Identification and phylogenetic analysis of heme synthesis genes in trypanosomatids and their bacterial endosymbionts

Abstract

It has been known for decades that some insect-infecting trypanosomatids can survive in culture without heme supplementation while others cannot, and that this capability is associated with the presence of a betaproteobacterial endosymbiont in the flagellate's cytoplasm. However, the specific mechanisms involved in this process remained obscure. In this work, we sequence and phylogenetically analyze the heme pathway genes from the symbionts and from their hosts, as well as from a number of heme synthesis-deficient Kinetoplastida. Our results show that the enzymes responsible for synthesis of heme are encoded on the symbiont genomes and produced in close cooperation with the flagellate host. Our evidence suggests that this synergistic relationship is the end result of a history of extensive gene loss and multiple lateral gene transfer events in different branches of the phylogeny of the Trypanosomatidae.

Figure 1. Synthetic pathway for heme.

Enzymes: gltX, glutamyl-tRNA synthetase (EC:6.1.1.17); hemA, glutamyl-tRNA reductase (EC:1.2.1.70); GSA, glutamate-1-semialdehyde 2,1-aminomutase (EC:5.4.3.8); ALAD, aminolevulinic acid dehydratase (EC:4.2.1.24); PBGD, porphobilinogen deaminase (EC:2.5.1.61); UROS, uroporphyrinogen III synthase (EC:4.2.1.75); UROD, uroporphyrinogen III decarboxilase (EC:4.1.1.37); CPOX, coproporphyrinogen III oxidase (EC:1.3.3.3); hemN, oxygen-independent coproporphyrinogen III oxidase (EC:1.3.99.22); PPOX, protoporphyrinogen oxidase (EC:1.3.3.4); FeCH, ferrochelatase (EC:4.99.1.1). Compounds: 1, L-glutamate; 2, L-glutamyl-tRNA; 3, glutamate-1-semialdehyde; 4, aminolevulinic acid; 5, porphobilinogen; 6, hydroxymethylbilane; 7, uroporphyrinogen III; 8, coproporphyrinogen III; 9, protoporphyrinogen IX; 10, protoporphyrin IX; H, heme.

Figure 2. Maximum-likelihood phylogeny of coproporphyrinogen III oxidase.

Inset shows details of the tree in the Kinetoplastida clade. Numbers on the branches represent bootstrap support from analysis of 100 pseudo-replicates (only values of 50 or more shown). Colors represent some of the major taxa.

Figure 3. Maximum-likelihood phylogeny of porphobilinogen deaminase from representative Betaproteobacteria, the endosymbiotic C. Kinetoplastibacterium sp., and Alpha- and Gammaproteobacteria as outgroups.

Numbers on the branches represent bootstrap support from analysis of 100 pseudo-replicates (only values of 50 or more shown).