Pokeweed Antiviral Protein, a Ribosome Inactivating Protein: Activity, Inhibition and Prospects

Abstract

Viruses employ an array of elaborate strategies to overcome plant defense mechanisms and must adapt to the requirements of the host translational systems. Pokeweed antiviral protein (PAP) from Phytolacca americana is a ribosome inactivating protein (RIP) and is an RNA N-glycosidase that removes specific purine residues from the sarcin/ricin (S/R) loop of large rRNA, arresting protein synthesis at the translocation step. PAP is thought to play an important role in the plant’s defense mechanism against foreign pathogens. This review focuses on the structure, function, and the relationship of PAP to other RIPs, discusses molecular aspects of PAP antiviral activity, the novel inhibition of this plant toxin by a virus counteraction—a peptide linked to the viral genome (VPg), and possible applications of RIP-conjugated immunotoxins in cancer therapeutics.

Insights into the Molecular Antiviral Mechanism of Pokeweed Protein from Phytolacca americana

“…In the instances where the 5ʹ-cap is absent, PAP directly binds to either the 5ʹ-or 3ʹ-UTRs (untranslated regions), containing either translational enhancer sequences or an internal ribosome entry site (K d for PAPuncapped full length TEV 5ʹ-leader RNA is 28.5 ± 3.7 nM; K d for PAP-m 7 GpppG-capped full length TEV 5ʹ-leader RNA is 87.5 ± 4.8 nM) [62]; binding of the eIFs increases PAP-RNA affinity, promoting depurination of RNA (presence of eIFiso4G increases 2.4-fold PAP-cap interactions) [53]. Additionally, PAP isoforms selectivity for different ribosomes and RNAs varies (e.g., PAP-I, found in spring leaves of the pokeweed plant, exhibits RC 50 of 1.5 nM towards rat liver ribosomes and 4.7 nM towards E. coli ribosomes [38]; PAP-S1, an isoform found in seeds of the plant, exhibits IC 50 of 3.2 nM towards rat liver ribosomes and 280 nM towards E. coli ribosomes [41,42]; whereas α-PAP, expressed in all organs of the plant, exhibits IC 50 of 1.3 nM towards rat liver ribosomes and 25 nM towards E. coli ribosomes [13,42]. In recent years, a viral protein (VPg), linked to genome of turnip mosaic virus (TuMV) was shown to inhibit PAP activity in vitro [13,57].…”

“…Additionally, PAP isoforms selectivity for different ribosomes and RNAs varies (e.g., PAP-I, found in spring leaves of the pokeweed plant, exhibits RC 50 of 1.5 nM towards rat liver ribosomes and 4.7 nM towards E. coli ribosomes [38]; PAP-S1, an isoform found in seeds of the plant, exhibits IC 50 of 3.2 nM towards rat liver ribosomes and 280 nM towards E. coli ribosomes [41,42]; whereas α-PAP, expressed in all organs of the plant, exhibits IC 50 of 1.3 nM towards rat liver ribosomes and 25 nM towards E. coli ribosomes [13,42]. In recent years, a viral protein (VPg), linked to genome of turnip mosaic virus (TuMV) was shown to inhibit PAP activity in vitro [13,57]. This viral peptide serves as an analog of the 5ʹ-m 7 G cap of viral RNA, and has been shown to play an important role in mRNA translation since it interacts with the cap-binding proteins (e.g., eIF4E, eIFiso4E, eIF4F, eIFiso4F) [64,65].…”

“…The biological effects credited to these protein toxins go back to early times, owing to the high toxicities of the castor bean and jequirity bean [10]. Yet other plants, such as American pokeweed (P. americana) and common soapwort, synthesize pokeweed antiviral protein (PAP) and saporin, that impose lower toxicity on intact cells while inhibiting protein production in cell-free lysates [11][12][13]. RIPs exert their cytotoxicity by impairing ribosomes [14]; specifically they interfere with the advanced stages of translation [15][16][17][18][19].…”

“…It has been postulated that a direct interaction of PAP with viral RNA (or DNA) is an alternative antiviral mechanism in play. The pokeweed plant produces several isozymes of PAP, all exerting potent antiviral properties [11,13,[36][37][38][39][40][41][42]. PAP isozymes evoke depurination of genomic HIV-1 RNA [43][44][45], TMV RNA [46], poliovirus [47], herpes simplex virus (HSV) [48], influenza virus [49], and brome mosaic virus (BMV) [50], among many others, showing a broad spectrum of antiviral activity [13].…”

Pokeweed (Phytolacca americana L.) antiviral protein inhibits Zucchini yellow mosaic virus infection in a dose-dependent manner in squash plants

“…Many plants contain ribosome-inactivating proteins (RIPs) catalytically inactivating prokaryotic and eukaryotic ribosomes (Domashevskiy and Goss, 2015). These enzymes remove a single adenine residue from the large rRNA through RNA N-glycosidase activity.…”