Leishmania species cause a spectrum of human diseases in tropical and subtropical regions of the world. We have sequenced the 36 chromosomes of the 32.8-megabase haploid genome of Leishmania major (Friedlin strain) and predict 911 RNA genes, 39 pseudogenes, and 8272 protein-coding genes, of which 36% can be ascribed a putative function. These include genes involved in host-pathogen interactions, such as proteolytic enzymes, and extensive machinery for synthesis of complex surface glycoconjugates. The organization of protein-coding genes into long, strand-specific, polycistronic clusters and lack of general transcription factors in the L. major, Trypanosoma brucei , and Trypanosoma cruzi (Tritryp) genomes suggest that the mechanisms regulating RNA polymerase II–directed transcription are distinct from those operating in other eukaryotes, although the trypanosomatids appear capable of chromatin remodeling. Abundant RNA-binding proteins are encoded in the Tritryp genomes, consistent with active posttranscriptional regulation of gene expression.

All organisms adapt to changes in their environment by adjustments in gene expression, and in all organisms, from Escherichia coli to man, the most important control point is at transcription initiation. All, that is, except those belonging to one very small family of early‐branching eukaryotes, which seems to have completely lost the ability to regulate transcription by RNA polymerase II. The organisms concerned are unicellular, spindle‐ like flagellates that flourish in the digestive systems of arthropods, in the blood, macrophages and brains of vertebrates from humans to lizards, and even in the sap of coconut palms and lemon trees. Many of them are able to multiply both in a vertebrate (or plant) and an invertebrate, which serves to transmit the parasites from one vertebrate (or plant) to the next. Adaptation to the two distinct environments, with different temperatures, nutrients and defences, requires major changes in gene expression. Yet this seems to be achieved in the total absence of any developmental regulation of RNA polymerase II; perhaps even without any specific polymerase II transcription initiation. This extraordinary state of affairs might be written off as an irrelevant evolutionary quirk (and, indeed, might have even gone unnoticed) if it were not for the fact that some of the organisms concerned, the trypanosomes and the leishmanias, kill millions of people every year (http://www.who.ch). The leishmanias cause a spectrum of diseases ranging from self‐resolving skin ulcers to lethal infection of the internal organs. One‐and‐a‐half to two million people are newly infected every year in the tropics and southern Europe. Leishmania has an extracellular form in the gut of the vector, the sand‐fly, but multiply as spherical aflagellate forms within the lysosomes of mammalian macrophages. Leishmania must be phagocytosed without activating the host macrophage and must combat oxidative, acidic and proteolytic stresses. The South …

In trypanosomes, RNA polymerase II transcription is polycistronic and individual mRNAs are excised by trans -splicing and polyadenylation. The lack of individual gene transcription control is compensated by control of mRNA processing, translation and degradation. Although the basic mechanisms of mRNA decay and translation are evolutionarily conserved, there are also unique aspects, such as the existence of six cap-binding translation initiation factor homologues, a novel decapping enzyme and an mRNA stabilizing complex that is recruited by RNA-binding proteins. High-throughput analyses have identified nearly a hundred regulatory mRNA-binding proteins, making trypanosomes valuable as a model system to investigate post-transcriptional regulation.

In Kinetoplastida, trypanothione and trypanothione reductase (TRYR) provide an intracellular reducing environment, substituting for the glutathione–glutathione reductase system found in most other organisms. To investigate the physiological role of TRYR in Trypanosoma brucei, we generated cells containing just one trypanothione reductase gene, TRYR, which was under the control of a tetracycline‐inducible promoter. This enabled us to regulate TRYR activity in the cells from less than 1% to 400% of wild‐type levels by adjusting the concentration of added tetracycline. In normal growth medium (which contains reducing agents), trypanosomes containing less than 10% of wild‐type enzyme activity were unable to grow, although the levels of reduced trypanothione and total thiols remained constant. In media lacking reducing agents, hypersensitivity towards hydrogen peroxide (EC₅₀ = 3.5 μM) was observed compared with the wild type (EC₅₀ = 223 μM). The depletion of TRYR had no effect on susceptibility to melarsen oxide. The infectivity and virulence of the parasites in mice was dependent upon tetracycline‐regulated TRYR activity: if the trypanosomes were injected into mice in the absence of tetracycline, no infection was detectable; and when tetracycline was withdrawn from previously infected animals, the parasitaemia was suppressed.

An inducible expression system was developed for the protozoan parasite Trypanosoma brucei . Transgenic trypanosomes expressing the tetracycline repressor of Escherichia coli exhibited inducer (tetracycline)-dependent expression of chromosomally integrated reporter genes under the control of a procyclic acidic repetitive protein (PARP) promoter bearing a tet operator. Reporter expression could be controlled over a range of four orders of magnitude in response to tetracycline concentration, a degree of regulation that exceeds those exhibited by other eukaryotic repression-based systems. The tet repressor-controlled PARP promoter should be a valuable tool for the study of trypanosome biochemistry, pathogenicity, and cell and molecular biology.