RNA

• 1. Introduction

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  Ribonucleic acid, known as RNA, is a polymeric molecule essential for most biological functions. It works as a chain of nucleotides, sometimes building proteins and sometimes performing jobs itself. Nucleic acids count among the four major macromolecules essential for all known forms of life. Yet RNA has long lived in the shadow of its more famous cousin, DNA. The two are close chemical relatives. They differ in only a handful of ways. But those small differences carry enormous weight.

  Consider a strange idea that has become widely accepted in science. Early in the history of life on Earth, before DNA and possibly before protein-based enzymes existed, an RNA world may have existed. In that world, RNA both stored genetic information and catalyzed reactions. Today DNA holds the genes and protein enzymes run the chemistry. RNA was once asked to do both jobs at once. How can a single molecule store a blueprint and also act as a tool? Why did nature settle on exactly four chemical letters and not more or fewer? And how did a molecule once dismissed as fragile become the basis of vaccines given to people during a global pandemic? Those are the questions ahead.

• 2. Four Letters

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  Each nucleotide in RNA carries a ribose sugar, its carbons numbered 1 prime through 5 prime. A base attaches at the 1 prime position, and a phosphate group bridges the 3 prime position of one ribose to the 5 prime position of the next. Those phosphate groups each hold a negative charge. That makes the whole molecule a polyanion, a charged chain that needs help to hold its shape.

  The bases come in four kinds: adenine, cytosine, guanine, and uracil, written as A, C, G, and U. Adenine and guanine are purines. Cytosine and uracil are pyrimidines. Cytosine pairs with guanine, and adenine pairs with uracil through standard hydrogen bonds. Guanine and uracil can also join through a non-canonical pairing called the G-U wobble. Other arrangements exist too, such as the GNRA tetraloop built around a guanine-adenine pair.

  Four bases is not an arbitrary number. To design RNA for any given secondary structure, two or three bases would not be enough. Four bases are enough. This is likely why nature chose a four-base alphabet. Fewer than four would not allow the creation of all structures, while more than four are not necessary. Because RNA is charged, metal ions such as magnesium are needed to stabilize many of its folded forms.

• 3. The DNA Difference

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  Three primary differences separate RNA from DNA, and each one matters. RNA is usually a single-stranded molecule in many of its biological roles, with much shorter chains of nucleotides than DNA. It can still form double-stranded RNA, and a single strand can fold back and pair with itself to make intrastrand double helices, as happens in transfer RNA.

  The sugar in the backbone tells the second part of the story. DNA contains deoxyribose, while RNA contains ribose. Ribose carries a hydroxyl group at the 2 prime position of the pentose ring, and deoxyribose does not. That single hydroxyl group makes RNA more chemically labile than DNA by lowering the activation energy of hydrolysis. In flexible regions not locked into a helix, that 2 prime hydroxyl can even attack the adjacent phosphodiester bond and cut the backbone.

  The third difference lives in the bases themselves. In DNA, adenine's partner is thymine. In RNA, that partner is uracil, an unmethylated form of thymine. The presence of the 2 prime hydroxyl also pushes RNA helices toward the A-form geometry, which produces a very deep and narrow major groove and a shallow and wide minor groove. Folded RNAs are not built from long double helices like DNA. They pack short helices together into shapes that resemble proteins, and that resemblance lets them do something proteins do.

• 4. RNA As Machine

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  Determination of the structure of the ribosome revealed a startling fact: its active site is composed entirely of RNA. The ribosome is an RNA-protein complex that catalyzes the assembly of proteins, yet the chemistry happens on RNA. An RNA molecule that catalyzes reactions is called a ribozyme. The ribosome is exactly that, a ribozyme, and it remains the notable exception to the rule that protein enzymes run the cell's chemistry.

  Protein synthesis is a universal function, and three kinds of RNA carry it out together. Messenger RNA carries information from DNA to the ribosome, with every three nucleotides forming a codon that corresponds to one amino acid. Transfer RNA, a small chain of about 80 nucleotides, delivers a specific amino acid to the growing chain and recognizes codons through an anticodon region. Ribosomal RNA is the catalytic component that hosts translation and links the amino acids together.

  Eukaryotic ribosomes contain four different rRNA molecules: the 18S, 5.8S, 28S, and 5S. Three of them are synthesized in the nucleolus and one elsewhere. Nearly all the RNA found in a typical eukaryotic cell is rRNA. Several ribosomes may be attached to a single mRNA at once. In bacteria and plastids, a molecule called transfer-messenger RNA steps in when something goes wrong, tagging proteins from mRNAs that lack stop codons for degradation so the ribosome does not stall.

• 5. The Silent Majority

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  About 97 percent of the transcriptional output in eukaryotes is non-protein-coding. That figure overturns an old assumption, since most RNA in these cells never spells out a protein at all. These non-coding RNAs can come from their own dedicated genes or derive from the introns of messenger RNA. Length offers one way to sort them. Small RNAs run shorter than 200 nucleotides, while long RNAs run greater than 200.

