Small Difference, Big Effect: How a single letter changes apple resistance to a pathogen
By Céline Caseys
You enter into your favorite grocery store and you see beautiful yellow ‘Golden Delicious’ apples. The fruits are unnaturally uniform and perfect in color and shape. These apples look so good that you probably want to pick one and eat it.
Seeing these gorgeous fruits, you probably can’t imagine the other side of the mirror: an orchard with sick apple trees, leaves covered with black spots and apples with moldy brown cores. That’s the horrible scene caused by a fungal pathogen, Alternaria blotch of apple (Alternaria alternata). This opportunistic pathogen invades tissue of wild and domesticated apple trees.
Different Varieties, Different Resistance
Fungicides are the usual treatment to control the spread of this tiny fungus, but it’s a costly solution. It would be better for the farmer, consumer, and the environment if the trees were able to fight back effectively with genetics rather than applied chemistry. That’s the goal of scientists. To achieve that, they investigate resistance in existing varieties.
You may prefer the ‘Red delicious’, maybe the ‘McIntosh’ or the ‘Pink Lady. Like all of them? Good, because, like the prism that reveals beautiful colors hidden in white light, genetic diversity can reveal mechanisms and — at least the beginning of — solutions: all of these apple varieties have different susceptibility to Alternaria, the ‘Golden delicious’ being quite susceptible.
So, what does it take for an apple tree to have genetic resistance against this invader?
That is the question that researchers from the China agricultural university in Beijing have partially answered in their article in The Plant Cell .
The Genetic Jungle
All living organisms come with an instruction book, their DNA, in which the genes are the specific orders that execute their instructions through an intermediate compound called RNA. Genes come in all sorts of flavors, each providing the blueprint that contributes to plant form and function.
Resistance genes (abbreviated R-genes) play a key role to immunity, detecting dangers and, like lieutenant colonels, they guide battalions on the battlefield. The apple tree has plethora of R-genes (1015 according to the latest count ) and is, in theory, well equipped to fight back against pathogens.
Variations in these R-genes are known to affect resistance to various plant pathogens, each providing a custom system to detect and fight a specific threat. But given the diversity of R-genes in the apple genome, how can scientists identify which gene matches which pathogen? In this case, what R-gene responds to the attack by Alternaria and why does the response (resistance) vary between apple varieties?
The researchers found the answer while tracking another phenomenon: interference from one element in the genome on others. Genetics is a huge and mysterious jungle and not all players always play fair.
In this case, researchers were tracking small RNA molecules that looked like pieces of R-genes. They found that these small bits of R gene mimic and match stretches of sequence with five R-genes. When present in the cell nucleus, they block the activity of (or interfere with) five R-genes that allow the apple to be susceptible to Alternaria infection.
It happens that these small interfering RNAs are all degradation products of a single gene, MdhpRNA277 (Md stand for Malus domestica, hpRNA for hairpin RNA). This gene is suspected to have emerged from duplication events (‘copy-paste’ events) in the genome. This is a common phenomenon, especially in immunity genes, and can generate diverse mechanisms to fight disease.
In this case, the duplicated gene slowly changed over millions of years, accumulating mutations and getting shorter, ending up as a disruptor of the gene family it descended from. This gene broke free from its original purpose and started a revolution on its own.
The glitch in the apple
Things are more complex than they seem at first glance: variation in susceptibility across apple varieties does not come from the presence or absence of MdhpRNA277; they all have it. Whether an apple gets infected or not depends on how much the gene is turned on, or “expressed”.
To understand why this gene is expressed in some apple varieties but not others, the researchers sequenced the whole region before that gene, a control region that act as regulator of ‘on/off’ and ‘how much’. This area contains the promoter region, the ‘landing area’ for proteins that read and translate the gene. What they found in this area is a simple single letter switch: a ‘G’ instead of a ‘T’ in the genetic code.
When the ‘G’ is present, the protein MdWHy — responsible for translating MdhpRNA277 — can do its job and produce plenty of RNAs. These RNAs degrade into pieces and interfere with the R-genes responding to Alternaria attack, making the plant susceptible.
When the ‘T’ is present, MdWHy doesn’t “understand” the instruction, gets confused, and doesn’t produce many RNA molecules. Given the disruptive nature of the gene, this ‘T’ glitch is a good thing. The immunity remains intact and the plant can fight back.
Here is the shocking reality: A single small letter switch is the only thing needed in the DNA to fix the resistance against Alternaria to our lovely apple trees. Elementary my dear Watson!
Another piece of good news: researchers can now produce genetic tests to estimates apple tree susceptibility to Alternaria even before planting the tree in the orchard. This ensures that apple breeders can tell if a tree will require fungicides or not years before it ever flowers or produces an apple, leading to more sustainable solutions in fruit production.
Department of plant sciences
University of California, Davis
 Zhang Q, Ma C, Zhang Y, Gu Z, Li W, Duan X, Wang S, Hao L, Wang Y, Wang S, Li T (2018). A single-nucleotide polymorphism in the promoter of a hairpin RNA contributes to Alternaria alternata leaf spot resistance in apple (Malus× domestica). Plant Cell 30: 1924–1942.
 Arya, P., Kumar, G., Acharya, V., & Singh, A. K. (2014). Genome-wide identification and expression analysis of NBS-encoding genes in Malus x domestica and expansion of NBS genes family in Rosaceae. PLoS One 9: e107987.
Herman, M., and Williams, M. (2015). Fighting for their lives: plants and pathogens. Teaching Tools in Plant Biology: Lecture Notes. Plant Cell, doi/10.1105/tpc.112.tt0612
Williams, M. (2016). The small RNA world. Teaching Tools in Plant Biology: Lecture Notes. Plant Cell, doi/10.1105/tpc.110.tt021