Virusoids: Viruses’ Very Own Parasites

(Posted on Friday, April 7, 2023)

This article is part of a series on subviral agents. The first three installments —which can be read here, here, and here, respectively— discussed viroids, the smallest known pathogens. Now, we turn to virusoids, which straddle the line between viroids and viruses. 

Great fleas have little fleas upon their backs to bite ’em,

And little fleas have lesser fleas, and so ad infinitum.

And the great fleas themselves, in turn, have greater fleas to go on;

While these again have greater still, and greater still, and so on.

             – Augustus De Morgan,  A Budget of Paradoxes

Viroids are tiny strands of circular, infectious RNA. They differ from viruses in that they do not have a protective protein coat and that they do not encode any proteins of their own. They are also much, much smaller than the average virus. Think of viroids as extreme minimalists, traveling as lightweight as possible. Viruses are generally more complex, with larger molecular “toolboxes” — all viruses produce at least one protein, but most produce a whole array of them, used for replication, and in some cases, suppression and evasion of the host immune system. 

Virusoids, in turn, straddle the line between viroids and viruses: like viroids, they are small circular RNAs that, for the most part, do not encode any proteins; like viruses, they are encapsidated in a protein coat, albeit not of their own making. The hallmark difference between viroids and virusoids is that the latter cannot replicate without assistance from a helper virus, which provides it with its protein coat and the necessary “machinery” for replication. So in order for a virusoid to infect a host, it needs to be carried in “upon the back of” a virus. If there is no helper virus to hitch a ride on, the virusoid is stuck outside, with no way into the host and no way to replicate. Curiously, virusoids share little to none of their genetic material with their helper viruses, which means they evolved independently of them and are not simply maladapted offshoots. This clearly differentiates them from another type of subviral agent, defective interfering particles (DIPs), which are small mutants of a parent virus generated in moments of defective replication.  

To summarize, virusoids (also called “viroid-like satellite RNAs”) are circular, single-stranded RNA molecules that exhibit the following traits: “(1) satRNA parasitizes on helper virus (HV) replication machinery (2) satRNA is encapsidated by the HV capsid protein to promote transmission and (3) satRNA and the HV genomic RNAs have no nucleotide sequence similarity.” 

Host Range and Classification

Virusoids can be separated into groups according to the genus of their helper virus (Table 1). In plant virusoids, five are linked to the sobemovirus genus, three are linked to the nepovirus genus, and one is linked to the polerovirus genus. In animals, we only know of one viroid-like satellite RNA to date: hepatitis delta virus (HDV), which depends on hepatitis B virus for transmission and replication.

TABLE 1. Overview of known virusoids (“viroid-like satellite RNAs”); sc-satRNA = small circular satellite RNA; nt = nucleotides. SOURCE: Adapted from Navarro et al. (2017), “Chapter 61 – Small Circular Satellite RNAs”

Plant Virusoid Replication

Like viroids, virusoid replication is a three-step affair, making use of the rolling-circle mechanism. First, the circular RNA is replicated into a long chain of RNAs containing multiple copies of the original genome; helper virus polymerases are likely hijacked to this end. Second, the long chain of “oligomeric” RNAs —to use technical speak— is then cut up into its individual units by host enzymes or, occasionally, by ribozymes encoded in the virusoid genome. Ribozymes are RNA sequences that, despite not encoding a protein, are “active” — they can themselves perform and catalyze tasks, such as cleavage, usually expected of protein enzymes. Finally, the individual strands of RNA have to be circularized again to fully form the virusoid particles. Known as ligation, this takes place either through the appropriation of host enzymes, or in rare instances, through self-ligation catalyzed by the ribozyme. 

Replication can take place in one of two ways: symmetric or asymmetric (Figure 1). The circular genome that viroids and virusoids start out with is positive-sense (+). During rolling-circle replication, however, the RNA is copied into a negative (-) strand — the mirror image of the original. In order to get the negative-strand RNA back to its original sense, it has to be copied a second time, flipping the mirror image back again. How the negative strand is copied back to the positive sense is what differentiates symmetric from asymmetric replication. In symmetric replication, this happens via a second instance of rolling-circle replication, aided by ribozyme cleavage. In the case of asymmetric replication, the negative-sense RNA gets copied back to its positive polarity through the help of host enzymes, which also serve to cleave the RNA into its unit-length strands. 

FIGURE 1. “Mechanism proposed for replication of viroids, viroid-like satellite rNAs and HDv. The asymmetric pathway with one rolling-circle (lower) is used by viroids of the family Pospiviroidae and by some viroid-like satellite rNAs, while the symmetric pathway with two rolling-circles (upper) is used by the other viroid-like satellite rNAs, viroids of the family Avsunviroidae and HDv. Solid and open lines refer to plus (+) and minus (-) polarities, respectively and processing sites are marked by arrowheads. Cleavage is alternatively mediated by ribozymes (rZ) or host enzymes (He), generating linear monomeric rNAs with characteristic termini that are subsequently ligated by host enzymes or autocatalytically.” SOURCE: FLORES ET AL. 2011, https://doi.org/10.4161/rna.8.2.14238