##RNA Building Blocks

​Every strand of RNA is a sequence of four building blocks called nucleotides. Each RNA nucleotide consists of three parts: a sugar, a phosphate group, and a nitrogen-containing base.

[[☃ image 1]]The four RNA bases are adenine, uracil, guanine, and cytosine—often referred to as A, U, G, and C. RNA shares three bases in common with DNA: adenine, guanine, and cytosine. Instead of uracil, DNA contains thymine.

The sugar and phosphate groups form the backbone of a strand of RNA, and the bases bond to each other. RNA nucleotides contain a sugar called ribose, whereas DNA nucleotides contain a sugar called deoxyribose. This is spelled out in the full names of RNA and DNA: ribonucleic acid and deoxyribonucleic acid.

The four RNA bases bond together in complementary pairs. Different bases form bonds of different strengths.  Guanine-Cytosine form the strongest bond, Adenine-Uracil form the next strongest bond, and Uracil-Guanine form the weakest bond.

Comparing RNA and DNA

DNA and RNA are both strings of chemical building blocks called nucleotides. And these nucleotides spell out a genetic code. That’s about where the similarities end though; these two molecules carry out very different functions in living things.

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Structural differences

DNA is double stranded, forms a double helix (the two strands spiral around each other) is stable and stays in the nucleus. RNA is single stranded (except in rare cases), can fold into complex 3D shapes, is relatively unstable and can leave the nucleus.

Functional differences

DNA stores information as genes. Genes contain the blueprints for RNAs and proteins. Many "non-coding" RNAs act as molecular machines. They carry out various functions that keep us alive. Some of their functions include facilitating cellular processes, protecting the cell from invading viruses, and altering other biomolecules.

Messenger RNA (mRNA)

[[☃ image 1]]Messenger RNAs (mRNAs) are messengers between DNA and a cell’s protein-assembly machines (ribosomes). They carry coded instructions for building proteins.

Simple structure

Unlike most of the VirtuaLab RNAs, mRNAs do not usually fold into 3D shapes (and when they do, they have to be unfolded in order to be read by a ribosome). Instead, they resemble long twisted strings.

mRNAs form from a DNA template in a process called transcription. When an mRNA is made, it often needs to be edited before it can be translated into a protein. This is carried out by a spliceosome, a molecular machine made of RNAs and proteins, which splices out unwanted RNA from newly made mRNA.

Translating the mRNA code

The four nucleotides, A, U, C, and G, are the basis of the code in mRNA. Every set of three nucleotides corresponds to a specific amino acid (amino acids are the building blocks of proteins). For example, the code UAC, CGA, GGA on mRNA means “tyrosine, arginine, glycine.” Every three nucleotides of mRNA are recognized by a particular transfer RNA (tRNA) that carries a particular amino acid. Amino acids from tRNAs are linked together to make a protein in a cell’s protein-assembly machine (ribosome) in the order specified in an mRNA.

Transfer RNA (tRNA)

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Transfer RNAs (tRNAS) have two jobs:  1. They carry a protein building block (an amino acid) to a cell's protein assembly machine, called a ribosome. 2. They translate the genetic code of messenger RNA (mRNA) into the amino acid sequence of proteins.

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tRNAs fold up into an upside-down L shape. The top end of the L has an amino acid bound to it, and the bottom end has three bases (nucleotides) that bind to mRNA when a protein is made.  mRNA carries the genetic code for a protein in a sequence of nucleotides. Every three nucleotides of mRNA are recognized by a particular tRNA that carries a particular amino acid.

When making a protein, the ribosome holds mRNA in place. When a tRNA enters the ribosome, it binds to a complementary section of mRNA. At this moment, the tRNA releases its amino acid to be incorporated into a growing chain of amino acids that will become a protein. The empty tRNA leaves the ribosome to make space for another amino acid-carrying tRNA.

RNA Cutter (RNase)

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RNA cutters, or ribonucleases (RNases), are enzymes made of RNA that break down other RNA molecules by chopping them up into smaller pieces.

RNases perform many roles, including: getting rid of unwanted RNA, refining RNA molecules by chopping off unwanted sections of RNA, and serving as a first defense against infection from RNA viruses by breaking down viral RNA.

