42 Organization of Genetic Material 3. Organellar Genomes (Mitochondrion, Chloroplast and Apicoplast) The genomes the three eukaryotic organelles Size and Organization of Genomes including mitochondria, chloroplasts and apicoplast have been studied in great detail ever The organeller genomes are relatively simple in since their discovery in 1960s and thereafter. organization and small in size (Table 42.1). Size Starting in 1995, now in 2017, whole genomw of mitochondrial genome differs greatly among sequences have become available for different organisms, ranging from 6 kb in mitochondrial genomes of ~8000 animal/plant Plasmodium (malarial parasite) to 2500 kb in species, chloroplast genomes of >800 plant melon (Cucumis melo). The variation in size of species and apicoplast genomes in >20 parasitic mitochondrial genomes is, however, unrelated to apicomplexan species (as in 2017). In this chapter, the variation in complexity of organisms we discuss the organization of mitochondrial, (Fig. 42.1). For instance, most multicellular chloroplast and apicoplast genomes. animals have small mitochondrial genomes with a compact genetic organization, the genes being close together with little space between them. This SIZE, SHAPE AND NUMBER is exemplified by human mitochondrial genome (Fig. 42.2), which is only 16,569 bp (~16.6 kb) (PER CELL) OF ORGANELLAR in length. In contrast, lower eukaryotes such as GENOMES S. cerevisiae (Fig. 42.3), as well as flowering plants, have larger and less compact mitochondrial genomes, with a number of genes, which contain Organellar genomes in mitochondria and introns. In humans, the two strands of double chloroplasts vary in their size, shape and number stranded mtDNA are also distinguished as heavy per cell. Same level of variation has not been strand (H-strand) and light strand (L-strand), reported yet for apicoplasts, perhaps because large since they differ in density, when measured on number of apicoplast genomes have not been CsCl density gradient. examined so far. 44. (BC–27) Mitochondria, Chloroplast and Apicoplast Genomes 705 Table 42.1. Sizes of mitochondrial and chloroplast genomes. Species Genome size (kb) Mitochondrial genomes Plasmodium falciparum 6 Chlamydomonas reinhardtii 16 Mus musculus 16 Homo sapiens 17 Metridium senile 17 Drosophila melanogaster 19 Chondrus crispus 26 Aspergillus nidulans 33 Reclinomonas americana 69 Saccharomyces cerevisiae 75 Suillus grisellus 121 Brassica oleracea 160 Arabidopsis thaliana 367 Zea mays 570 Cucumis melo 2500 Chloroplast genomes Pisum sativum 120 Fig. 42.1. Mitochondrial genomes, showing variation in Marchantia polymorpha 121 size. For comparison, genome of mitochondria-like Oryza sativa 136 proteobacteria of the Rickettsia group (Reclinomonas Nicotiana tabacum 156 americana) is also shown. Chlamydomonas reinhardtii 204 Apicoplast genomes Leucocytozoon caulleryi 34.8 apicoplast genome within a species and within the Plasmodium falciparum 34.7 same cell has not been reported, but may become P. chabuadi chabaudi 29.6 known, when many more genomes are sequenced. Toxoplasma gondii 35.0 Eimeria tenella 34.8 Theileria parva 39.6 Shape of Genome (DNA Molecules) Babesia bovis 33.4 Babesia microti 28.7 Cyclospora cayetanensis 33.2 The mtDNA and cpDNA were initially believed to be circular (like bacterial chromosomes) and In comparison of mitochondrial genomes, any linear DNA molecules observed, were actually chloroplast and apicoplast genomes are less attributed to broken circles (‘broken circle variable in size. The chloroplast genome ranges theory’). Both in bacteria and the cell organelles, from 120 kb in pea to 204 kb in Chlamydomonas the proposal of circular shape was actually also and the genomes of apicoplast range from ~27 kb based on the circular nature of their genetic maps, in Babesia microti to ~39.6 kb in Theileria parva without realizing that circular genetic maps could (Table 42.1). Most chloroplast genomes have a also result from linear head-to-tail concatamers. structure similar to that shown in Figure 42.4 for For almost 30 years (1970-2000), the ‘broken the rice chloroplast genome. Similarly, most circle theory’ remained a strong barrier to the apicoplast genomes have a structure shown in discovery of linear mtDNA/cpDNA. The Figure 42.5 for Plasmodium falciparum. However, apicoplast DNA is also circular in nature with the size of different DNA molecules within a some exceptions. chloroplast may vary, so that in many eukaryotes, During 1990s and in the early years of the larger molecules coexist with smaller molecules present century (2000-2004), it was demonstrated that contain subcomponents of the larger that a large fraction of in vivo mtDNA and multigenome molecules. The variation in size of cpDNA in many (and perhaps all) eukaryotes