Small­est Things Con­sid­ered

by Merry

What do stunted co­conut palms, mis­shapen potato tu­bers, and peach trees with necrotic branches have in com­mon? They are three of the nu­mer­ous crops stricken with dis­eases caused by vi­roids, an as­ton­ish­ing group of min­i­mal­ist plant pathogens. There isn't much to a vi­roid, just one sin­gle-stranded, cir­cu­lar RNA mol­e­cule. The lar­gest vi­roid genome so far is 399 nu­cleotides, the small­est a mere 246 – about one tenth the size of the small­est vi­ruses (he­pad­navirus) and one hun­dredth the size of more typ­i­cal viruses. Be­ing la­beled as "sub­vi­ral," they are even less likely than the viruses to be granted a place on the tree of life. They get by with­out cap­sid or mem­brane shell. They en­code no pro­teins. They don't re­verse-tran­scribe in­to DNA when they repli­cate. They never in­sert into the host genome. Some of them cause dis­ease symp­toms, some don't. They sim­ply repli­cate in­side plant cells and then their prog­eny move on to the next lo­ca­tion to re­peat the process. Their very ex­is­tence raises quest­ions, many with­out an­swers.

Leaflets of oil palm in­fected with cadang-ca­­dang-like vi­roid and show­ing symp­toms of or­ange spot­ting. Source

What does a vi­roid look like? Well, that de­pends on the vi­roid. The thirty-plus species known so far fall into two groups. Most be­long to the Pospivi­roidae (PSTVd), named af­ter the Potato Spin­dle Tuber ViroiD. Four species, in­clud­ing the Avo­cado SunBlotch ViroiD, make up their own group, the Avsunvi­roi­dae (AS­BVd). Al­though all of them lack pro­tec­tive cap­sids, they are nev­er­the­less highly struc­tured.

Most of their RNA genome base-pairs with com­ple­men­tary se­quences in other parts of the same RNA mol­e­cule to form sec­tions of dou­ble-he­lix. The PSTVds form rod-shaped struc­tures, the AS­B­Vds have more elab­o­rate forms. (See fig­ure.) For both groups, the bulges and loops are im­portant for sur­vival, repli­ca­tion, and traf­fick­ing. (For a re­cent re­view, click here.)

Pre­dicted low­est free en­ergy sec­ondary struc­tures of (a) the po­tato spin­dle tu­ber vi­roid, a Pospi­vi­roidae, and (b) peach la­tent mo­saic vi­roid, an Av­sun­vi­roidae. Source

How do they repli­cate when they en­code no pro­teins? Vi­roids rely on host cell fac­tors to help with just about every­thing. Once the vi­roid RNA en­ters a cell, it re­cruits fac­tors for "RNA traf­fick­ing" to the com­part­ment where it repli­cates; that's the nu­cleus for the PSTVds, the chloro­plast for the AS­B­Vds. Then, for copy­ing their RNA, both groups co-opt host DNA-de­pen­dent RNA poly­merases, redi­rect­ing them to use the vi­roid RNA as tem­plate. This works, but is er­ror-prone, giv­ing one of the vi­roids the fastest known rate of mu­ta­tion.

Both types repli­cate us­ing a rolling cir­cle mech­a­nism. Al­though the de­tails dif­fer, they both pro­duce long, mul­ti­meric strands of RNA that must be cut into monomers, each monomer then li­gated to form an in­di­vid­ual cir­cu­lar vi­roid. For the cut­ting AS­B­Vds em­ploy a do-it-your­self strat­egy. They ded­i­cate 40 nu­cleotides of their se­quence to form the small­est known nat­u­rally-oc­cur­ring ri­bozyme. This ham­mer­head ri­bozyme cat­alyzes the self-cleav­age needed to trans­form the long mul­ti­meric strands into in­di­vid­ual genomes. Af­ter repli­ca­tion, the prog­eny RNAs use host fac­tors to help them move to neigh­bor­ing cells through the con­nect­ing plas­mod­es­mata or to dis­tant parts of the plant via the phloem.

Vi­roids un­der the elec­tron mi­cro­scope. Source

Next ques­tion. How do they cause dis­ease if they don't en­code any pro­teins? Di­rectly, by RNA in­ter­fer­ence. RNAi is a mech­a­nism that de­grades dou­ble-stranded RNAs (dsRNA). In plants, this de­fends against RNA viruses by at­tack­ing their dou­ble-stranded repli­ca­tion intermedi­ates. Not only does this strat­egy not work against vi­roids, but it ac­tu­ally causes the symp­toms as­so­ci­ated with vi­roid in­fec­tion. How does this hap­pen? Short lengths of RNA clipped from the vi­roid genome can base pair with com­ple­men­tary host mRNA to form dsR­NAs that are then de­graded. De­stroy­ing the mR­NAs ef­fec­tively turns off the cor­re­spond­ing genes in the host. You might won­der why host-de­rived RNAi doesn't de­grade the dou­ble-stranded vi­roids. This is likely due to the viroid's com­plex sec­ondary struc­ture and to the fact that all of their dou­ble-stranded re­gions are too short to ac­ti­vate RNAi.

Where did vi­roids come from? Vi­roids show no re­la­tion­ship to viruses, but there is some ev­i­dence – all seem­ingly a bit of a stretch – sug­gest­ing evo­lu­tion­ary links to trans­pos­able el­e­ments, or may­be to mi­to­chon­dr­ial plas­mids, or even to self-splic­ing in­trons. Take your pick. All vi­roids form a mono­phyletic group along with the satel­lite RNAs also found in plants. Are they an­cient? Those who pic­ture early life on the planet as based on RNA, not DNA, would like to think so.

If vi­roids are an­cient, or even just a cen­tury old, why the sud­den ap­pear­ance of vi­roid dis­eases in our crops in re­cent decades? At least some vi­roids were ac­ci­den­tally in­tro­duced by hu­mans from wild rel­a­tives where of­ten their in­fec­tions are asymp­to­matic. We be­come aware of them in our crops when they cause dis­ease. Once in­tro­duced, we have in­sured that they spread. They used to have to rely pri­mar­ily on di­rect trans­mis­sion to the next gen­er­a­tion via seed or pollen. Some were oc­ca­sion­ally car­ried from one host to the next by chew­ing in­sects. Nowa­days, our prac­tice of mono-crop­ping makes in­ad­ver­tent trans­mis­sion by in­sects far more likely. Bet­ter yet, most of­ten we trans­port the vi­roids our­selves, in­tro­duc­ing them di­rectly into their next host as we prune or graft.

Why are there no vi­roids that in­fect an­i­mals? Maybe even bac­te­ria? Not a clue.

An­cient or not, vi­roids are to­day ex­pand­ing their hori­zons through global com­merce, meet­ing new hosts through mod­ern agri­cul­ture. Life is good for vi­roids to­day – if in­deed it would oc­cur to any­one to de­scribe vi­roids as alive.

Ref­er­ence

Ding B (2009). The bi­ol­ogy of vi­roid-host in­ter­ac­tions. An­nual Re­view of Phy­topathol­ogy, 47, 105–131. PMID 19400635