RNases work like recycling plants. They stop cells from accumulating unwanted RNAs by breaking them down into pieces that can be recycled into new RNA molecules.

There are two types of RNase: the first bites off one nucleotide at a time from the ends of an RNA molecule; the other works in the middle of an RNA molecule, splitting it apart or chopping out chunks of RNA.

RNases break down all the unprotected RNAs that they encounter in a cell. An RNA can protect itself from RNase attack by forming complexes with proteins or by adding particular chemical groups to either of its ends. And an RNase can be stopped in its tracks when an RNase inhibitor protein binds to it.

All living things that have been studied contain RNases. So, RNases must have evolved in early life-forms and been conserved through evolution. This tells us that breaking down RNA is a very important process.

Ribosomes and rRNA

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Ribosomes are a cell’s protein-assembly machines. Their job is to link protein building blocks (amino acids) together to make proteins in an order spelled out in messenger RNA (mRNA).

Two subunits

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Ribosomes are made of two parts: a small subunit and a large subunit. These subunits are made of ribosomal RNA (rRNA) and proteins.

Protein-making machines

When making a protein, the small ribosome subunit holds mRNA in place. Transfer RNAs (tRNA) enter the ribosome carrying individual amino acids. The large ribosome subunit bonds these amino acids together to make a protein. When a tRNA releases its amino acid, it exits the ribosome and another tRNA takes its place to add its amino acid to the growing protein. When a ribosome finishes translating an mRNA, the small and large ribosome subunits split apart.

It’s the rRNA in the ribosome that links amino acids together when making a protein, which means that rRNA is an RNA enzyme. The role of the ribosome proteins seems to be to stabilize the ribosome structure.

All living things that have been studied contain rRNA. So rRNA must have evolved in early life-forms and been conserved through evolution. Evolutionarily speaking, rRNAs are far older than the proteins in a ribosome. This suggests that in very early life-forms, ribosomes operated without proteins to stabilize them.

This is good evidence for the RNA World hypothesis, which suggests that RNA allowed life to form on Earth, and that it came before DNA and proteins. Evolution of rRNA as a cell’s protein-assembly machine would have made the existence of proteins possible.

mRNA-Splicing Machine

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mRNA-splicing machines chop out unwanted sections of newly made messenger RNA (mRNA) to create mature mRNA that can be translated to make a protein.

When a strand of mRNA is first copied from DNA, the genetic code it carries isn’t ready to be translated to make a protein. First, it needs to be edited by a machine made of RNAs and proteins, called a spliceosome. It’s like an editor that proofreads the first draft of an mRNA. Like scissors and glue, it chops out unwanted sections of mRNA, called introns, and sticks the remaining pieces back together again. This is called splicing.

A single mRNA copied from one gene can code for lots of different proteins if it’s spliced in different ways. This explains how humans are able to produce about 100,000 proteins from only about 20,000 genes.

Self-Copying RNA

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The RNA world hypothesis suggests that life on Earth began with a simple RNA molecule that could copy itself without help from other molecules.

DNA, RNA, and proteins are central to life on Earth. DNA stores the instructions for building living things—from bacteria to bumble bees. And proteins drive the chemical reactions needed to keep cells alive and healthy. Until recently, RNA was thought of as little more than a messenger between DNA and proteins, carrying instructions as messenger RNA (mRNA) to build proteins. However, RNA can do far more. It can drive chemical reactions, like proteins, and carries genetic information, like DNA. And because RNA can do both these jobs, most scientists think life as we know it began in an RNA world, without DNA and proteins.

The first RNAs

So how did RNA evolve on Earth? Scientists think RNA building blocks (nucleotides) emerged in a chaotic soup of molecules on early Earth. These nucleotides bonded together to make the first RNAs. No sooner were they made than they broke down; however, new ones were made in their place. Some RNAs turned out to be more stable than others. These RNA strands grew longer and bonded nucleotides more quickly. Eventually, RNA strands grew faster than they broke down. This was RNA’s chance to start life.

