presence of common motifs within the promoters of this unique set of genes, which may function to coordinate expression in cells of the vascular system. A bioinformatics analysis of the upstream sequences from 150 Arabidopsis genes homologous to members of the pumpkin phloem transcriptome identified degenerate sequences containing CT/GA- and GT/CA-rich motifs that were common to many of these promoters. Parallel studies performed on genes shown previously to be expressed in phloem tissues identified similar motifs. An expanded analysis, based on homologs of the pumpkin phloem transcriptome from cucumber (Cucumis sativus), identified similar sets of common motifs within the promoters of these genes. Promoter analysis offered support for the hypothesis that these motifs regulate expression within the vascular system. Our findings are discussed in terms of a role for these motifs in coordinating gene expression within the companion cell/sieve element system. These motifs could provide a useful bioinformatics tool for genome-wide screens on plants for which phloem tissues cannot readily be obtained.

Systemic Signaling: A Role in Propelling Crop Yield.

Food security has become a topic of great concern in many countries. Global food security depends heavily on agriculture that has access to proper resources and best practices to generate higher crop yields. Crops, as with other plants, have a variety of strategies to adapt their growth to external environments and internal needs. In plants, the distal organs are interconnected through the vascular system and intricate hierarchical signaling networks, to communicate and enhance survival within fluctuating environments. Photosynthesis and carbon allocation are fundamental to crop production and agricultural outputs. Despite tremendous progress achieved by analyzing local responses to environmental cues, and bioengineering of critical enzymatic processes, little is known about the regulatory mechanisms underlying carbon assimilation, allocation, and utilization. This review provides insights into vascular-based systemic regulation of photosynthesis and resource allocation, thereby opening the way for the engineering of source and sink activities to optimize the yield performance of major crops.

The Mobile Small RNAs: Important Messengers for Long-Distance Communication in Plants.

Various species of small RNAs (sRNAs), notably microRNAs and small interfering RNAs (siRNAs), have been characterized as the major effectors of RNA interference in plants. Growing evidence supports a model in which sRNAs move, intercellularly, systemically, and between cross-species. These non-coding sRNAs can traffic cell-to-cell through plasmodesmata (PD), in a symplasmic manner, as well as from source to sink tissues, via the phloem, to trigger gene silencing in their target cells. Such mobile sRNAs function in non-cell-autonomous communication pathways, to regulate various biological processes, such as plant development, reproduction, and plant defense. In this review, we summarize recent progress supporting the roles of mobile sRNA in plants, and discuss mechanisms of sRNA transport, signal amplification, and the plant's response, in terms of RNAi activity, within the recipient tissues. We also discuss potential research directions and their likely impact on engineering of crops with traits for achieving food security.

Pumpkin eIF5A isoforms interact with components of the translational machinery in the cucurbit sieve tube system.

In yeast, eIF5A, in combination with eEF2, functions at the translation step, during the protein elongation cycle. This result is of significance with respect to functioning of the enucleate sieve tube system, as eIF5A was recently detected in Cucurbita maxima (pumpkin) phloem sap. In the present study, we further characterized four CmeIF5A isoforms, encoding three proteins, all of which were present in the phloem sap. Although hypusination of CmeIF5A was not necessary for entry into the sieve elements, this unique post-translational modification was necessary for RNA binding. The two enzymes required for hypusination were detected in pumpkin phloem sap, where presumably this modification takes place. A combination of gel-filtration chromatography and protein overlay assays demonstrated that, as in yeast, CmeIF5A interacts with phloem proteins, like eEF2, known to be involved in protein synthesis. These findings are discussed in terms of a potential role for eIF5A in regulating protein synthesis within the enucleate sieve tube system of the angiosperms.

A zinc finger protein Tsip1 controls Cucumber mosaic virus infection by interacting with the replication complex on vacuolar membranes of the tobacco plant.

• In Cucumber mosaic virus (CMV) RNA replication, replicase-associated protein CMV 1a and RNA-dependent RNA polymerase protein CMV 2a are essential for formation of an active virus replicase complex on vacuolar membranes. • To identify plant host factors involved in CMV replication, a yeast two-hybrid system was used with CMV 1a protein as bait. One of the candidate genes encoded Tsi1-interacting protein 1 (Tsip1), a zinc (Zn) finger protein. Tsip1 strongly interacted with CMV 2a protein, too. • Formation of a Tsip1 complex involving CMV 1a or CMV 2a was confirmed in vitro and in planta. When 35S::Tsip1 tobacco (Nicotiana tabacum) plants were inoculated with CMV-Kor, disease symptom development was delayed and the accumulation of CMV RNAs and coat protein was decreased in both the infected local leaves and the uninfected upper leaves, compared with the wild type, whereas Tsip1-RNAi plants showed modestly but consistently increased CMV susceptibility. In a CMV replication assay, CMV RNA concentrations were reduced in the 35S::Tsip1 transgenic protoplasts compared with wild-type (WT) protoplasts. • These results indicate that Tsip1 might directly control CMV multiplication in tobacco plants by formation of a complex with CMV 1a and CMV 2a.

A Plant SMALL RNA-BINDING PROTEIN 1 Family Mediates Cell-to-Cell Trafficking of RNAi Signals.

In plants, RNA interference (RNAi) plays a pivotal role in growth and development, and responses to environmental inputs, including pathogen attack. The intercellular and systemic trafficking of small interfering RNA (siRNA)/microRNA (miRNA) is a central component in this regulatory pathway. Currently, little is known with regards to the molecular agents involved in the movement of these si/miRNAs. To address this situation, we employed a biochemical approach to identify and characterize a conserved SMALL RNA-BINDING PROTEIN 1 (SRBP1) family that mediates non-cell-autonomous small RNA (sRNA) trafficking. In Arabidopsis, AtSRBP1 is a glycine-rich (GR) RNA-binding protein, also known as AtGRP7, which we show binds single-stranded siRNA. A viral vector, Zucchini yellow mosaic virus (ZYMV), was employed to functionally characterized the AtSRBP1-4 (AtGRP7/2/4/8) RNA recognition motif and GR domains. Cellular-based studies revealed the GR domain as being necessary and sufficient for SRBP1 cell-to-cell movement. Taken together, our findings provide a foundation for future research into the mechanism and function of mobile sRNA signaling agents in plants.

A novel methyltransferase methylates Cucumber mosaic virus 1a protein and promotes systemic spread.

In mammalian and yeast systems, methyltransferases have been implicated in the regulation of diverse processes, such as protein-protein interactions, protein localization, signal transduction, RNA processing, and transcription. The Cucumber mosaic virus (CMV) 1a protein is essential not only for virus replication but also for movement. Using a yeast two-hybrid system with tobacco plants, we have identified a novel gene encoding a methyltransferase that interacts with the CMV 1a protein and have designated this gene Tcoi1 (tobacco CMV 1a-interacting protein 1). Tcoi1 specifically interacted with the methyltransferase domain of CMV 1a, and the expression of Tcoi1 was increased by CMV inoculation. Biochemical studies revealed that the interaction of Tcoi1 with CMV 1a protein was direct and that Tcoi1 methylated CMV 1a protein both in vitro and in vivo. The CMV 1a binding activity of Tcoi1 is in the C-terminal domain, which shows the methyltransferase activity. The overexpression of Tcoi1 enhanced the CMV infection, while the reduced expression of Tcoi1 decreased virus infectivity. These results suggest that Tcoi1 controls the propagation of CMV through an interaction with the CMV 1a protein.