  The earliest known regulators of gene expression were proteins, the repressors and activators that bind short sites within enhancer regions. Later studies showed that RNAs regulate genes too. RNA interference represses genes after transcription, using the RNA-induced silencing complex and a microRNA guide to either degrade an mRNA or block its translation. Long non-coding RNAs work differently, shutting down whole blocks of chromatin. Xist and other long noncoding RNAs were linked to X chromosome inactivation, and Jeannie T. Lee and others showed they silence chromatin by recruiting the Polycomb complex.

  Bacteria and archaea were once thought to lack such systems. As soon as researchers looked, they found them. Enterobacterial small RNAs appear in stress responses to membrane stress, starvation, phosphosugar stress, and DNA damage. Riboswitches change shape when they bind metabolites, gaining or losing the ability to regulate genes. And the CRISPR system, now used to edit DNA in place, originally acts through regulatory RNAs in archaea and bacteria to defend against virus invaders.

• 6. Made And Remade

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  Synthesis of RNA usually occurs in the cell nucleus, catalyzed by RNA polymerase using DNA as a template in a process called transcription. The enzyme binds a promoter sequence, unwinds the double helix, then moves along the template strand in the 3 prime to 5 prime direction while building the new RNA in the 5 prime to 3 prime direction. The DNA sequence also dictates where synthesis will stop.

  Primary transcripts rarely stay as first written. A poly(A) tail and a 5 prime cap are added to eukaryotic pre-mRNA, and introns are removed by the spliceosome. RNA is transcribed with only four bases, but those bases and sugars can be modified in numerous ways as the RNA matures. Pseudouridine changes the linkage between uracil and ribose from a C-N bond to a C-C bond. Inosine, a deaminated adenine, plays a key role in the wobble hypothesis of the genetic code. There are more than 100 other naturally occurring modified nucleosides.

  Some RNAs exist to modify other RNAs. Small nuclear RNAs sit inside the spliceosomes that cut out introns, and some introns are ribozymes that splice themselves. In eukaryotes, small nucleolar RNAs of 60 to 300 nucleotides guide enzymes to specific spots by base pairing, directing nucleotide modifications. In ribosomal RNA, many post-transcriptional modifications cluster in highly functional regions like the peptidyl transferase center, hinting that they matter for normal function.

• 7. Genomes Of RNA

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  Like DNA, RNA can carry genetic information, and many viruses encode their genes in an RNA genome. Some of the virus's own proteins replicate that genome while others protect it as the particle moves to a new host cell. Viroids strip the idea even further. They consist only of RNA, encode no protein, and are replicated by a host plant cell's polymerase.

  Reverse transcription runs the usual flow of information backward. Reverse transcribing viruses make DNA copies from their RNA, which are then transcribed back into new RNA. Retrotransposons spread by copying DNA and RNA from one another. Telomerase, the enzyme that builds the ends of eukaryotic chromosomes, contains an RNA used as its template.

  Double-stranded RNA carries two complementary strands, much like cellular DNA but with uracil in place of thymine and an added oxygen atom. It forms the genetic material of some viruses, and in eukaryotes it can trigger RNA interference and help activate the innate immune system against viral infections. In vertebrates it can also set off an interferon response. In the late 1970s, researchers found a covalently closed, circular form of RNA expressed across the animal and plant kingdoms. The function of these circular RNAs is largely unknown, though a few act as microRNA sponges.

• 8. A Century Of Discovery

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  Nucleic acids were discovered in 1868 by Friedrich Miescher, who called the material nuclein because he found it in the nucleus. The role of RNA in protein synthesis was suspected as early as 1939. In 1956, Alex Rich and David Davies hybridized two separate strands of RNA to form the first crystal of RNA whose structure could be solved by X-ray crystallography. Severo Ochoa won the 1959 Nobel Prize in Medicine, shared with Arthur Kornberg, though the enzyme he found was later shown to degrade RNA rather than build it.

  Robert W. Holley sequenced the 77 nucleotides of a yeast tRNA in 1965, winning the 1968 Nobel Prize shared with Har Gobind Khorana and Marshall Nirenberg. In the early 1970s, David Baltimore, Renato Dulbecco, and Howard Temin discovered retroviruses and reverse transcriptase, earning a Nobel in 1975. In 1976, Walter Fiers and his team determined the first complete nucleotide sequence of an RNA virus genome, that of bacteriophage MS2. Introns and RNA splicing, found in 1977, brought a 1993 Nobel to Philip Sharp and Richard Roberts.

  Catalytic RNA molecules discovered in the early 1980s led to a 1989 Nobel for Thomas Cech and Sidney Altman. In 1990, work in Petunia showed that introduced genes can silence the plant's own, a result of RNA interference. Andrew Fire and Craig Mello won a Nobel in 2006 for studies of RNA interference, and Roger Kornberg won that same year for the transcription of RNA. In 2009, Venki Ramakrishnan, Thomas A. Steitz, and Ada Yonath were honored for the atomic structure of the ribosome. The story reaches the present with the 2023 Nobel awarded to Katalin Kariko and Drew Weissman for modified nucleosides that enabled effective mRNA vaccines against COVID-19, whose first successful large-scale use came during the COVID-19 pandemic.