RNA offspring

All living things reproduce. They copy their genetic information and pass it onto their offspring. And for RNAs to start life, they needed to reproduce too. This is why scientists think that the RNA world took off when an RNA emerged that could make copies of itself. As it did, new self-copying RNAs emerged. Some were better at copying themselves than others. The RNAs competed against each other, and the most successful won out. Over millions of years, these RNAs multiplied and evolved to create an array of RNA machines. At some stage, DNA and proteins evolved. Proteins began to drive chemical reaction in cells, and DNA—which is more stable than RNA—took the job of storing genetic information.

Nucleotide-Building RNA

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Maintaining enough RNA building blocks (nucleotides) would have been a top priority in the RNA world. Scientists think nucleotide-building RNAs evolved on early Earth to provide nucleotides for building new RNAs.

Supplying the RNA world

According to the RNA world theory, the first RNAs were made using free-floating nucleotides that emerged in a primordial soup of molecules. They bonded together to make strands of RNA that weren’t very stable and degraded quickly. But some were more stable than others; these RNAs grew longer and bonded nucleotides more quickly. Eventually, RNA strands grew faster than they broke down—and this was RNA’s foot in the door. Over millions of years, these RNAs multiplied and evolved to create an array of RNA machines that are the basis of life as we know it today. But for RNA molecules to take hold, they would have needed an abundant supply of nucleotides. And scientists think nucleotide-building RNAs evolved to provide these RNA building blocks.

Test tube evolution

Scientists attempting to re-create the conditions of early Earth in a test tube have managed to evolve a number of RNA machines that can drive chemical reactions to make some parts of a nucleotide. This proves that RNA can drive nucleotide-building chemical reactions. But researchers have yet to create an RNA machine that can create whole nucleotides using ingredients that would have been available on primitive Earth.

Grabber RNA

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In an RNA world, scientists think that simple RNAs grabbed onto other RNAs or molecules to form complexes that could change, or enhance, their function. This was a step towards more complex life.

Ribosomes, which are a cell’s protein-assembly machines, are made of ribosomal RNAs (rRNAs) and proteins. But the rRNAs in a ribosome evolved long before ribosomal proteins. Back in the RNA world, it is possible that one RNA may have grabbed onto another RNA to create an RNA machine that—for the first time ever—linked amino acids together to make a protein. Thus, the first version of a ribosome emerged.

Some messenger RNAs (mRNAs) in bacteria and some plants contain a section of code called a riboswitch that can grab onto a specific molecule. Binding this molecule controls whether the mRNA is translated to make a protein. The molecule could be a nutrient that binds to an mRNA riboswitch and triggers the mRNA to be translated to make a protein that breaks this nutrient down. So mRNAs that contain riboswitches can regulate themselves in response to specific molecules. It was previously thought that only proteins regulated the production of protein from mRNA; however, riboswitches hint of a regulation system that may have existed in an RNA world long before proteins existed.

Protein-Directing RNA

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Our cells contain a protein-directing machine, made of RNA and proteins, that sends newly made proteins to where they’re needed in a cell. This machine is called the signal recognition particle (SRP).

The SRP is on the lookout for proteins being made in a cell’s protein assembly machine (ribosome). When it spots the beginning of a protein poking out of the ribosome, it binds to it. The ribosome halts protein production while the SRP brings the ribosome and its partly-built protein to where it’s needed in the cell. On arrival, the SRP is released and protein synthesis starts up again.

The RNA in the SRP is found in all living things, which suggests that it evolved in very early life-forms. When proteins first emerged on Earth, an early version of this protein-directing RNA may have helped organize proteins in a cell. It could have enhanced primordial cells by directing proteins to form a cytoskeleton. A cytoskeleton helps a cell keep its shape and is like a highway system for transporting molecules around a cell.

Gene-Manipulating RNA

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One messenger RNA (mRNA) can be remixed in different ways so that its genetic code can be translated to make lots of different proteins. The ability to make more than one protein from one mRNA sped up the evolution of multicellular life.

Newly made mRNA is spliced by a molecular machine called the spliceosome that is like the scissors and glue of the cell. Made of RNAs and proteins, this machine chops out unwanted sections of mRNA code and sticks the remaining mRNA back together again, to create mature mRNA that can be translated to make a protein.

