The New Microbiology. Pascale Cossart

the sugar ribose (which gives RNA its name, ribonucleic acid), whereas DNA has deoxyribose (which likewise gives DNA its name, deoxyribonucleic acid).

      Replication begins in the part of a chromosome called the origin of replication and moves in two directions along the chromosome. As the chromosome is composed of two strands of DNA, each is thus duplicated. Once replication begins, the whole chromosome is replicated. Transcription, in contrast, is a process that moves in one direction only. It can begin at any point in a chromosome but only “upstream” of the gene(s) being transcribed, in regions that François Jacob and Jacques Monod named promoters. Only certain regions of each DNA strand are transcribed onto RNA. The RNA transcript—the mRNA—is then read during translation.

Figure 5. Schematic representation of a double strand of DNA, of its transcribed messenger RNA (mRNA), and of the small protein encoded by the mRNA.

      Translation is a fairly sophisticated process that is achieved by several bacterial actors and in particular a big machine called the ribosome. Ribosomes read the nucleotide sequence of the mRNA transcript by recognizing successive triplets of nucleotides or codons. Each of the 64 possible triplets of nucleotides (AUG, UAC, or ACC, for example) corresponds to one of the 20 amino acids that are the building blocks for proteins; thus each amino acid can be encoded by more than one codon. This genetic code is the same for all bacteria and most other organisms. Thus, from their DNA, bacteria produce RNA that is ultimately translated into proteins by this universal code. Bacteria produce thousands of mRNAs, each of which produce proteins.

      The results that led François Jacob, André Lwoff, and Jacques Monod to their 1965 Nobel Prize in Physiology or Medicine concerned the discovery that several successive genes, generally involved in the same physiological function, are transcribed together from a single promoter in a single mRNA and thus form a cluster of coregulated genes that they termed an operon.

      Transcription is not a permanent process. It depends on many environmental factors (such as pH or temperature). It also depends on bacterial factors. In the simplest situation, transcription is regulated by a single “regulator” protein encoded by a gene situated elsewhere on the chromosome. This regulator interacts with the DNA region located upstream of the first gene of the operon; it can either prevent (“repress”) or stimulate the expression of mRNA. This model of regulation was studied by François Jacob and his colleagues in the lactose (lac) operon, which is involved in the utilization of the sugar lactose, in Escherichia coli. The lac operon in E. coli encodes, among other things, the lactose-metabolizing protein LacZ. Transcription of the lac operon is generally repressed, or silenced, because the protein LacI (also called the Lac repressor) binds to the chromosome upstream of the lac operon genes and prevents that DNA from being read. When lactose is present in the bacterium’s environment, the organism can convert it into allolactose, a molecule that binds to and alters the form of the repressor LacI protein (a conformation, or allosteric change) and prevents it from binding to the DNA. In the absence of LacI repression, the lac operon genes are transcribed and then translated, allowing the organism to metabolize lactose.

Many researchers have forgotten that in the original model for the operon published in the Journal of Molecular Biology in 1961, Jacob and Monod proposed that the repressor is the product of a regulatory gene—that is, an RNA repressor that would act either in a region of the DNA upstream of the operon, to repress its transcription, or at the start of the operon mRNA, to prevent translation of the message (Fig. 6). Following the publication of this model, researchers analyzed in detail the regulation of the lactose operon and found that the Lac repressor is not in fact an RNA molecule but instead a protein that works upstream of operon genes, as discussed earlier. For years, this discovery was followed by numerous successful discoveries of other repressors, forgetting a key part of Jacob and Monod’s hypothesis— repressors could also be RNA.

      Repressors and activators

      The operon model has been shown to occur in all bacteria, other prokaryotes, and some eukaryotes (particularly in worms such as the nematode Caenorhab-ditis elegans). It has thus been refined and enriched. We now know that genes can be repressed by a variety of repressors that more or less resemble the Lac repressor. Certain genes can be repressed by several repressors but others may also be activated. In this case, it is not a negative regulation that occurs, as for the lac operon, but a positive regulation, with an activator protein that binds upstream of the operon when specific conditions require genes to be expressed. One of the best-known activator proteins in bacteria is the protein CAP or CRP in Escherichia coli, which binds cyclic AMP, a molecule that acts as a hormone and can activate genes involved in the use of sugars other than glucose in bacteria.

      Some repressors and activators present in bacteria can also be found in bacteriophages, viruses that attack bacteria. Bacteriophages, or phages for short, are essentially made of DNA and a few proteins packaged within an outer coat of proteins. The protein cl is a major player in the life of the phage lambda.

      Phages, such as lambda phage, can be lytic or lysogenic. A phage is lytic if it causes lysis (from the Greek lysein, meaning to dissolve) of the infected host cell. Lysis occurs when a phage injects its DNA into a bacterium. The DNA circularizes and proceeds to replicate into new bacteriophages until the sheer number of them makes the bacterium explode. In contrast, a phage is lysogenic if the phage's DNA, injected into the bacterium, becomes integrated into the host bacterial cell's chromosome and becomes silent. In some conditions, it can excise and again act as a lytic phage.

      The cI protein is a regulator at the heart of both the lytic and lysogenic situations in the lambda phage. cI is able to behave like a repressor when the phage DNA integrates itself into the bacterial genome. cI inhibits the expression of bacterial excision genes that allow the phage DNA to excise from the bacterial chromosome. If bacteria are stressed, irradiated, or grown in certain nutrient deficiencies, the bacterial protein RecA is activated. It cleaves the cI protein, allowing excision to occur.

Figure 6. (Left) The three 1965 Nobel Prize winners François Jacob, Jacques Monod, and André Lwoff. (Right) Jacob, Monod, and Lwoff's famous model proposing two possible alternatives for a simultaneous repression of the three genes in the lactose operon. Note that RNA here plays the role of the repressor.

      However, in the beginning of the 1980s, it was found that the replication of plasmids—small circular minichromosomes that carry certain accessory genes, such as those for virulence or antibiotic resistance—is controlled by small RNAs that attach (or hybridize) to single strands of the plasmid’s DNA and prevent the plasmid’s replication. The concept of antisense RNA was thus born and the RNA revolution began.

      Several other antisense RNAs were then discovered in bacteria, but no one could have foreseen the explosion of knowledge in the early 2000s following the discovery of microRNAs in eukaryotes. MicroRNAs are small RNAs with 22 nucleotides that attach to the 3' region of eukaryotic RNA and affect its translation. This surge of new discoveries was made possible by revolutionary new technologies, in particular the DNA chips called tiling arrays and new ultrarapid sequencing methods for genomes and RNA. These techniques allowed for the analysis of the ensemble of RNA transcripts of bacteria grown under a variety of conditions and led to the discovery that bacteria express a large number of RNA transcripts that do not encode proteins. These transcripts, called noncoding RNAs, are often the product of the intergenic regions of DNA and can act as regulators. Bacteria can have as many as several hundred distinct noncoding RNAs.

      Many