are 45. (BC–27) 706 Mitochondria, Chloroplast and Apicoplast Genomes Fig. 42.2. The human mitochondrial genome showing (i) 2 rRNA genes, (ii) 13 protein coding regions and (iii) regions containing genes for 22 tRNAs (14 transcribed in the clockwise direction and 8 transcribed in the anticlockwise direction). Fig. 42.3. The Saccharomyces cervisiae (budding yeast) mitochondrial genome. In the yeast genome, the genes are more spaced out than in the human mitochondrial genome and some of the genes have introns. This type of organization is typical of many lower eukaryotes and plants. (Abbreviations : ATP6, ATP8, ATP9, genes for ATPase subunits, 6, 8 and 9, respectively; COI, COII, COIII, genes for cytochrome C oxidase subunits I, II and III respectively; Cyt b, gene for apocytochrome b; var I, gene for a ribosome-associated protein. Ribosomal RNA and transfer RNA are two types of non-coding RNA. The 9S RNA gene specifies the RNA component of the enzyme ribonuclease P. 45. (BC–27) Mitochondria, Chloroplast and Apicoplast Genomes 707 Fig. 42.4. The rice chloroplast genome. Only genes with known functions are shown. A number of genes contain introns which are not indicated on the map. These discontinuous genes include several of those for tRNAs; this is why the tRNA genes are of different lengths although the tRNAs that they specify are all of similar size. essentially linear in shape, with only a small protozoa (e.g., Paramecium), and some fraction occurring as circular DNA. Linear apicomplexans (e.g., Plasmodium). Similarly, mtDNA has been reported in some algae linear cpDNA was reported in maize. For some (e.g., Chlamydomonas), several fungi (including apicoplast genomes also, linear DNA has been yeasts), plants (e.g., Chenopodium), some reported.. 708 Mitochondria, Chloroplast and Apicoplast Genomes Table 42.2. Relative amounts of organellar DNA in some cells and tissues. Organelle Tissue or DNA molecules Organelle Organellar DNA and organism cell type (genomes) per organelle per cell (as % total cell DNA) I Mitochondrial DNA Rat Liver 5-10 1000 1 Yeast Vegetative 2-50 1-50 15 Frog Egg 5-10 107 99 II Chloroplast DNA Chlamydomonas Vegetative 80 1 7 Maize Leaves 20-40 20-40 15 Number of Genomes per Organelle For instance, each human cell contains 800 mitochondria, and each mitochondrion contains as The number of genomes in an individual organelle many as 10 identical genomes, so that each cell also varies. A large number of mitochondria or contains about 8000 mitochondrial genomes. In chloroplasts are also available within an individual yeast, although the number of genomes per cell, thus increasing many-fold the number of mitochondrion is about 100, but there are fewer organellar genomes within a cell (Table 42.2). mitochondria per cell, thus making the number Fig. 42.5. Apicoplast genome from Plasmodium falciparum; genes are indicated as arrows indicating the direction of transcription; broken lines indicate functional regions; ribosomal RNA genes are labeled as LSU (large subunit) and SSU (small subunit); tRNA are shown by single letters indicated their corresponding amino acids. Mitochondria, Chloroplast and Apicoplast Genomes 709 of genomes per cell closer to that in humans (~5000). In case of plants, the number of GENES IN ORGANELLAR chloroplast genomes per cell is approximately GENOMES 5000, but that in photosynthetic microorganisms like Chlamydomonas, it is only 1000 per cell. Apicoplasts appear to contain, each a single Organellar genomes are much smaller than their genome, although future work may show presence nuclear counterparts. Therefore, it is anticipated of more than one genome per apicoplast, and that the gene contents in these organellar genomes more than one apicoplasts per cell. are much more limited, which is indeed the case. Mitochondrial genomes exhibit greater variability in gene content. Mitochondrial REPLICATION OF genomes display greater variability, the number of genes ranging from 5 for the malaria parasite ORGANELLAR DNA (P. falciparum) to 92 for the protozoan, Reclinomonas americana (Table 42.3). All mitochondrial genomes contain genes for the non- Several mechanisms for the replication of coding rRNAs and for at least some of the protein organellar DNA have been proposed in the past. components of the respiratory chain, the latter The most popular of these models is the model, being the main biochemical feature of the involving initial formation of D-loops mitochondrion. Some gene-rich mitochondrial followed by the formation of intermediate genomes also code for tRNAs, ribosomal proteins θ shaped structures and finally rolling circles and proteins involved in transcription and (see Chapter 39). This model was later translation; they also code for proteins questioned, and recombination-dependent