One gene and lots of proteins

Back in the 1970s, scientists thought that one gene coded for one mRNA, which in turn coded for one protein. This is largely true for bacteria and other single-celled life; however, for multicellular life one gene codes for one mRNA that can be spliced in different ways to create many different proteins. This is called alternative splicing.

Alternative splicing is an ingenious way of creating a diverse range of proteins from a relatively small number of genes. One of the surprises of the Human Genome Project was that the human genome codes for so few genes. Scientists predicted that there would be about 100,000 human genes, but the number is closer to 20,000. These genes are spliced in different ways to create a huge number of human proteins.

Speeding up evolution

The arrival of alternative splicing in multicellular life-forms likely sped up evolution. It meant that an organism could create new proteins without going through the lengthy process of evolving new genes. Chance mutations would have arisen that caused existing mRNAs to be spliced in different ways. These alternatively spliced mRNAs coded for brand new proteins that may have driven brand-new cellular processes, driving forward the evolution of complex life.

Cell-Growth-Directing RNA

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RNA machines were likely central to the evolutionary leap from single-celled to multicellular life forms.

Multicellular life starts with a single fertilized egg. This cell divides into two cells, which divide again… and so it goes on. Soon, the cells in this developing life-form start carrying out different jobs. In a plant, they could become leaf cells or root cells. In an animal, they could become blood cells or nerve cells. There are about 200 different types of cells in a human, and it is essential that each is made in the right place at the right time in a human embryo. What a cell becomes is determined by what molecular machines—RNAs and proteins—are operating in that cell. And those RNAs and proteins that are present in a cell are determined by transcription factors that switch genes on and off.

Telling heads from tails

We get clues about the molecular machines that could have driven the leap to multicellular life by studying how embryos develop in organisms now. A favorite creature for developmental biologists is the fruit fly. And an important RNA in fruit fly development is called bicoid. It plays a vital role in organizing the body plan of a developing fruit fly. In an unfertilized fruit fly egg, bicoid RNA is found in the end of the egg that will become the fly’s head. Once the egg is fertilized, the bicoid mRNA is translated to make a protein. Bicoid protein switches on genes that make head-making proteins and switches off genes that make tail-making proteins. So, bicoid tells fruit fly embryos exactly where to make the head.

Viral RNA

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Viral RNA contains the blueprints for all of the components of a particular virus. Examples of RNA viruses include the flu, polio, measles, and SARS.

What is a virus?

A virus is a collection of genetic material (DNA or RNA) inside a protein shell. There is much scientific debate about whether viruses are alive. Like living creatures, they carry genes and evolve. But they are unable to reproduce on their own and must infect living cells and hijack cellular machinery (such as ribosomes) in order to copy themselves.

Though viruses are tiny, they are fierce. Influenza (the flu) alone has killed tens of millions of humans in the last 100 years.

Viral genes

Viruses carry their genetic information as either DNA or RNA. RNA is less stable and more prone to mutation than DNA, so RNA viruses generally change and evolve faster. This sometimes allows RNA viruses to jump from species to species (notable recent examples include the avian flu and swine flu, which jumped to humans from birds and pigs, respectively).

Viruses hijack cellular machinery in many ways, but in almost all cases, the virus tricks the cell into treating the virus’s genetic information as its own and transcribing and/or translating it. In those instances, perhaps the best way the cell can defend itself is to learn to recognize and destroy viral genes.

Virus-Packing RNA

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pRNA, a component of the strongest nanomotor on Earth, packs genetic information into a virus’s protein shell.

When a virus hijacks a cell, it forces the cell to produce copies of the virus. This process includes creating capsids—the outer protein shell of a virus. The genetic information of some viruses is so long and complex that it has to be folded and packed tightly to fit inside the capsid. This requires energy—it’s like trying to put a Jack-in-the-box back inside the box.

ATP: The molecular battery

The energy comes in the form of ATP (adenosine triphosphate), a molecule that stores energy in the bonds between its atoms. ATP acts like a rechargeable battery, carrying energy throughout the cell to power its molecular machines. But to deliver the energy, ATP needs something to bind to.

Gears in a motor

This is where pRNA, or packaging RNA, comes into play. pRNA molecules together with ATPase motors bind ATP molecules and extract their energy. By using this energy, pRNAs form a ring that acts like a gear in a powerful motor, stuffing the genetic strands into the capsid and assembling the virus.

Viral RNA Chopper and Destroyer

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The RNA chopper and RNA destroyer are parts of a cell’s pathway for recognizing and destroying viral RNA.

In the Virus chapter of NOVA’s RNA Lab game, the process used to fight invading viruses is based on an immune response observed in bacteria, plants, and some animals (but not yet in humans) called RNA interference (RNAi). RNAi has two main steps.

RNAi Step 1: Intruder detection
Specially evolved RNAs and proteins detect strands of invading viral RNA and chop them up.

RNAi Step 2: Border guards
Another set of RNAs and proteins pick up some of the chopped-up viral RNA from Step 1. You can think of them as setting up security checkpoints near a cell’s ribosome, where translation occurs. These molecular border guards compare incoming messenger RNA (mRNA) that comes from either the virus or the host cell against their small pieces of the virus’s genetic code. Anything that matches codes for something that the virus is trying to make, so they slice it up before it gets through.

RNA chopper

The “RNA chopper” in the NOVA RNA Lab stands in for the whole group of RNAs and proteins involved in Step 1—detecting and chopping up invading viral RNA. This design is modeled on an RNA found in mosquitoes that helps identify invading RNA from the dengue virus.

RNA destroyer

Cells can sometimes use a process similar to RNAi to quickly turn off their own genes. The ”RNA destroyer” that players design in the NOVA RNA Lab comes from an RNA involved in one such process in nematodes (roundworms). This RNA (called microRNA or miRNA) finds nematode mRNA with a matching sequence and signals to proteins that they should destroy this mRNA. This silences the target gene because its mRNA is destroyed before it is translated to make a protein.

NOVA’s RNA destroyer is doing double duty in a way that probably wouldn't happen in real cells—our RNA destroyer is both detecting viral mRNA and cutting it up. Proteins would normally carry out the actual destruction/cutting.

Two-Shape RNA Switch

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A special class of RNAs called “riboswitches” can switch from one shape to another when they detect specific molecules.

When it comes to riboswitches, the structural weakness of RNA is its greatest strength. Because some of RNA’s bonds (such as U-Gs) are weak, they can easily break and allow the RNA to transform. Molecules like DNA and proteins can’t transform as easily or as drastically as RNA, because their structures are much more stable.

Riboswitches “switch” shapes when they detect a specific molecule. Their 3D shape fits that molecule in such a way that they naturally bind to it. But binding to the molecule changes the way that RNA folds, because some of the base pairs will no longer be able to reach each other. This means that the RNA breaks some of its bonds and takes on a new 3D shape. Since molecular shape determines molecular function, that RNA now has a different function in the cell.

It’s like the effect that the light of the full moon has on a werewolf.* Just by being exposed to moonlight, a harmless human undergoes a rapid transformation to a feral creature. Riboswitches do not become monsters, but their transformations are swift, easily triggered, and functionally drastic.

*NOVA does not endorse, suggest, or encourage the existence of werewolves.

RNA computers

The processes shown in the last two sections of the Virus chapter in NOVA's RNA Lab are not yet being used for medicine, but they demonstrate a possible, and promising, future treatment that could fight viruses, cancer, and who knows what else.

Here’s a simplified version of how it might work. Let’s say you want to design an RNA-based treatment for HIV (the virus that leads to AIDS).

Step 1: Identify a molecule that is only present in cells that have been infected by HIV. We’ll call it Molecule X.
Step 2: Design a riboswitch that can detect Molecule X by binding to it and switching shape.
Step 3: Tweak your RNA switch so that when it switches to its second shape, that shape activates “cell death.” Cell death is a natural process that your body already uses to eliminate sick cells.

The same principle might be used to fight cancer—but instead of finding Molecule X, the RNAs look for Molecule Y, which is only found in cancerous cells.

And why stop with just one RNA switch at a time? Why not design a whole RNA computer with many different RNA switches that can look for many different molecules in the cell and respond accordingly?

Perhaps RNA engineers could program a single RNA computer that fights viruses, defeats cancer, and trims out genetic defects from DNA. It sounds fanciful, and may be decades away, but scientists are exploring this as a real possibility. And if it works, it will change the world.